Mechanisms of

Cocaine Abuse

and Toxicity

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES • Public Health Service • Alcohol, Drug Abuse, and Mental Health Administration I
Mechanisms of Cocaine Abuse

and Toxicity

Editors:

Doris Clouet, Ph.D.

Khursheed Asghar, Ph.D.

Roger Brown, Ph.D.

Division of Preclinical Research

National Institute on Drug Abuse

NIDA Research Monograph 88

1988

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES

Public Health Service

Alcohol, Drug Abuse, and Mental Health Administration

National Institute on Drug Abuse

5600 Fishers Lane

Rockville, MD 20857

For sale by the Superintendent of Documents, U.S. Government Printing Office

Washington, DC 20402
NIDA Research Monographs are prepared by the research divisions of the

National Institute on Drug Abuse and published by its Office of Science. The

primary objective of the series is to provide critical reviews of research problem

areas and techniques, the content of state-of-the-art conferences, and

integrative research reviews. Its dual publication emphasis is rapid and targeted

dissemination to the scientific and professional community.

Editorial Advisors

MARTIN W. ADLER, Ph.D.

Temple University School of Medicine

Philadelphia,Pennsylvania

SYDNEY ARCHER, Ph.D.

Rensselaer Polytechnic lnstitute

Troy, New York

RICHARD E. BELLEVILLE, Ph.D.

NB Associates, Health Sciences

RockviIle, Maryland

KARST J. BESTEMAN

Alcohol and Drug Problems Association

of North America

Washington, D.C.

GILBERT J. BOTVIN, Ph.D.

Cornell Unvirsity Medical College

New York, New York

JOSEPH V. BRADY, Ph.D.

The Johns Hopkins University School of

Medicine

Baltimore, Maryland

THEODORE J. CICERO, Ph.D

Washington University School of

Medicine

St Louis, Missouri

MARY L. JACOBSON

National Federation of Parents for

Drug-Free Youth

Omaha, Nebraska

REESE T. JONES, M.D.

Langley Porter Neuropsychiatric lnstitute

San Francisco, California

DENISE KANDEL, Ph.D.

College of Physicians and Surgeons of

Columbia University

New York, New York

HERBERT KLEBER, M.D.

Yale University School of Medicine

New Haven, Connecticut

RICHARD RUSSO

New Jersey State Department of Health

Trenton, New Jersey

NIDA Research Monograph Series

CHARLES R. SCHUSTER, Ph.D.

Director, NIDA

THEODORE M. PINKERT, M.D.. J.D.

Acting Associate Director for Science, NIDA

Parklawn Buildlng, 5600 Fishers Lane, RockviIle, Maryland 20857
Mechanisms of Cocaine Abuse

and Toxicity
ACKNOWLEDGMENT

This monograph is based upon papers and discussion from a technical

review on mechanisms of cocaine abuse and toxicity that took place

on September 21-23, 1987, at Rockville, MD. The review meeting was

sponsored by the Office of Science and the Division of Preclinical

Research, National Institute on Drug Abuse.

COPYRIGHT STATUS

The National Institute on Drug Abuse has obtained permission from

the copyright holders to reproduce certain previously published

material as noted in the text. Further reproduction of this copyrighted material is permitted only as part of a reprinting of the

entire publication or chapter. For any other use, the copyright

holder’s permission is required. All other material in this volume

except quoted passages from copyrighted sources is in the public

domain and may be used or reproduced without permission from the

Institute or the authors. Citation of the source is appreciated.

Opinions expressed in this volume are those of the authors and do

not necessarily reflect the opinions or official policy of the National

Institute on Drug Abuse or any other part of the U.S. Department of

Health and Human Services.

The U.S. Government does not endorse or favor any specific commercial product or company. Trade, proprietary, or company names

appearing in this publication are used only because they are considered essential in the context of the studies reported herein.

DHHS publication number (ADM)89-1585

Printed 1988, Reprinted 1989

NIDA Research Monographs are indexed in the Index Medicus. They

are selectively included in the coverage of American Statistics Index,

BioSciences Information Service, Chemical Abstracts, Current Contents, Psychological Abstracts, and Psychopharmacology Abstracts.

iv
Contents

Page

Foreword ix

Pharmacological Effects of Cocaine Relevant

to Its Abuse

Robert L. Balster 1

Cocaine Receptors on Dopamine Transporters Mediate

Cocaine-Reinforced Behavior

Michael J. Kuhar, Mary C. Ritz, and John Sharkey 14

Cocaine Receptors on Monoamine Transporters

and Sodium Channels

Maarten EA. Reith 23

Modulation of Cocaine Receptors

lngeborg Hanbauer 44

Sensitization to Cocaine in the Nigrostriatal

Dopamine System

Nancy R. Zahniser, Joanna Peris, Linda P. Dwoskin,

Pamela Curella, Robert P. Yasuda, Laurie O’Keefe,

and Sally J. Boyson 55

The Interaction of Cocaine With Central Serotonergic

Neuronal Systems: Cellular Electrophysiologic

Approaches

Joan M. Lakoski and Kathryn A. Cunningham 78

v
Page

The Effects of Cocaine on Local Cerebral

Metabolic Activity

Linda J. Porrino and Conan Kornetsky 92

Behavioral Studies of the Reinforcing Properties

of Cocaine

Chris E. Johanson 107

Pharmacological Modifications of Cocaine and

Opioid Self-Administration

Gail Winger 125

Reinforcement Pathways for Cocaine

George F. Koob and Carol B. Hubner 137

Multiple Dopamine Receptors and the Behavioral

Effects of Cocaine

W.L. Woolverton and M.S. Kleven 160

Neurobehavioral Pharmacology of Cocaine

Steven I. Dworkin and James E. Smith 185

Intracranial Cocaine Self-Administration

Nick E. Goeders 199

Psychomotor Stimulant vs. Local Anesthetic Effects

of Cocaine: Role of Behavioral Sensitization

and Kindling

Robert M. Post and Susan R.B. Weiss 217

Genetic Differences in Responses

to Cocaine

Frank R. George and Steven R. Goldberg 239

Hepatotoxicity of Cocaine

Louis Shuster, Christine A. Garhart, James Powers,

Yona Grunfeld, and Gary Kanel 250

Lack of Toxic Effects of Cocaine on Dopamine

or Serotonin Neurons in the Rat Brain

Lewis S. Seiden and Mark S. Kleven 276

vi
Page

Developmental Effects of Cocaine

Diana L. Dow-Edwards 290

Cardiovascular Toxicity of Cocaine

R. Douglas Wilkerson 304

Arteriosclerotic Toxicity of Cocaine

Ronald O. Langner, Collette L. Bement,

and Lynda E. Perry 325

Mechanisms of Cocaine Abuse and Toxicity:

An Overview

Thomas K. Dunwiddie 337

List of NIDA Research Monographs 354

vii

Preface

The seriousness of cocaine use is illustrated by population statistics

on this drug. The most recent National Household Survey on Drug

Abuse1 shows that, in 1985, an estimated 22 million persons in the

continental United States had at least tried cocaine. Of these,

approximately 12 million individuals used it within the past year and

6 million at least once in the month prior to the survey (prior month

use is considered an indication of current use). “Freebasing” has

become a common occurrence and is more dangerous than snorting

because of the increased pharmacological effects of the drug and the

resulting drug craving. The propensity toward freebasing increases

with the frequency of use. Seven percent of the population that

reported using cocaine only one or two times reported smoking it,

whereas 57 percent of those who reported using it 100 times or more

had freebased. The popularity of the drug is underscored by figures

on teenage use.2 Over half (57 percent) of the graduating high

school class of 1987 had tried illicit drugs at least once; 15.2 percent

used cocaine, and crack use is estimated as 1 out of every 18 seniors

(5.6 percent).2

Although progress is being made through biobehavioral research, a

full understanding of the compulsion to keep using cocaine is, as yet,

not available. Cocaine produces a number of physiological effects,

including psychomotor stimulation, hypertension, tachycardia, anorexia,

pupillary dilation, and euphoria. These effects have been related to

the two major actions of cocaine: local anesthesia and actions in the

central nervous system. Such general pharmacological knowledge,

although necessary for further studies, does not provide an understanding of the basic mechanisms underlying cocaine reinforcement

and its abuse.

A technical review meeting on “Mechanisms of Cocaine Abuse and

Toxicity,” held September 21 to 23,1987, in Rockville, MD, addressed

ix
the main problems encountered in treating cocaine abusers, which are

twofold: to reverse or treat toxic effects of drug overdose and to

develop methods of dealing with the drug craving that leads to

cocaine abuse. Both of these problems have been examined in their

basic biomedical aspects in this review. In particular, the review

contains state-of-the-art knowledge of factors that influence drugseeking behavior in laboratory animals, including drug vulnerability,

basic neurochemical mechanisms, and neuronal sites of the “reward”

pathway, as well as toxicology, perinatal consequences, and cardiovascular actions. The proceedings of this conference are presented in

the following chapters, concluding with a review of the research

described in the conference and, especially, areas where future

efforts will be both feasible and productive.

The Editors

FOOTNOTES

1. National Household Survey on Drug Abuse: Population Estimates

7985. DHHS Pub. No. (ADM) 87-1539. Rockville, MD: National

Institute on Drug Abuse, 1987. 79 pp.

2. Johnston, LD.; O’Malley, P.M.; and Bachman, J.G. Illicit Drug

Use, Smoking, and Drinking by America’s High School Students,

College Students, and Young Adults, 1975-1987. Rockville, MD:

National Institute on Drug Abuse, in preparation.

x
Pharmacological Effects of Cocaine

Relevant to Its Abuse

Robert L. Balster

lNTRODUCTlON

The purpose of this paper is to provide an introduction to the

proceedings of a review sponsored by the National Institute on Drug

Abuse (NIDA) on the neurobiology of cocaine. This meeting was

called to survey some of what we know and to identify fruitful areas

for future inquiry. Armed with a better knowledge of the neurobiology of cocaine and a fuller understanding of the relationships of

the cellular actions to the pharmacological properties of cocaine that

are relevant to its abuse, we should be able to design better and

more targeted interventions for treatment and prevention.

In this context, then, one of the goals of this paper is to give an

overview of some of the pharmacological and behavioral effects of

cocaine for which we seek cellular mechanisms. The general

pharmacological profile of cocaine is fairly well known. it is a

short-acting central sympathomimetic stimulant with effects very

similar to those of amphetamines. However, unlike amphetamines,

cocaine also has local anesthetic effects. One would find little

disagreement with the assertion that it is the amphetaminelike

effects of cocaine that are relevant to its abuse; its local anesthetic

effects are responsible only for certain side effects commonly

encountered with insufflation or oral use and possibly for certain

aspects of cocaine toxicity. I would like to discuss these two aspects

of cocaine’s pharmacology and raise some questions that I hope will

be worthwhile to consider.

1
AMPHETAMINE LIKE EFFECTS OF COCAINE

Table 1 provides a list of some of the pharmacological and behavioral

effects shared by cocaine and the amphetamines. The first three

effects are particularly relevant for explaining the motivation to use

and abuse cocaine; the other effects may be more relevant to some of

the behavioral and physiological toxicity accompanying cocaine abuse.

TABLE 1. In vivo effects shared by cocaine and amphetamines

1. Subjective effects 5. Stereotyped behavior

in humans

6. Increased locomotor

2. Discriminative stimulus activity

effects

7. Rate-dependent increases

3. Reinforcing effects in operant behavior

4. Sympathomimetic 8. Cross-tolerance

physiological profile

9. Cross-sensitization

Subjective Effects

It is not as widely appreciated as it should be that the intoxication

produced by cocaine and d-amphetamine is very similar, even if the

duration of cocaine’s effects may be somewhat shorter. For example,

in the study by Fischman and colleagues (1976), 16 to 32 mg of IV

cocaine given to human research subjects produced a profile of

subjective effects very similar to that produced by 10 mg of damphetamine. In fact, in these experienced IV cocaine abusers, damphetamine was frequently identified incorrectly as cocaine.

Discriminative Stimulus Effects

Drug discrimination studies in animals are widely believed to be

measuring properties of drugs that are relevant to the production of

their subjective effects in humans (Schuster and Johanson 1988).

Animals can be readily trained to discriminate an active cocaine

injection from a vehicle injection. Similarly, animals can be trained

to discriminate amphetamine from vehicle injections. Under these

2
conditions, cocaine and amphetamines often substitute for one another

(Woods et al. 1987). For example, pigeons (Jarbe 1984), mice (Snoddy

and Tesset 1985), and rats (Colpaert et at. 1978; Woolverton and

Cervo 1986; Huang and Wilson 1986) trained to discriminate damphetamine from vehicle respond as if they had received d-amphetamine after cocaine injections. Similarly, d-amphetamine substites

for cocaine in pigeons (de la Garza and Johanson 1985). rats

(Colpaert et al. 1978; Huang and Wilson 1986; Wood and EmmettOglesby 1986), and rhesus monkeys (de la Garza and Johanson 1983)

trained to discriminate cocaine from vehicle. These data are

consistent with the similarities in the subjective effects of cocaine

and amphetamines reported in humans.

Reinforcing Effects

A number of the chapters in this volume will address the extensive

data on the reinforcing effects of cocaine and amphetamines.

Cocaine and d-amphetamine result in similar patterns of IV selfadministration in rhesus monkeys (Deneau et al. 1969; Johanson et al.

1978). Among those working in the area of drug self-administration,

it is widely believed that cocaine may be among the most effective

drug reinforcers there are. Some of the evidence for this is the

wide range of experimental conditions under which cocaine can be

shown to function as a reinforcer (Woods et al. 1987). A demonstration of cocaine’s powerful reinforcing effects was provided by a

study in my laboratory by Thomas Aigner (Aigner and Balster 1978).

In this study, rhesus monkeys were given a choice between IV

cocaine injections and a food pellet every 15 minutes, 24 hours a day,

for 8 days. All three animals tested chose cocaine on nearly every

trial and even relearned within a single day to perform the necessary

response to obtain cocaine after we reversed the response requirements for cocaine and food. They lost quite a lot of weight, so we

terminated the experiment after 8 days to prevent more serious

health effects.

lt is these powerful reinforcing effects of cocaine that are most in

need of being understood, since they are probably the most relevant

to the compulsive abuse of cocaine and its considerable attractiveness

to users. I will have more to say about what is known about the

neural substrates for these effects a little later, but first I will digress to a related subject that should be of some importance in the

present context.

3
"Crack"

One of the factors that has undoubtedly contributed to the current

crisis of coners about cocaine abuse la the emergence of a new

usage pattern, cocaine freebase in the form known as “crack.” Crack

is typically heated and the resulting vapor inhaled. This new form of

cocaine abuse has raised some important research questions as

summarized in table 2.

TABLE 2. Research questions raised by the abuse of "crack"

1.

2.

3.

4.

5.

What is the physical form of cocaine available for

absorption after heating?

Where in the tracheobronchial tree does absorption take

place?

Does oxidation occur and are there pyrolysis products?

What special toxicological problems result from the

inhalation route?

How do the effects of cocaine differ when it is inhaled?

Many factors have contributed to the popularity of crack, but paramount among these is the belief that the intoxication experienced

after inhaling crack is more desirable than that after insufftation. It

may be that higher doses can be self-administered by the inhalation

route. On the other hand, there may be a more profound rush by

the inhalation route. This rush may be as intense as that after IV

use. Laboratory studies comparing the timecourse for cocaine after

inhalation and IV administration confirm that the drug effects occur

very rapidly after both routes (Perez-Reyes et al. 1982).

Why should a rapid onset of drug effects produce a greater euphoric

response? There is evidence from animal research that the rate of

cocaine administration is a very important factor in its reinforcing

efficacy. In rhesus monkeys equipped with IV catheters and trained

to lever-press under a fixed-interval schedule for cocaine injections,

the rate of responding varied dramatically with changes in infusion

rate. Figure 1 shows this relationship (Balster and Schuster 1973).

4
FIGURE 1. Effects of infusion rate on the reinforcing effects of

cocaine in rhesus monkeys

NOTE: The duration of the infusion of two different doses of cocaine was varied from

5 to 200 seconds. The ordinate is the average change in response rate from

the rate with a 10-second infusion duration.

SOURCE: Modified from Balster and Schuster 1973, Copyright 1873. the Society for

the Experimental Analysis of Behavior, Inc.

The ordinate can be taken as a measure of the reinforcing strength

of each of the conditions tested. When the rate at which the same

dose of cocaine was given was decreased, i.e., when the infusion

duration was increased, response rates declined. In fact, infusion

rate was as important a determinant of response rate as dose. For

example, the response rate when responding was maintained by 0.4

mg/kg/injection delivered over 200 seconds was similar to that

maintained by 0.2 mg/kg/injection delivered in 100 seconds. Although

the dose differed by twofold in these tests, the rate of cocaine

delivery was the same, 0.002 mg/kg/second.

Why would the rate of cocaine infusion be such an important factor

in cocaine’s reinforcing effects? It could be hypothesized that, with

a more rapid rate of infusion, cocaine concentrations at critical sites

5
of action in the brain change more rapidly, and perhaps the rate of

change has important cellular consequences. At a biochemical level,

rate theory of drug-receptor interactions (Goldstein et al. 1974)

provides a model for considering this type of phenomenon. As a

more practical matter, biochemical and pharmacological studies

conducted under equilibrium or steady-state conditions cannot yield

information on the second-by-second changes that occur in the brain

when drugs like cocaine are taken intravenously or inhaled. Cellular

approaches may need to be expanded to include more dynamic

preparations capable of this temporal resolution.

BEHAVlORAL EFFECTS OF LOCAL ANESTHETICS

Cocaine, unlike the amphetamines, also has prominent local anesthetic

effects; however, the most wmmon assumption is that these local

anesthetic effects have little to do with the abuse of cocaine. On

the other hand, certain other local anesthetics have been shown to

have behavioral effects and are capable of producing an intoxication

in humans. Could the central nervous system (CNS) effects that

cocaine shares with local anesthetics play a role in cocaine abuse?

Could other local anesthetics have abuse potential? In the context of

this conference, it may be useful to briefly review some of the

behavioral pharmacology of local anesthetics.

Reinforcing Effects

Perhaps the most interesting behavioral action of certain local

anesthetics is their ability to function as reinforcers in IV drug selfadministration studies in monkeys. This was initially shown for procaine (Ford and Balster 1977; Hammerbeck and Mitchell 1978), but,

subsequently, a number of other esteratic local anesthetics were

found to have reinforcing effects as well (Johanson 1980; Woolverton

and Balster 1979; Woolverton and Balster 1982). lt is important to

note that not all local anesthetics are reinforcers. Some, such as

lidocaine and procainamide, are not.

Discimintaive Stimulus Effects

A number of drug discrimination studies have compared the discriminative stimulus effects of cocaine and local anesthetics. When animals

are trained to discriminate cocaine from vehicle, most drugs other

than sympathomimetic stimulants do not substitute for cocaine. On

the other hand, when procaine is tested, partial substitution often

results (de la Gana and Johanson 1983; de la Garza and Johanson

6
1985; Jarbe 1984; McKenna and Ho 1980), indicating some overlap in

the effects of these two drugs. Similarly, when animals are trained

to discriminate procaine from vehicle, cocaine almost completely

substitutes (Woolverton and Balster 1982). These data are shown in

figure 2.

Tests with cocaine resulted in dose-dependent generalization from

procaine; it was about tenfold more potent. Interestingly, damphetamine also substituted for procaine. lf one were to assume

FlGURE 2. Effects of substitutiing cocaine and &hetamine in rats

trained to discriminate procaine from saline

SOURCE: Modified from Woolverton and Balster 1982, Copyright 1982. Pergamon

Journals, Ltd.

that the reason cocaine substituted for procaine was because cocaine

produced local anesthetic effects similar to procaine, the results with

d-amphetamine would be difficult to explain unless one assumed that

d-amphetamine also had local anesthetic effects. Although this study

would need to be repeated before definite conclusions could be drawn,

the most parsimonious explanation at this point is that procaine,

cocaine, and d-amphetamine share some common effects not related to

local anesthesia.

7
Subjective Effects Procaine

Because of the behavioral effects of procaine seen in animals,

Fischman et al. (1983) compared it to cocaine after IV administration

in human subjects. Although procaine resufled in a different profile

of subjective effects and was considerably less efficacious than cocaine, a high dose of procaine did produce a “high,” and three of

the four subjects misidentified this dose of procaine as cocaine.

Thus, even in human subjects there is some evidence to support the

hypothesis that cocaine is not the only local anesthetic to have

behavioral effects that may be related to abuse.

NEURAL BASICS OF COCAINE’S EFFECTS

The subject of this monograph is the cellular basis of cocaine’s

actions. Any discussion of this topic must have clearly in mind the

effect for which a mechanism is sought. Cocaine has a complex

pharmacology and toxicology, and different mechanisms will undoubtedly be found for different effects. In this introduction, I

have focused on two aspects of the behavioral pharmacology of cocaine: its amphetaminelike effects and effects that are shared by

other local anesthetics. The amphetaminelike actions of cocaine are

generally well accepted and fairly well understood. On the other

hand, the basis for the commonalities with local anesthetics is less

clear.

The dopaminergic system undoubtedly plays an important role in most,

if not all, of the amphetaminelike effects of cocaine and has also

been widely implicated as a basis for its reinforcing effects (Woods

et al. 1987; Kometsky and Bain 1987). On the other hand, evidence

for an exclusive role for dopamine mediation of the discriminative

stimulus effects of cocaine is less clear, since selective dopamine

receptor blockers are not always completely effective antagonists of

the cocaine stimulus (Woods et al. 1987). It should be pointed out in

this context that the behavioral pharmacology of cocaine differs

considerably from selective dopamine agonists such as apomorphine

and amantadine, which leads to the suspicion that many behavioral

effects of cocaine, including its reinforcing and discriminative

stimulus effects, may result from cocaine’s interactions with a number

of neurochemical systems, not just dopamine systems. The relative

importance of these other biochemical effects of cocaine will be

addressed in this monograph.

8
Another question that will be addressed is the specific mechanism for

cocaine’s dopaminergic actions. Unlike the amphetamines, cocaine has

traditionally not been thought to release catecholamines but rather to

block their reuptake, particularly that of dopamine (Heikkila et al.

1975). Yet the pharmacology of cocaine is much more similar to that

of amphetamines than that of tricyclic antidepressants, perhaps due

to its higher affinity for the dopamine carrier than most antidepressants. Comparison of the behavioral pharmacology of cocaine and

antidepressants such as buproprion and nomifensine, which are also

potent dopamine uptake blockers, should be useful in determining the

role of dopamine uptake inhibition in cocaine’s neuropharmacology.

Nonetheless, until more is known about the pharmacological and

behavioral relevance of cocaine’s potent interactions with other

neurotransmitter systems, it would seem unwise to focus exclusively

on dopamine systems. This diversity of neurochemical actions of

cocaine is a prominent feature in this monograph.

Assuming for the moment that dopaminergic effects are an important

component of cocaine’s behavioral effects that are relevant to its

abuse, questions need to be asked about the possibility of specific

dopamine pathways that may be involved and dopamine receptor sub

types that may be important. This topic, too, will be an important

area of consideration in this conference.

Finally, I should comment on the problem of interpreting the data

showing similarities in the effects of cocaine and local anesthetics.

To put the matter most simply, either cocaine’s local anesthetic

effects are more important to its behavioral pharmacology than is

normally believed or certain local anesthetics have some effects,

possibly shared by cocaine, that account for their behavioral effects.

The former option would seem unlikely. Cocaine analogs have been

synthesized that allow a better separation of CNS effects and local

anesthetic effects (Clarke et al. 1973; Heikkila et al. 1979), and those

compounds with lower anesthetic potencies are nonetheless more

potent in self-administration studies (Spealman and Kelleher 1981) and

for altering schedule-controlled behavior (Spealman et al. 1977). In

addition, not all local anesthetics have similar behavioral effects

(Woolverton and Balster 1982), suggesting that some action of these

drugs other than their local anesthetic effect may be important.

Some resolution of this question was provided by the recent data of

Ritz et al. (1987), which will also be discussed in this monograph.

They found that certain local anesthetics bind to the same site as

cocaine, presumably on the dopamine transporter. In general, there

9
was a good correlation between those local anesthetics that have

reinforcing effects in animal self-administration studies and those

with a high affinity for this site. A similar correspondence is seen

between the effects on dopamine uptake and reinforcing effects for

the few cocaine analogs that have been studied in both procedures

(Spealman and Kelleher 1981; Reith et al. 1986; Ritz et al. 1987).

More extensive structure-activity studies of cocaine analogs and local

anesthetics for biochemical and behavioral effects will be an

important research direction for the future.

The fact that certain local anesthetics have affinity for the dopamine

transporter and potentially inhibit dopamine uptake (Ritz et al. 1987)

suggests that their behavioral effects may have a strong dopaminergic

component. This is consistent with the finding that some of these

drugs share discriminative stimulus effects with cocaine and amphetamine and that some serve as reinforcers. On the other hand, there

are differences in the behavioral effects of local anesthetics and

dopaminergic agonists. For example, there is evidence that haloperidol, which increases cocaine self-administration presumably due to

antagonism of the dopamlnergic effects of cocaine, fails to elicit

antagonist effects in procaine self-administration (de la Garza and

Johanson 1982). In our studies of procaine self-administration, we

did not see overt signs of dopaminergic effects such as stereotyped

behavior after the animals had received high doses (Ford and Balster

1977). What is clear from the studies of local anesthetics is that

further insight into their behavioral and biochemical effects may be

an important tool for understanding the neurobiology of cocaine.

CONCLUSION

In these introductory remarks, I have highlighted some of the behavioral effects of cocaine whose neural basis is the subject of this

volume. Cocaine has many amphetaminelike effects but also has

actions shared by certain local anesthetics. Comparisons among these

diverse drugs should prove to be important in discovering the cellular

basis for the actions of cocaine that are most relevant to its abuse.

Although dopaminergic systems appear to play an important role,

much needs to be learned about the exact nature of cocaine’s interactions with this important neurotransmitter. In addition, certain

aspects of the behavioral pharmacology of cocaine suggest that

dopamine may not be solely responsible for cocaine’s actions. Much

more needs to be learned about the functional significance of cocaine’s involvement with noradrenergic and serotonergic systems, and

perhaps others. In addition, the cellular mechanisms for cocaine’s

10
toxic effects may be quite different from those effects that result in

its abuse. These and other related topics are the subjects of the

papers that follow.

REFERENCES

Aigner, T.G., and Balster, R.L Choice behavior in rhesus monkeys:

Cocaine versus food. Science 201:534-535, 1978.

Balster, R.L, and Schuster, C.R. Fixed-interval schedule of cocaine

reinforcement: Effect of dose and infusion duration. J Exp Anal

Behav 20:119-129, 1973.

Clarke, R.L; Daum, S.J.; Gambino, A.J.; Aceto, M.D.; Pearl, J.;

Levitt, M.; Cuminsky, W.R.; and Bogado, E.F. Compounds affecting

the central nervous system. 4. 3 beta-phenyltropane-2-carboxylic

esters and analogs. J Med Chem 16:1260-1267, 1973.

Colpaert, F.C.; Niemegeers, C.J.; and Janssen, P.A. Discriminative

stimulus properties of cocaine and d-amphetamine, and antagonism

by haloperidol: A comparative study. /Veuruopharmacology 17:937-

942, 1978.

de la Gana, R., and Johanson, C.E. Effects of haloperidol and

physostigmine on self-administration of local anesthetics.

Pharmacol Biochem Behav 17:1295-1299, 1982.

de la Garza, R., and Johanson, C.E. The discriminative stimulus

properties of cocaine in the rhesus monkey. Pharmacol Biochem

Behav 19:145-146, 1983.

de la Garza, R., and Johanson, C.E. Discriminative stimulus

properties of cocaine in pigeons. Psychophannacology (Berlin)

85:23-30, 1985.

Deneau, G.; Yanagita, T.; and Seevers, M.H. Self-administration of

psychoactive substances by the monkey: A measure of psychological dependence. Psychopharmacologia 16:30-48, 1969.

Fischman, M.W.; Schuster, C.R.; and Rajfer, S. A comparison of the

subjective and cardiovascular effects of cocaine and procaine in

humans. Pharmacol Biochem Behav 18:711-716, 1983.

Fischman, M.W.; Schuster, C.R.; Resnekov, L.; Schick, J.F.E.;

Krasnegor, N.A.; Fennell, W.; and Freedman, D.X. Cardiovascular

and subjective effects of intravenous cocaine administration in

humans. Arch Gen Psychiatry 33:983-989, 1976.

Ford, R.D., and Balster, R.L Reinforcing properties in intravenous

procaine in rhesus monkeys. Pharmacol Biochem Behav 6:284-296,

1977.

Goldstein, A.; Aronow, L.; and Kalman, S.M. Principles of Drug

Action. 2nd ed. New York: John Wiley & Sons, 1974. pp. 104-106.

11
Hammerbeck, D.M., and Mitchell, C.L The reinforcing properties of

procaine and d-amphetamine compared in rhesus monkeys.

J Phamracol Exp Ther 204:558-569, 1978.

Heikkiia, R.E.; Cabbat, F.S.; and Duvoisin, R.C. Motor activity and

rotation behavior after analogs of cocaine: Correlation with

dopamine uptake blockade. Commun Psychopharmacol 3:285-290,

1979.

Heikkila, R.E.; Ortansky, H.; and Cohen, G. Studies on the distinction

between uptake inhibition and release of 3H-dopamine in rat brain

tissue slices. Biochem Pharmacol 24:847-852, 1975.

Huang, D., and Wilson, M.C. Comparative discriminative stimulus

properties of dl-chathinone, d-amphetamine, and cocaine in rats.

Pharmecol Biochem Behav 24:205-210, 1986.

Jarbe, T.U.C. Discriminative stimulus properties of cocaine. Effects

of apomorphine, haloperidol, procaine and other drugs.

Neuropharmocology 23:899-907, 1984.

Johanson, C.E. The reinforcing properties of procaine, chloroprowine, and proparacaine in rhesus monkeys. Psychopharmocology

(Berlin) 67:189-194, 1980.

Johanson, C.E.; Balster, R.L; and Bonese, K. Self-administration of

psychomotor stimulant drugs: The effects of unlimited access.

Pharmacol Biochem Behav 4:45-51, 1976.

Kometsky, C., and Bain, G. Neuronal bases for hedonic effects of

cocaine and opiates. in: Fisher, S.; Raskin, A.; and Uhlenhuth,

E.H., eds. Cocaine: Clinical and Biobehavioral Aspects. New York:

Oxford University Press, 1987. pp. 66-108.

McKenna, M.L, and Ho, B.T. The role of dopamine in the discriminative stimulus properties of cocaine. Neuropharmacology 19:298-

303, 1980.

Perez-Reyes, M.; DiGuiseppi, S.; Ondrusek, G.; Jeffcoat, A.R.; and

Cook, C.E. Freebase cocaine smoking. Clin Pharmacol Ther

32:459-485, 1982.

Reith, M.AE.; Meisier, B.E.; Sershen, H.; and Lajtha, A. Structural

requirements for cocaine congeners to interact with dopamine and

serotonin uptake sites in mouse brain and to induce stereotyped

behavior. Biochem Pharmacol 35:1123-1129, 1986.

Ritz, MC.; Lamb, R.J.; Goldberg, S.R.; and Kuhar, M.J. Cocaine

receptors on dopamine transporters are related to selfadministration of cocaine. Science 237:1219-1223, 1987.

Schuster, C.R., and Johanson, C.E. Relationship between the

discriminative stimulus properties and subjective effects of drugs.

In: Colpaert, F.C., and Balster, R.L, eds. Transduction

Mechanisms of Drug Stimuli. Berlin: Springer-Verlag, 1988.

pp. 161-175.

12
Snoddy, A.M., and Tessel, R.E. Prazocin: Effect on psychomotorstimulant cues and locomotor activity in mice. Eur J Pharmacol

116:221-228, 1985.

Spealman, R.D.; Goldberg, S.R.; Kelleher, R.T.; Goldberg, D.M.; and

Charlton, J.P. Some effects of cocaine and two cocaine analogs on

schedule-controlled behavior of squirrel monkeys. J Pharmecol Exp

Ther 202:500-509, 1977.

Spealman, R.D., and Kelleher, R.T. Self-administration of cocaine

derivatives by squirrel monkeys. J Pharmacol Exp Ther 216:532-536,

1981.

Wood, D.M., and Emmett-Oglesby, M.S. Characteristics of tolerance,

recovery from tolerance and cross-tolerance for cocaine used as a

discriminative stimulus. J Pharmacol Exp Ther 237:120-125, 1986.

Woods, J.H.; Winger, G.D.; and France, C.P. Reinforcing and

discriminative stimulus effects of cocaine: Analysis of pharmacological mechanisms. In: Fisher, S.; Raskin, A; and Uhlenhuth,

E-H., eds. Cocaine: Clinical and Biobehevioral Aspects. New York:

Oxford University Press, 1987. pp. 21-65.

Wooiverton, W.L. and Balster, R.L Reinforcing properties of some

local anesthetics in rhesus monkeys. Pharmacol Biochem Behav

11:669-672, 1979.

Woolverton, W.L, and Balster, R.L Behavioral pharmacology of local

anesthetics: Reinforcing and discriminative stimulus effects.

Pharmacol Biochem Behav 16:491-500, 1982.

Woolverton, W.L, and Cervo, L. Effects of central dopamine

depletion on the d-amphetamine discriminative stimulus in rats.

Psychopharmacology (Berlin) 88:196-200, 1986.

ACKNOWLEDGMENTS

Preparation of this paper and much of the research from my

laboratory discussed herein were supported by U.S. Public Health

Service grant DA 00490 from the National Institute on Drug Abuse.

AUTHOR

Robert L. Balster, Ph.D.

Professor

Department of Pharmacology

and Toxicology

Medical College of Virginia

Virginia Commonwealth University

Richmond, VA 23298-0613

13
Cocaine Receptors on Dopamine

Transporters Mediate

Cocaine-Reinforced Behavior

Michael J. Kuhar, Mary C. Ritz, and John Sharkey

INTRODUCTlON

Cocaine is a powerful reinforcing drug that has a variety of

pharmacological effects on the central nervous system. While several

binding sites for 3H-cocaine have been identified in the brain and in

the periphery (Reith et al. 1980; Kennedy and Hanbauer 1983;

Schoemaker et al. 1985; Calligaro and Eldefrawi 1987), these have not

been associated with the addictive or reinforcing properties of

cocaine. Recently, such a receptor for cocaine has been identified in

the brain (Ritz et al. 1987a). This receptor appears to be the

cocaine-binding site on the dopamine transporter on dopaminergic

nerve terminals.

GOALS AND METHODS

The goal of the study was to determine the relative importance of

various receptors in mediating cocaine reinforcement. To do so

required a correlation between the affinities of cocaine and related

drugs at the various receptor sites and the pharmacological profile of

these drugs in a drug self-administration paradigm. The study

included drugs that were closely related to cocaine, such as cocaine

analogs, other ester-linked local anesthetics, and some other closely

related psychostimulants such as methylphenidate (table 1). Other

drugs, such as amphetamines, that produce similar behavioral effects

were excluded because of evidence that they had different pharmacokinetic properties or functioned through somewhat different

mechanisms (McMillen 1983).

lt is well established that cocaine inhibits monoamine uptake (Iversen

1973; Ritz et al. 1987a), and these sites appear to be labeled by

14
TABLE 1. Potencies of cocaine and related compounds in selfadministration and biochemical studies

Dopamine Uptake

Relative Relative

Compound Behavioral Potency K1

(M) Potency

1. WIN 35, 065-2 0.26 0.26 0.41

2. WIN 35, 981 0.58 0.36 0.56

3. (-)Cocaine 1 0.64 1

4. Dimethocaine 0.67 1.29 2.02

5. (+/-)Norcocaine 2.82 1.21 1.89

6. Procaine 14.1 104 164

7. Chloroprocaine 2 9 65 102

8 . (+)Pseudococaine 3 3 116 183

9. Mazinol 0.17 0.023 0.036

10. Methlyphenidate 1 0.39 0.61

11. (+)Ampehtamine 0.20 3.6 5.63

12. Lidcaine n.d 3,298 5,153

13. Procainamide n.d 1,943 3,036

14. Diethylaminoethanol n.d 18,197 28,433

15. WIN 35, 065-3 n.d 385 602

16. (+)Cocaine n.d 136 213

17. WIN 35, 428 - 0.17 0.27

18. (+/-)Cocaine - 1.51 2.36

NOTE: The relative behavioral potencies In self-administration studies were determined by averaging values obtained from studies of cocaine reinforcement.

These could not be determined (n.d.) for compounds 12 to 18 due to their low

potencies or toxic side effects. Compound 17 was tested only in discrimination studies. Except in two cases, the animals utilized were monkeys. 3Hmazindol was used to label the dopamine transporter (105

 M nomifensine

blank) in striatum. Tissues were homogenized then incubated for 1 hour in

buffer (50 mM tris, 120 mM NaCl. 5 mM KCI; pH 7.8; 4 °C) containing a final

3H-mazindol concentration of 4 nM. Behavioral potendes were determined as

described previously (Ritz et al. 1987a).

3H-cocaine (Reith et al. 1980; Kennedy and Hanbauer 1983; Schoemaker et al. 1985; Calligaro and Eldefrawi 1987). Accordingly, we

decided to examine these 3H-cocaine-binding sites as well as a large

number of other drug-binding sites in brain to determine their

15
possible relationship to drug self-administration. Of the more than

25 blnding sites examined, cocaine had relatively high affinity only at

uptake sites for dopamine, serotonin, and norepinephrine (Ritz et al.

1987a); at sigma opiate receptors; and at muscarlnic cholinergic

receptors. The potency of cocaine at these sites was in the

micromolar range, which is similar to the blood levels in human

subjects when they experience the subjective feeling of “high”

(Fiihman et al. 1983; Javaid et al. 1978). Thus, these affinities were

considered appropriate for a physiologically relevant cocaine receptor.

Choline uptake sites and a large number of other binding sites had

relatively low affinities for cocaine (Ritz et al. 1987a).

RESULTS

In general, drugs that are potent in self-administration studies are

also potent inhibitors of binding at the transport sites for dopamine

(table 1). Compounds that are weak in self-administration studies are

correspondingly weak at the binding site. Moreover, drugs that have

no detectable potency in drug reinforcement studies, despite chemical

similarities (such as optical isomers), show extremely weak binding

affinities.

A statistical evaluation of the data indicated that inhibition of

3H-mazindol binding to the dopamine transporter was significantly

related to drug self-administration of cocaine and related compounds.

A multiple regression analysis relating the logarithms of the relative

concentrations to inhibit binding and the logarithms of the relative

behavioral doses for compounds 1 through 10 was significantly and

positively associated (slope=0.72±0.123, p<.001) (figure 1).

Additional studies using 3H-GBR 12935, a more selective dopamine

uptake blocker than mazindol (Van der Zee et al. 1980), gave results

that were consistent with these conclusions. We have shown that

3H-GBR 12935 was competitively displaced by mazindol and cocaine

(Sharkey and Kuhar 1987). The affinities of cocaine and related

compounds are similar at the 3H-mazindol and 3H-GBR 12935 sites

3

(r=.96, p<.001). Thus, the relative potencies of these drugs at the

H-GBR 12935 site are significantly correlated with their potencies in

drug self-administration studies (r=.92, p<.001). Taken together, these

findings strongly suggest that the dopamine transport mechanism is

the primary mechanism associated with the reinforcing effects of

cocaine.

16
RELATIVE CONCENTRATION TO INHIBIT BINDING

FIGURE 1. Relation between the relative behavior doses of cocaine

and related compounds in self-administration studies and

their relative inhibitory concentrations for 3H-mazindol

binding at the dopamine transporter

NOTE: Linear regressions of logarithms of relative behavioral doses on logarithms of

relative inhibitory concentrations for compounds 1 through 10 from table 1

were calculated using the BMDP statistical package. Value shown Is slope ±

SD. Compound 11 (open circle) was not induced in the regression analysis. A

two-tailed test of significance was applied (p<.0001).

SOURCE: Ritz et al. 1987a, Copyright 1987, American Association for the Advancement of Science.

Cocaine also has an affinity for sigma receptors in the low micromolar range (Sharkey et al., in preparation). Since sigma agonists are

also reinforcing in animal models (Slifer and Balster 1983), it could

be argued that cocaine’s reinforcing properties could be mediated via

sigma receptors. However, the relative potencies of the cocaine

analogs tested at the sigma site do not correlate with their potencies

in producing reinforcement in animals. For example, procaine and

17
lidocaine, which display only weak reinforcing properties in animal

models (Woolverton and Balster 1979). are approximately equipotent

with cocaine in their affinities at the sigma site (Sharkey et al., in

preparation).

In our studies of the binding properties of cocaine at various

receptors, we investigated the effects of cocaine at muscarinic

cholinergic receptors in both brain and heart tissues (Sharkey et al.,

in preparation). These binding studies revealed that the (+)

enantiomer of cocaine displayed an eightfold higher affinity than the

(-) enantiomer. This stereospecificity is opposite to that shown by

cocaine at dopamine uptake sites and in cocaine reinforcement

studies. Thus, cocaine binding at muscarinic receptors does not

appear to mediate its reinforcing effects. However, (-)cocaine has a

k1 of about 18 M at cholinergic muscarinic receptors in the heart.

Thus, these receptors would be blocked at high blood levels of

cocaine that appear to be in the toxic range. Consistent with this,

the average blood level of cocaine in autopsy studies where cocaine

has been implicated as a cause of death was about 20 μM Mittleman

and Wetli 1984). Our data indicate that cocaine binding to 3H-QNB

sites is not shifted in the presence of 10 μM Gpp(NH)p, suggesting

that cocaine does not act as an agonist at these sites. Thus, it is

possible that inhibition of muscarinic cholinergic receptors, which

would functionally enhance the sympathomimetic properties of cocaine

by blocking the parasympathetic cholinergic receptors, might be

involved in the cardiotoxicity of cocaine.

DISCUSSION

Many other reports are consistent with our findings relating cocaine

binding to the dopamine transporter with drug self-administration. In

animal studies, dopamine-containing neuronal systems, particularly in

the limbic region, have been strongly implicated in the self-administration of cocaine (Wise 1984; Goeders et al. 1986; Roberts et al.

1980; Roberts and Koob 1982). Intravenous substitution of dopaminergic agonists can maintain cocaine-reinforced behavior (Woolverton et

al. 1984). Self-administration of cocaine has been modulated by

dopamine receptor blockers but not by drugs that affect other

neurotransmitter systems. In clinical studies, it has been found that

compounds that potentiate or mimic dopaminergic transmission,

including methylphenidate and bromocriptine, decrease craving for

cocaine (Dackis et al. 1985-1986; Khantzian et al. 1984; Gawin and

Kleber 1984). By contrast, other neurotransmitter systems are

apparently not involved in the mediation of the positive reinforcing

18
effects of cocaine. Serotonin agonists, in contrast to dopamine

agonists, decrease cocain-reinforced behavior (Mattia et al. 1986).

Depletion of brain norepinephrine has little influence on selfadministration of cocaine (Roberts et al. 1977). Thus, although

cocaine effects norepinephrine transport and serotonin transport as

well as dopamine transport, the above studies indicate that the

reinforcing properties of the drug are associated with dopamine

uptake inhibition and not serotonin uptake or norepinephrine uptake

inhibiiion. While we cannot rule out the involvement of other,

unknown sites in producing the reinforcing effects of cocaine, it

should be emphasized that binding to the dopamine transporter

accounts for 94 percent of the variance in cocaine self-administration

(Ritz et al. 1987a).

At present, there is some question concerning the reinforcing prop

erties of mazindol. Mazindol has been shown to be self-administered

in animals (Wilson and Schuster 1976; Risner and Silcox 1981) and,

therefore, has positive reinforcing properties. However, mazindol may

have other properties, possibly even aversive properties, which could

become apparent under other conditions (Chait et al. 1987). Nevertheless, the presence of other properties does not preclude the

existence of quantifiable reinforcing properties of mazindol and its

inclusion in our study. Also, its behavioral properties do not detract

from its value as a biochemical marker for the dopamine transporter

in vitro (Javitch et al. 1984).

We have demonstrated that 3H-mazindol and 3H-GBR 12935 are inhibited by cocaine and related compounds (Ritz et al. 1987b; Sharkey

and Kuhar 1987). Thus, the sites recognized by 3H-mazindol and 3HGBR 12935 are presumed to be the same as or very close to the site

that recognizes cocaine. However, studies with 3H-mazindol and 3HGBR 12935 indicate that the site labeled by cocaine, mazindol, and

GBR 12935 may be distinct from the dopamine recognition site on the

dopamine transporter (Javitch et al. 1984; Anderson 1987).

CONCLUSION

lt has been shown that a cocaine-binding site on the dopamine

transporter in brain is significantly related to the positive reinforcing

properties of cocaine and some related compounds. Presumably, the

main physiological effect of binding to this site is inhibition of

dopamine uptake and potentiation of dopaminergic transmission. This

suggestion that the dopaminergic nerve terminal is the primary site of

action of cocaine is supported by the large body of evidence showing

19
that dopaminergic systems are affected by cocaine. In fact, intact

limbic dopaminergic systems are required for the reinforcing

properties of the drug. Thus, the cocaine receptor related to its

dependence-producing properties is proposed to be the cocainebinding site on the dopamine transporter.

REFERENCES

Andersen, P.H. Biochemical and pharmacological characterization of

[

3H]GBR 12935 binding in vitro to rat striatal membranes: Labeling

of the dopamine uptake complex. J Neurochem 46:1887-1896, 1987.

Calligaro, D.O., and Eldefrawi, M.E. Central and peripheral cocaine

receptors. J Pharmacol Exp Ther 243:61-68, 1987.

Chait, L.D.; Uhlenhuth, E.H.; and Johanson, C.E. Reinforcing and

subjective effects of several anorectics in normal volunteers.

J Pharmacol Exp Ther 242:777-783, 1987.

Dackis, C.A.; Gold, M.S.; Davies, R.K; and Sweeney, D.R.

Bromocriptine treatment for cocaine abuse: The dopamine depletion

hypothesis. Int J Psychiatry Med 15: 125-135, 1985-1986.

Fischman, M.; Schuster, C.R.; and Rajfer, S. A comparison of the

subjective and cardiovascular effects of cocaine and procaine in

humans. Pharmacol Biochem Behav 202:711-716, 1983.

Gawin, F.H., and Kleber, H.D. Cocaine abuse treatment: Open pilot

trial with desipramine and lithium carbonate. Arch Gen Psychiatry

41:903-909, 1984.

Goeders, N.E.; Dworkin, S.L; and Smith, J.E. Neuropharmacological

assessment of cocaine self-administration into the medial prefrontal

cortex. Pharmacol Biochem Behav 24:1429-1440, 1986.

Iversen, L.L. Catecholamines. Br Med Bull 29:91-178, 1973.

Javaid, J.I.; Fischman, M.W.; Schuster, C.R.; Dekirmenjian, H.; and

Davis, J.M. Cocaine plasma concentration: Relation to physiological and subjective effects in humans. Science 202:227-228,

1978.

Javitch, J.A.; Blaustein, R.O.; and Snyder, S.H. 3H-mazindol binding

associated with neuronal dopamine and norepinephrine uptake sites.

Mol Pharmacol 26:35-44, 1984.

Kennedy, L.T., and Hanbauer, I. Sodium-sensitive cocaine binding to

rat striatal membrane: Possible relationship to dopamine uptake

sites. J Neurochem 41:172-178, 1983.

Khantzian, E.J.; Gawin, F.H.; Kleber, H.D.; and Riordan, C.E.

Methylphenidate (Ritalin) treatment of cocaine dependence--A

preliminary report. J Subst Abuse Treat 1:107-112, 1984.

20
Mattia, A.; Leccesse, A.P.; and Morton, J.E. Effects of quizazine on

cocaine self-administration in rats. The Pharmacologist 28:151,

1986.

McMillen, B.A. CNS stimulants: Two distinct mechanisms of action

for amphetamine-like drugs. Trends Pharmacol Sci 4:429-432, 1983.

Mittleman, R.E., and Wetli, C.V. Death caused by recreational

cocaine use. JAMA 252:1889-1893, 1984.

Reith, M.E.; Sershen, H.; and Lajtha, A. Saturable 3H-cocaine

binding in central nervous system of mouse. Life Sci 27:1055-

1062, 1980.

Risner, M.E., and Silcox, D.L Psychostimulant self-administration by

beagle dogs in a progressive-ratio paradigm. Psychopharmacology

(Berlin) 75:25-30, 1981.

Ritz, M.C.; Lamb, R.J.; Goldberg, S.R.; and Kuhar, M.J. Cocaine

receptors on dopamine transporters are related to selfadministration of cocaine. Science 237:1219-1223, 1987a.

Ritz, M.C.; Sharkey, J.; and Kuhar, M.J. Cocaine inhibits QNB

binding to muscarinic receptors in rat brain and heart. Abstr Soc

Neurosci 13:42.5, 1987b.

Roberts, D.C.S.; Corcoran, M.E.; and Fibiger, H.C. On the role of

ascending catecholamine systems in intravenous self-administration

of cocaine. Pharmacol Biochem Behav 6:615-620, 1977.

Roberts, D.C.S., and Koob, G.F. Disruption of cocaine selfadministration following 6-hydroxydopamine lesions of the ventral

tegmental area in rats. Pharmacol Biochem Behav 17:901-904,

1982.

Roberts, D.C.S.; Koob, G.F.; Klonoff, P.; and Fibiger, H.C.

Extinction and recovery of cocaine self-administration following

6-hydroxydopamine lesions of the nucleus accumbens. Pharmacol

Biochem Behav 12:781-787, 1980.

Schoemaker, C.P.; Pimoule, C.; Arbilla, S.; Scatton, B.; Javoy-Agid,

F.; and Langer, S.Z. Sodium-dependent [

3H]cocaine binding

associated with dopamine uptake sites in the rat striatum and

human putamen decrease after dopaminergic denervation and in

Parkinson’s disease. Naunyn Schmiedebergs Arch Pharmacol

329:227-235, 1985.

Sharkey, J., and Kuhar, M.J. 3H-GBR 12935 labels the cocaine

binding site associated with dopamine uptake inhibition. The

Pharmacologist 29:294, 1987.

Sharkey, J.; Glen, K.A.; Wolfe, S.; and Kuhar, M.J. Cocaine

binding at sigma receptors. In preparation.

21
Slifer, B.L, and Balster, R.L Reinforcing properties of stereoisomers of the putative sigma agonists N-allylnormetazocine and

cyclazocine in rhesus monkeys. J Pharmacol Exp Ther 225:522-

528, 1983.

Van der Zee, P.; Koger, H.S.; Gootjes, J.; and Heope, W. Aryl 1,4-

dialk (en)yipiperazines as selective and very potent inhibitors of

dopamine uptake. Eur J Med Chem 15(4):363-370, 1980.

Wilson, M.C., and Schuster, C.R. Mazindol self-administration in the

rhesus monkey. Pharmacol Biochem Behav 4:207-210, 1976.

Wise, R.W. Neural mechanisms of the reinforcing action of cocaine.

in: Grabowski, J., ed. Cocaine: Pharmacology, Effects, and

Treatment of Abuse. National Institute on Drug Abuse Research

Monograph 50. DHHS Pub. No. (ADM)84-1326. Washington, DC:

Supt. of Docs., U.S. Govt. Print. Off., 1984. pp. 15-33.

Woolverton, W.L, and Balster, R.L Reinforcing properties of some

local anesthetics in rhesus monkeys. Pharmacol Biochem Behav

11:669-672, 1979.

Woolverton, W.L; Goldberg, L.; and Ginos, J.Z. Intravenous selfadministration of dopamine receptor agonists by rhesus monkeys.

J Pharmacol Exp Ther 230:678, 1984.

ACKNOWLEDGMENTS

Steven R. Goldberg and R.J. Lamb, Preclinical Branch, NIDA Addiction

Research Center, and A. Kimball, Johns Hopkins University, are

acknowledged for helpful discussions and collaborative efforts.

AUTHORS

Michael J. Kuhar, Ph.D., Chief, Neuroscience Research Branch

Mary C. Ritz, Ph.D., Staff Fellow

John Sharkey, Ph.D., Visiting Fellow

Addiction Research Center

National Institute on Drug Abuse

4940 Eastern Avenue

P.O. Box 5180

Baltimore, MD 21224

22
Cocaine Receptors on Monoamine

Transporters and Sodium

Channels

Maarten E.A. Reith

lNTRODUCTlON

Cocaine receptors can be divided into two categories.

(1) Sites that bind cocaine with high affinity and that can be

detected with [

3H]cocaine by standard receptor-binding

techniques. Two types have been characterized so far:

(a) NA+-sensitive receptors associated with dopaminergic nerve

terminals; and

(b) NA+-insensitive receptors associated with serotonergic

nerve terminals.

(2) Sites that bind cocaine with high affinity and that cannot be

detected with [

3H]cocaine by standard receptor-binding

techniques. The following sites are in this category:

(a) norepinephrine uptake sites;

(b) nonmonoaminergic sites; and

(c) sites on sodium channels.

This chapter reviews the work done in our laboratory on the sites

(1)(a), (1)(b), and (2)(c), and the implication of these sites in the

effect of cocaine and cocaine analogs on locomotor behavior and

stereotyped behavior in mice.

23
LOCALIZATION OF COCAINE RECEPTORS: TOPOGRAPHICAL

[

3H]Cocaine binds with high affinity to brain membranes under

conditions routinely used in most receptor-binding assays, such as the

use of crude particulate fractions and 50 mM Tris-HCI at pH 7.4 to

7.7 (Reith et al. 1980a; Reith et al. 1981). Kinetic experiments

indicated an extremely rapid rate of dissociation of [

3H]cocaine

binding from mouse cerebrocortical membranes that was not related

to the use of Tris-HCI buffer, which we later found to be inhibitory

for cocaine binding and to act like a competitive inhibitor of cocaine

binding (Reith et al. 1984a). The rate of dissociation of [

3H]cocaine

binding, however, was the same in sodium phosphate buffer as in

Tris-HCI buffer, with a half-life of approximately 25 seconds (Reith

et al. 1986a).

There were distinct differences between various mouse brain regions in

the effect of Na+ on the binding of [

3H]cocaine (figure 1). The

striatum showed pronounced Na+-stimulated binding: a maximal

fourfold increase in 25 mM Na+. There was relatively little stimulation by Na+

 in the olfactory tubercle, another dopaminergic area.

These results are similar to those reported by Kennedy and Hanbauer

(1983) for Sprague-Dawley rats. Concentrations of Na+ up to 50 mM

had little or no effect in cortex and midbrain+(hypo)thalamus, slightly

inhibited binding in hippocampus and pons-medulla, and decreased

binding by 60 percent in the cerebellum (figure 1). Higher concentrations of Na+

(100 to 200 mM) were inhibitory in all brain regions.

Thus, Na+ dependent binding is abundant in the striatum and present

to some extent in the olfactory tubercle, and Na+-independent binding

is present in all brain regions.

The studies by Kennedy and Hanbauer (1983) indicated an association

of NA+-dependent cocaine binding in the striatum with dopaminergic

nerve terminals. In contrast, the NA+-independent binding in the

cerebral cortex had the pharmacological profile of serotonin uptake

sites, and was reduced by lesions caused by neurotoxins aimed at

serotonergic nerve terminals (Reith et al. 1983a).

Cocaine binding observed in the striatum in the absence of NA+ represented the NA+

-independent site of the serotonergic type observed

in other brain areas as shown by the effect of 5,7-dihydroxytryptamine (Reith et al. 1985a).

24
FIGURE 1. Binding of [

3H]cocaine to mouse brain membranes as a

function of Na+ concentration

NOTE: embranes from mouse brain regions were incubated in triplicate with 16 nM

[

3H]cocaine and varying amounts of Na+

, in the presence and absence of 30

μM cocaine for defining nonspecific binding. For the range of 0 to 50 mM

Na+, membranes were buffered with 5 mM Tris-HCI, pH 7.7; for the range of 50

to 200 mM Na+, with 25 mM sodium phosphate buffer, pH 7.7, which contributes

48 mM Na+ from its buffer salts. The range of 16 to 66 mM Na+ was also

studied with 25 mM HEPES buffer, pH 7.7, with similar results. The SEM in

triplicate detetminations was less than 7 percent. OLF TUBER, olfactory

tubercle; CEREB, cerebdlum; MIDBRAIN, mid brain+(hypo)thafamus;

PONS-MED, pons-medulla; and HIPPO, hippocampus.

25
There was a significant correlation between the Na+

-independent

binding of [

3H]cocaine at a low concentration and the neuronal

uptake of [ 3H]serotonin in various brain regions (figure 2, top

left panel). Likewise, there was a good correlation between the

Bmax of NA+

-independent [

3H]cocaine binding and [3H]serotonin

uptake in the regions (figure 2, bottom left panel). There was no

relationship between NA+-independent [

3H]cocaine binding and uptake

of [

3H]dopamine into noradrenergic and dopaminergic nerve terminals

(figure 2, middle and right panels). There was no relationship

between cocaine binding and the uptake of norepinephrine, as

indicated by the lack of regional correlation (figure 2) and the

ineffectiveness of lesioning by 6-hydroxydopamine in reducing the

binding in the cerebral cortex (Reith et al. 1983a).

In general, the high-affinity binding of ligands to uptake sites

requires the presence of sodium if sodium is required by the

transport of the substrate that those sites subserve. However,

there are exceptions to this rule. For instance, [

3H]cocaine binds

with high affinity to serotonin-related sites that are Na+-

insensitive, and 8-hydroxy-2-(di-N-propylamino)tetralin binds to

serotonin uptake sites in the striatum in the absence of sodium

(Schoemaker and Langer 1986).

LOCALIZATION OF COCAINE RECEPTORS: SUBCELLULAR

Subcellular fractions from mouse cerebral cortex were separated to

localize cocaine receptors (Whittaker 1969). The heavy and light

synaptosomal mitochondrial fractions accounted for 50 percent of the

recovered cocaine binding without enrichment in specific binding

activity (Reith et al. 1984b). Most of the binding was recovered in

the synaptosomal subfraction, although a small amount of binding in

myelin had the highest relative specific activity. Since, in these

experiments, Tris-HCI was used, thus reducing high-affinity binding,

this localization requires cautious interpretation. However, the

results are consonant with the idea that a substantial portion of

cocaine binding observed in brain homogenates or crude membranes is

associated with nerve terminals (Reith et al. 1984b).

INTERACTION OF COCAINE WITH THE SEROTONIN

TRANSPORTER: STRUCTURE-ACTIVITY RELATIONSHIP

The following profile of cocaine congeners (table 1) was the same for

Na+-independent cocaine binding measured in synaptosomal membranes

from the cerebral cortex and for [ 3H]serotonin uptake into

26
FIGURE 2. Uptake of monoamines, and Na+

-independent binding of

[

3H]cocaine in various mouse brain regions

NOTE: The top panel shows Na+-independent binding of [

3H]cocaine in one-point

assays at 16 nM of labeled ligand; the bottom panel shows Bmax, estimates,

Values are the average ± range of tw8 independent preparations assayed in

triplicate. 5-HT, neuronal uptake of [

3H]serotonin; NE, uptake of [3H]dopamine into nomdrenergic nerve terminals (defined as total uptake minus

uptake in the presence of 1 μM desipramine); DA, uptake of [

3H]dopamine

into dopaminergic terminals (estimated from the difference between uptake

into both dopaminergic and noradrenergic terminals (as defined with 50 μM

benztropine) and uptake into noradrenergic terminais (as defined with 1 μM

desipramine)). 1, cerebellum (CER); 2, hippocampus (HIP); 3, striatum

(STRIA); 4, cerebral cortex (CORT); 5, olfactory tuberde (OLF); 6, ponsmedulla (POMED); and 7, midbrain+(hypo)thalamus (MID+(HY)THA).

synaptosomes from the cerebral cortex. Removal of the ester

linkage between the tropane and phenyl rings of cocaine (WIN 5,428

and WIN 35,065-2) did not reduce the affinity of the molecule for

the serotonin uptake sites. Moving the carbomethoxy group (R2) on

C2 from an axial (cocaine and WIN 35,065-2) to an equatorial

27
((+)-pseudococaine and WIN 35,140) position diminished the potency

from fortyfold to a hundredfold. Likewise, moving the O-benzoyl

group (R4) on C3 from an equatorial (benzoylpseudotropine) to an

axial (benzoyltropine) position reduced the activity of the

molecule threefold. Therefore, the position of the C2 constituent

seems to be more important than that of the C3 constituent (Reith

et al. 1986b). Removal of the C3 constituent (benzoylpseudotropine

vs. cocaine) reduced the activity tenfold, but had a less profound

effect than removal of the methylester from the C3 position of

cocaine (benzoylecgonine), which rendered the molecule inactive.

An isomeric pair, WIN 35,065-3(d) and WIN 35,065-2(l), showed a

500-fold stereospecificity for the l-form.

Mouse brain extracts contained material that inhibited the binding

of [

3H]cocaine to mouse cerebrocortical membranes (Reith et al.

1980b). The material was only partially purified, and many

attempts (unpublished) at increasing the specific activity of the

inhibitor(s) were unsuccessful. It is not known whether the

inhibitory material that we studied is specific toward cocainebinding sites associated with serotonin uptake. Hanbauer and

colleagues (1985) have reported the existence of endogenous

inhibitors of the dopamine-related binding sites.

INTERACTION OF COCAlNE WITH THE SEROTONIN

TRANSPORTER: A MODEL SYSTEM

Although brain slices and synaptosomes have been used successfully

for demonstration of high-affinity uptake of serotonin and the effect

of cocaine, the description of the precise properties of the transporter is confounded in these preparations by the presence of storage

granules, enzymes involved in the metabolism of serotonin, and Na+,

K

+

-ATPase, which plays a role in the maintenance of the Na+

gradient. A less complex preparation of plasma membrane vesicles

can be obtained after osmotic shock of synaptosomes that has been

proven extremely useful for the study of the transport of gammaaminobutyric acid (Kanner 1978), glutamate (Kanner and Sharon 1978;

Kanner and Marva 1982), and tryptophan (Herrero et al. 1985).

Plasma membrane vesicles from blood platelets have often been used

as a model system for brain serotonergic nerve terminals (Stahl and

Meltzer 1978). However, recent studies have emphasized differences

that exist between blood platelets and brain serotonergic nerve

terminals.

28
TABLE 1. Cocaine congeners used in study of structure-activity

relationships

R1 R2

R3

R4

R5

R2(3)/R3(2)

H/COOCH3

or H

R4(5)/R5(4)

H/(OCO)-Phe or

OCOCHCH2OHPHE

Cocaine Axial Equatorial

(+)-pseudococaine Equatorial Equatorial

Benzolypseudotropine Equatorial

Benzoyltropine Axial

Allfocoine Axial Axial

Allopseudococaine Equatorial Axial

Atropine Axial

Win 35, 065-2 (1) Axial Equatorial

Win 35,065-3 (d) Axial Equatorial

Win 35,140 Equatorial Equatorial

Therefore, in our laboratory, plasma membrane vesicles from mouse

cerebrocortical synaptosomes were used as a tool to examine the

serotonin transporter. Synaptosomes were isolated and osmotically

shocked and resealed as described by Kanner (1978). In 10 seconds,

the concentration of serotonin doubled in assays of uptake in

membrane vesicles. The uptake of serotonin was saturable, with a

Km in the range of the values reported for synaptosomes (50 to 150

nM) (Ross 1982). In contrast to synaptosomes, plasma membrane

vesicles have no exogenous energy source for uptake except for ion

gradients. It was necessary to have an Na+ gradient ([Na+]out

greater than [Na+]in, and uptake was stimulated by the presence of

internal K

+

 and by a K

+

 gradient ([K+]in greater than [K+]out). A

membrane potential per se did not drive uptake. Cocaine inhibited

uptake of serotonin, with an IC50 value of 347 nM and with a Hill

number of 0.9, consonant with a mechanism of competitive inhibition.

In future experiments, this model system will be used as a tool to

study in more detail the relationship between the site where cocaine

29
interacts to inhibit uptake and the substrate recognition site for

serotonin.

INTERACTION OF COCAINE WITH THE DOPAMINE TRANSPORTER:

STRUCTURE-ACTIVITY RELATlONSHIP AND BEHAVIOR

The structure-activity relationship of cocaine congeners for the

dopamine uptake transporter was generally the same as that for the

serotonin transporter. The following were exceptions to this rule:

removal of the ester linkage between the tropane and phenyl rings

made the molecule more potent toward the dopamine transporter

(Reith et al. 1986b); N-demethylation (to norcocaine) resulted in some

loss of activity; and C3 epimerization of benzoylpseudotropine

(yielding benzoyltropine) had a greater effect in reducing the potency

toward the dopamine transporter (sixfold) than toward the serotonin

transporter (threefold). Such differences may indicate that cocaine

and cocaine congeners interact at different sites on the dopamine and

serotonin transporters, or that there are different environments

around the cocaine-binding sites on the two types of transporters

(Reith et al. 1986b). The correlation we observed between the

potencies of cocaine congeners in inhibiting Na+-dependent cocaine

binding and in inducing stereotyped sniffing (Reith et al. 1986b) is in

agreement with the notion that the Na+-dependent cocaine-binding

sites are involved in cocaine-induced increases in stereotyped

behavior; this, in turn, fits in with the hypothesis that striatal

dopamine is involved in the generation of such behavior (Beninger

1983). It should be emphasized here that it was necessary to

administer the cocaine congeners intracerebroventricularly in order to

observe the stereotyped behavior, because intraperitoneal administration of most cocaine congeners exhibited a depressant rather than a

central stimulatory effect on behavior (see below).

RELATlONSHlP OF THE CHARACTERISTICS OF COCAINE BINDING

TO ITS EFFECTS ON MONOAMINE UPTAKE

If an inhibitor competes for binding or uptake by a competitive

mechanism, its IC50 can be converted to an inhibition constant

(Cheng and Prusoff 1973). However, caution should be exercised in

comparing the absolute values of potencies of compounds in uptake

and binding assays. Conditions under which the assays are carried

out should be considered. For instance, Tris-HCI buffer inhibits

[

3H]cocaine binding and shifts the potencies of drugs toward higher

values (Reith et al. 1986b), and brain slices generally give higher

IC50 values for inhibitors of monoamine uptake than synaptosomal

30
preparations (Ross and Renyi 1975). Thus, in the striatum, there is

good correspondence between the absolute value of drugs in inhibiting

[

3H]cocaine binding to membranes and in inhibiting dopamine uptake

into the slices (Kennedy and Hanbauer 1983), while an example of

lack of correspondence is the binding of [

3H]imipramine to serotonin

uptake sites in brain, which has a binding affinity an order of

magnitude smaller (Langer et al. 1980; Reith et al. 1983b) than the

inhibition constant of imipramine in inhibiting [

3H]serotonin uptake

into brain synaptosomes (Koe 1976). We have shown that this

discrepancy is probably caused by the different temperatures at which

the two assays are routinely carried out: 0 °C for imipramine

binding and 37 °C for serotonin uptake (Reith et al. 1984c).

In the absence of Tris-HCI and presence of 50 mM Na+, there is a

good agreement between the absolute values for cocaine and cocaine

congeners in inhibiting [

3H]cocaine binding to cerebrocortical

membranes and the values in inhibiting [

3H]serotonin uptake into

cerebrocortical synaptosomes; there is also a good agreement between

the inhibition of [

3HIcocaine binding to striatal membranes and that

of [

3H]dopamine uptake into striatal synaptosomes (Reith et al.

1986b). These results are consonant with the suggestion that cocaine

analogs are competitive inhibitors of monoamine uptake and are not

translocated themselves by the uptake carrier. In contrast, substrates

such as serotonin or dopamine can require higher levels in inhibiting

the binding of a ligand to the uptake carrier than for inhibition in

the uptake assay because transport is a multistep reaction whereas

binding is not. More serotonin and dopamine are required to inhibit

[

3H]cocaine binding than to inhibit monoamine uptake into synaptosomes (Reith et al. 1986b). lt is not necessary to invoke an

allosteric linkage between the binding sites and the substrate

recognition sites for uptake. Attempts at demonstrating an allosteric

relationship have not been successful. For instance, there is little or

no effect of the substrates serotonin and dopamine on the dissociation rates of [

3H]cocaine binding (Reith et al. 1986a). Drugs that

have releasing effects in addition to uptake inhibitory effects, such

as amphetamine, may produce an IC50 in inhibiting uptake that is

lower than the true value because of the dopamine-releasing effect

of amphetamine.

INTERACTlON OF COCAlNE WlTH THE DOPAMINE TRANSPORTER:

THE EFFECT OF LONG-LASTING BLOCKADE BY METAPHlT

Metaphit (1-(1-(3-isothiocyanathophenyl)-cyclohexyl)piperidine), an

analog of phencyclidine (PCP), has been proposed to specifically

31
acylate PCP receptors and not opioid, muscarinic, or benzodiazepine

receptors (Rafferty et al. 1985). Since PCP has appreciable affinity

for the dopamine uptake sites (Kennedy and Hanbauer 1983;

Schoemaker et al. 1985), we examined the possibility of using

metaphit as a long-acting blocker of dopamine uptake sites for the

antagonism of effects of cocaine mediated by the dopamine transporter. Treatment of striatal membranes with metaphit produced a

concentration-dependent loss of [

3H]cocaine binding, due to an

effect on maximal binding rather than to the affinity (Berger et al.

1986). Since the membranes were washed three times after metaphit

treatment, these results suggest that metaphit binds irreversibly to,

or dissociates slowly from, the Na’dependent cocaine-binding sites.

Similar results have been repotted recently by Schweri et al. (1987)

for the binding of [

3H]methylphenidate to rat striatal membranes.

The behavioral effects of PCP in inducing stereotypy and ataxia are

antagonized by pretreating rats intracerebroventricularly with

metaphit 24 hours prior to PCP administration (Contreras et al. 1985),

and the density of PCP receptors is reduced by pretreating animals 24

hours prior with metaphit both intracerebroventricularly and

intravenously (Contreras et al. 1986). Pretreatment of mice with

metaphit (20 mg/kg intravenously) antagonizes the locomotor

stimulation caused by various psychostimulant drugs tested 24 hours

later (Sershen et al. 1988). Metaphit antagonizes drugs that exert

their hypermotive effect primarily by inhibition of the uptake of

dopamine, such as methylphenidate, mazindol, cocaine, and GBR 12909.

Subsequent experiments did not demonstrate a blockade of the

dopamine transporter at the time of the behavioral testing. First,

pretreatment with metaphit did not reduce the in vivo labeling of the

dopamine transporter with [

3H]GBR 12935. Second, pretreatment with

metaphit did not antagonize the locomotor stimulation evoked by

amphetamine, in spite of the fact that the effect of amphetamine

involves the release of dopamine by the same carrier that translocates dopamine into the terminal. Third, ex vivo experiments did

not indicate any difference between metaphit- and saline-pretreated

animals in the uptake of dopamine into synaptosomes or slices

(Sershen et al. 1988).

The possibility was considered that metaphit affects the locomotor

responses to stimulant drugs by a mechanism unrelated to uptake

blockade. A study of levels of monoamines and their metabolites in

various brain regions showed a consistent increase in homovanillic

acid (HVA) in all regions studied--striatum, olfactory tubercle, and

cerebral cortex-24 hours after pretreatment with metaphit (Sershen

et al. 1988). Metaphit had no effect on the disappearance rate of

32
3, 4-dihydroxyphenylacetic acid and HVA from the striatum during

inhibition of monoamine oxidase with pargyline. lf the increase in

HVA reflects a greater rate of dopamine catabolism in metaphitpretreated animals, it could explain the lack of locomotor stimulation

of blockers of dopamine uptake in these animals, resulting from a

rapid breakdown of extracellular accumulated dopemine.

INTERACTlON OF COCAlNE WlTH SODlUM CHANNELS:

STRUCTURE-ACTIVITY RELATIONSHIP AND BEHAVlOR

As do other local anesthetic drugs, cocaine can be expected to

interfere with the uptake of sodium through voltage-regulated sodium

channels. For studying the interaction of cocaine and cocaine

congeners with sodium channels, we used [

3H]batrachtochotoxininA20 benzoate (BXT-B) as a ligand for the proteins involved in the

gating of the channel. It is known that local anesthetic drugs inhibit

BTX-B binding to sodium channel preparations from brain, most likely

by an allosteric mechanism shifting the equilibrium of the channel

toward the nonconducting state (Postma and Catterall 1984; Reith

et al. 1986c). We assessed the potency of cocaine congeners by two

methods that gave the same result: (1) determination of the

inhibition constant for the equilibrium binding of [

3H]BTX-B, and (2)

estimation of the concentration that is required to enhance by a

factor of two the dissociation rate of [

3H]BTX-B binding initiated by

an excess of aconitine, an alkaloid that acts at the same site as

BTX-B (Catterall 1977). The observed enhancement of the dissociation rate agrees with the suggestion that cocaine and cocaine

congeners act at a site in the sodium channel that is allosterically

linked with the site labeled by [3H]BTX-B (Reith et al. 1986c). As

found for the activity toward monoamine transporters, the ester

linkage between the tropane and phenyl rings of cocaine was not

necessary for local anesthetic activity: WIN 35,140 and WIN 35,094

(table 1) had the same potency as cocaine, whereas WIN 35,428 and

WIN 35,065-3 were approximately threefold to fourfold less potent

than cocaine. Moving the C-benzoyl group on C3 from an equatorial

(benzoylpseudotropine) to an axial (benzoyltropine) position reduced

the potency threefold, and removal of the O-benzoyl group from C3

(ecgonine methylester) had a greater effect in reducing potency

(1 ,000-fold) than modifying the group (atropine, sixfold reduction).

In contrast to the structure-activity relationships for monoamine

uptake inhibition, however, N-demethylation (norcocaine) increased

the inhibitory activity fourfold. Moving the carbomethoxyl group on

C2 from an axial (cocaine and WIN 35,428) to an equatorial ((+)-

pseudococaine and WIN 35,140) position increased the potency by a

33
factor of 4; (-)-pseudococaine, which is the enantiomer of (+)-

pseudococaine and has the C2- and C3-substituents also in the

equatorial positions, was three times less potent than (+)-pseudococaine.

To our surprise, intraperitoneal administration of most cocaine

congeners did not induce locomotor stimulation as cocaine itself did.

Instead, spontaneous locomotor behavior was inhibited; the animals

were less alert, but had the eyes open and did not assume postures

normally associated with sleep (Reith et al. 1985b). There was a

reasonable agreement between the rank order of potencies of cocaine

congeners and other local anesthetics in depressing locomotor

behavior and in inhibiting [

3H]BTX-B binding (Reith et al. 1986c).

Such a comparison has to be judged with caution, since drugs applied

in vivo must be transported to their presumed sites of actions, are

susceptible to metabolizing enzymes, and may have multiple effects.

Never the less, the observed correlation is consonant with an involvement of sodium channels in the inhibition of locomotor behavior by

cocaine congeners. The present results do not indicate whether

central sodium channels are involved; there is some evidence

suggesting action at the peripheral level (Reith and Lajtha 1986). It

is possible that multiple targets at the peripheral levels are involved

in the effect of local anesthetics, such as the cardiovascular system,

the neuromuscular junction, and the lungs (Ritchie and Greene 1980).

INTERACTION OF COCAlNE WlTH SODIUM CHANNELS: THE

PHOSPHATlDYLlNOSlTOL RESPONSE

Hydrolysis of inositol phospholipids has received much recent

attention as an effector system coupled to alpha1-adrenergic, 5-HT2-,

muscarinic-cholinergic, and certain peptidergic receptors, with

diacylglycerol and inositol 1,4,5-triphosphate as the second messengers (Berridge et al. 1983; Nahorski et al. 1986). In addition to

agonists acting on these receptors, various agents that activate

voltage-dependent sodium channels have been found to stimulate

polyphosphoinositide turnover (Gusovsky et al. 1986), suggesting that

sodium channel activity per se can have an input to phosphatidylinositol systems. The blockade of monoamine uptake by cocaine may lead

to an enhanced polyphosphoinositide turnover in neurons innervated

by norepinephrine and serotonin, while the inhibition of sodium flux

by cocaine may lead to reductions in inositol phospholipid hydrolysis.

Experiments were carried out in our laboratory to test the latter

possibility. Production of inositol phosphates in the presence of

lithium slices was taken as an index of phosphatidylinositol phosphate

34
hydrolysis, and [

14C]guanidine flux through activated sodium channels

in synaptosomes served as a model for sodium flux. lt was found

that cocaine indeed interacts with the phosphoinositide system at

concentrations that inhibit sodium flux. The stimulation of inositide

hydrolysis by the sodium channel activators batrachotoxinin (BXT),

aconitine, and scorpion venom in mouse cerebrocortical slices was

reduced by the presence of 5 μM tetrodotoxin, a blocker of sodium

channels. Cocaine and tetracaine were found to shift the doss

response curve of BTX in stimulating inositide hydrolysis to the right.

In addition, cocaine and tetracaine inhibited BTX-elicited (0.05 μM)

inositide hydrolysis, with IC50 values of approximately 100 and 2 μM,

respectively. Cocaine and tetracaine also inhibited veratridinestimulated (60 μM) [

14C]guanidine uptake into synaptosomes, with

IC50 values of approximately 140 and 25 μM, respectively. These

results do not negate the possibility that cocaine can lead to

increased inositide hydrolysis by monoamine uptake blockade. The

data, however, do show that the increase in inositide hydrolysis

induced by a sodium channel activator can be counteracted by

concentrations of cocaine that inhibit the flux of guanidine through

activated sodium channels (Kim and Reith 1988).

REPEATED COCAlNE ADMINISTRATION. TOLERANCE, AND

SENSITlZATlON, AND THE CONVERSION TO N-HYDROXYNORCOCAlNE

Repeated intraperitoneal injections of cocaine in rodents can produce

both sensitization and tolerance to its behavioral effects (Post and

Contel 1983). These results are related to the type of behavior under

study, such as locomotion and stereotypy, which are usually susceptible to the sensitization phenomena. Cocaine discrimination and

operant conditioning produce tolerance effects. Three other variables

that have been considered in studies of a related psychostimulant

drug, amphetamine, have not been addressed in cocaine studies:

(1) the size of the dose, (2) the regimen of administration (intermittent vs. continuous), and (3) pharmacodynamics.

(1) The size of the dose was studied with cocaine and WIN 35,065-2,

one of the few phenyltropane analogs of cocaine that, upon

intraperitoneal administration, stimulate locomotor behavior of

mice. Locomotor sensitization to the locomotor stimulatory

effects developed during daily administration of either compound,

with similarly shaped dose-response curves (Reith 1986).

Dosages giving optimal sensitization were 25 mg/kg/day for

cocaine and 3 mg/kg/day for WIN 35,065-2. At higher doses,

35
(2)

(3)

cocaine produced tolerance to stimulation on day 3, as did WIN

35,065-2 (Reith 1986). This tolerance at the higher dose did not

appear to result from an increase in nonambulatory (or stereotyped) behavior. Therefore, the size of the dose of cocaine is a

crucial variable in dosage schedules for studying the sensitization to its effect on locomotor behavior.

Intermittent SC and IP injections of cocaine (20 mg/kg: total

of 21 injections, 430 mg/kg for each mouse) for 18 days resulted

in locomotor stimulation of mice upon challenge with cocaine on

the 25th or 26th day, compared with no locomotor stimulation in

a saline-pretreated group. In contrast to the sensitization by

intermittent cocaine administration, tolerance was found upon

challenge after continuous administration of cocaine by minipumps implanted SC (25 mg/kg/day; total 450 mg/kg for each

animal) on a similar schedule (Reith et al. 1987). Therefore,

the mode of administration, intermittent vs. continuous, is an

important factor determining the behavioral response at the

endpoint.

No differences were found between the above sensitized and

tolerant groups in the levels of cocaine and benzoylecgonine in

plasma and brain 12 minutes after IP administration of a

challenge dose of cocaine, suggesting that, in these chronic

experiments, the changes in the locomotor response are not

accounted for by the dispositional effects. In contrast, in

animals treated daily for 2 or 3 days IP with cocaine and

challenged with cocaine 1 day later, there was both a greater

locomotor stimulation and a higher level of cocaine in brain

than in saline-pretreated animals, suggesting a dispositional

effect. Among individual animals, there was a positive

correlation between their locomotor stimulation by the challenge

dose and their brain concentration (Reith et al. 1987). lt

suffices to consider the brain level of cocaine alone for

dopamine-related effects of cocaine on locomotion, because

the only active metabolite in this respect, norcocaine, reaches

concentrations in the brain that are less than 20 percent of

those of cocaine at all timepoints after IP administration of

cocaine (Benuck et al. 1987).

At the same time that levels of cocaine and norcocaine peak in the

brain after IP administration of cocaine, another metabolite reaches

its highest concentration in the brain: N-hydroxynorcocaine. The

amounts of this metabolite are at least as high as norcocaine itself

36
(Benuck et al. 1988). This is of considerable interest because, in

liver, N-hydroxynorcocaine has been found to be the precursor of

norcocaine nitroxide, a free radical that produces liver toxicity

(Shuster et al. 1983).

CONCLUSION

The potent effect of cocaine in inhibiting uptake of monoamines has

been known for a long time. A new approach for studying how

cocaine interacts with these uptake systems has opened up with the

development of receptor-binding assays of [

3H]cocaine binding. The

binding assays are a measure of the site where cocaine acts to inhibit

monoamine uptake, enabling us to study the relationship between this

site and the substrate recognition site where the monoamine binds

before it is translocated. The interesting possibility exists that these

sites are distinct and are coupled by an allosteric linkage. If so, it

is possible to envision (1) a putative cocaine antagonist that prevents

the action of cocaine but does not interfere with the normal uptake

of monoamines; and (2) regulation of monoamine uptake by a site that

is acted upon by an endogenous ligand different from the monoamine

itself. Evidence for regulation has come from studies by Hanbauer

and colleagues (1985) with the dopamine transporter. In our

laboratory, we have not been successful in obtaining evidence to

support the allosteric model, as illustrated by the following observations. First, the absolute potencies of cocaine congeners and

antidepressant drugs in inhibiting [

3H]cocaine binding agree closely

with those in inhibiting monoamine uptake into synaptosomes,

consonant with the simple model in which drugs bind to the same site

as the monoamine but are not themselves translocated. Second, for

most inhibitors, the competition curves in the binding and uptake

assays are steep, displaying Hill slopes not very different from unity.

Third, no evidence was found for changes in the dissociation rate of

[

3H]cocaine-binding as a result of the presence of monoamines, an

effect that is often associated with allosteric systems. Fourth, long

lasting or irreversible blockade of [

3H]cocaine binding sites in the

striatum by metaphit, an isothiocyanate derivative of PCP, causes a

decrease in both [

3H]cocaine-binding and [3H]dopamine uptake.

Like other local anesthetic drugs, cocaine inhibits sodium flux

through voltage-regulated sodium channels. The blockade of flux is

probably not due to mere occlusion of the channel, but involves an

allosteric effect in shifting the equilibrium between the nonconducting

and conducting state of the sodium channel towards the nonconducting state. This state is associated with closed gates, blocking

37
passage of sodium, and has a low affinity for [3H]BTX-B associated

with a rapid dissociation of [

3H]BTX-B from its binding site. The

decrease in polyphosphoinositide turnover by cocaine’s inhibition of

sodium influx possibly interferes with processes further downstream in

the sequence of events triggered by second messengers generated by

inositide hydrolysis. Changes in protein kinase C-catalyzed protein

phosphorylation by cocaine in the brain could be important for

processes specific for the central nervous system, such as learninginduced alterations in potassium channel conductance involving second

messenger systems.

lt is clear that there are many more sites of action of cocaine than

there are binding sites with high affinity for [

3H]cocaine. Furthermore, there are probably more binding sites for [

3H]cocaine than

have been demonstrated so far. lt is possible that there are discrete

areas in the brain where [

3H]cocaine binds with characteristics

different from those of the known sites. For instance, recent reports

of self-administration of cocaine in the frontal cortex make it of

interest to reinvestigate [

3H]cocaine-binding in this area. The

behavioral models described above allow us to compare animals that

are sensitized or tolerant to the locomotor stimulatory effect of

cocaine, and it will be of great interest to study the regulation of

t

3H]cocaine binding sites relating behavioral and biochemical measures

derived from the same animal.

REFERENCES

Beninger, R.J. The role of dopamine in locomotor activity and

learning. Brain Res Rev 6:173-196, 1983.

Benuck, M.; Reith, M.E.A.; and Lajtha, A. Pharmacokinetics of

systemically administered cocaine and locomotor stimulation in mice.

J Pharmacol Exp Ther 243: 144-149, 1987.

Benuck, M.; Reith, M.E.A.; and Lajtha, A. Presence of the toxic

metabolite N-hydroxy-norcocaine in brain and liver of the mouse.

Biochem Pharmacol 37:1169-1172, 1988.

Berger, P.; Jacobson, A.E.; Rice, K.C.; Lessor, R.A.; and Reith, M.E.A.

Metaphit, a receptor acylator, inactivates cocaine binding sites in

striatum and antagonizes cocaine-induced locomotor stimulation in

rodents. Neuropharmacology 25:931-933, 1986.

Berridge, M.J.; Dawson, R.M.C.; Downes, C.P.; Heslop, J.P.; and Irvine,

R.F. Changes in the levels of inositol phosphates after agonistdependent hydrolysis of membrane phosphoinositides. Biochem J

212:473-482, 1983.

38
Catterall, W.A. Activation of the action potential Na+

 ionophore by

neurotoxins. An allosteric model. J Biol Chem 252:8669-8676, 1977.

Cheng, Y.-C., and Prusoff, W.H. Relationship between the inhibition

constant (Ki) and the concentration of inhibitor which causes 50

percent inhibition (IC50) of an enzymatic reaction. Biochem

Pharmacol 22:3099-3108, 1973.

Contreras, P.C.; Johnson, S.; Freedman, R.; Hoffer, B.; Olsen, K;

Rafferty, M.F.; Lessor, R.A.; Rice, K.C.; Jacobson, A.E.; and

O’Donohue, T.L. Metaphit, an acylating ligand for phencyclidine

receptors: Characterization of in vivo actions in the rat. J

Pharmacol Exp Ther 238:1101-1107, 1986.

Contreras, P.C.; Rafferty, M.F.; Lessor, R.A.; Rice, KC.; Jacobson,

A.E.; and O’Donohue, T.L. A specific alkylating ligand for

phencyclidine (PCP) receptors antagonizes PCP behavioral effects.

Eur J Pharmacol 111:405-406, 1985.

Cotman, C.W. Isolation of synaptosomal and synaptic plasma

membrane fractions. In: Fleischer, S., and Packer, L., eds.

Methods of Enzymology. Vol. 31. New York: Academic Press,

1974. pp. 445-452.

Gusovsky, F.; Hollingworth, E.B.; and Daly, J.W. Regulation of

phosphatidylinositol, turnover in brain synaptoneurosomes:

Stimulatory effects of agents that enhance influx of sodium ions.

Proc Natl Acad Sci USA 83:3003-3007, 1986.

Hanbauer, I.; Kennedy, L.T.; Missale, M.C.; and Bruckwick, E.C.

Cocaine binding sites located in striatal membranes are regulatory

sites for dopaminergic synapses. In: Langer, S.Z.; Takahashi, R.;

Segawa, T.; and Briley, M., eds. New Vistas in Depression. Vol.

40. Oxford: Pergamon Press, 1985. pp. 4149.

Herrero, E.; Aragon, M.C.; Diez-Guerra, J.; Valdivieso, F.; and

Gimenez, C. Ontogenetic studies on tryptophan transport into

plasma membrane vesicles derived from rat brain synaptosomes:

Effects of thyroid hormones. Neurochem Res 10:579-589, 1985.

Kanner, B.I. Active transport of aminobutyric acid by membrane

vesicles isolated from rat brain, Biochemistry 17:1207-1211, 1978.

Kanner, B.I., and Marva, E. Efflux of L-glutamate by synaptic plasma

membrane vesicles isolated from rat brain. Biochemistry 21:3143-

3147, 1982.

Kanner, B.I., and Sharon, I. Active transport of L-glutamate by

membrane vesicles isolate from rat brain. Biochemisty 17:3949-3953,

1978.

Kennedy, L.T., and Hanbauer, I. Sodium-sensitive cocaine binding to

rat striatal membrane: Possible relationship to dopamine uptake

sites. J Neurochem 41:172-178, 1983.

39
Kim, S.S., and Reith, M.E.A. Inhibition by cocaine of inositol

phospholipid hydrolysis induced by the sodium channel activator

batrachotoxin in mouse cerebral cortex. Biochem Pharmacol 37:773-

775, 1988.

Koe, B.K. Molecular geometry of inhibitors of the uptake of

catecholamines and serotonin in synaptosomal preparations of rat

brain. J Pharmarcol Exp Ther 199:649-661, 1976.

Langer, S.Z.; Moret, C.; Raisman, R.; Dubocovich, M.L, and Briley, M.

High affinity [

3H]imipramine binding in rat hypothalamus:

Association with uptake of serotonin but not of norepinephrine.

Science 210:1133-1135, 1980.

Nahorski, S.R.; Kendall, D.A.; and Batty, I. fieptors and phosphoinositide metabolism in the central newous system. Biochem

Pharmacol 35:2447-2453, 1986.

Post, R.M., and Contel, N.R. Human and animal studies of cocaine:

Implications for development of behavioral pathology. In: Creese,

I., ed. Stimulants: Neurochemical, Behavioral and Clinical

Perspectives. New York: Raven Press, 1983. pp. 169-203.

Postma, S.W., and Catterall, W.A. Inhibition of binding of

[

3H]batrachotoxinin A20 benzoate to sodium channels by local

anesthetics. MOl Pharmacol 25:219-227, 1984.

Rafferty, M.F.; Mattson, M.; Jacobson, E.A.; and Rice, KC. A

specific acylating agent for the [

3H]phencyclidine receptors in rat

brain. FEBS Lett 181:318-322, 1985.

Reith, M.E.A. Effect of repeated administration of various doses of

cocaine and WIN 35,065-2 on locomotor behavior of mice. Eur J

Pharmacol 130:65-72, 1986.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Saturable [

3H]cocaine

binding in central nervous system of mouse. Life Sci 27:1055-1062,

1980a.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Endogenous peptide

inhibiting [

3H]cocaine binding in mouse brain. Neurochem Res

5:1291-1299, 1980b.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Binding of [

3H]cocaine in

mouse brain: Kinetics saturability. J Recept Res 2:233-243, 1981.

Reith, M.EA; Sershen, H.; Allen, D.L; and Lajtha, A. A portion of

[

3H]cocaine binding in brain is associated with serotonergic

neurons. MOl Pharmacol 23:600-606, 1983a.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. High- and low-affinity

binding of [

3H]imipramine in mouse cerebral cortex. J Neurochem

40:389-395, 1983b.

Reith, M.E.A.; Meislet, BE.; Sershen, H.; and Lajtha, A. [

3H]cocaine

binding in brain is inhibited by Trias (hydroxymethyl)aminomethane.

J Neurosci Methods 12:151-154, 1984a.

40
Reith, ME-A.; Allen, D.L.; Sershen, H.; and Lajtha, A. Similarities

and differences between high affinity binding sites for cocaine and

imipramine in mouse cerebral cortex. J Neurochem 43:249-255,

1984b.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Thermodynamics of the

interactions of tricyclic drugs with binding sites for [

3H]imipramine

in mouse cerebral cortex. Biochem Pharmacol 33:4101-4104, 1984c.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Binding of imipramine and

cocaine to a model lipid membrane. Neurochem Res 9:965-977,

1984d.

Reith, M.E.A; Meisler, B.E.; Sershen, H.; and Lajjha, A Sodiumindependent binding of [

3H]cocaine in mouse striatum is serotonin

related. Brain Res 342:145-148, 1985a.

Reith, M.E.A.; Meisler, B.E.; and Lajjha, A Locomotor effects of

cocaine, cocaine congeners, and local anesthetics in mice.

Pharmacol Biochem Behav 23:831-836, 1985b.

Reith, M.E.A., and Lajtha, A Locomotor depression in mice by

norcocaine does not involve central a2-adrenergic or presynaptic

dopamine receptors. Pharmacol Biochem Behav 24:305-307, 1986.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Binding sites for

[

3H]cocaine in mouse striatum and cerebral cortex have different

dissociation kinetics. J Neurochem 46:309-312, 1986a.

Reith, M.E.A.; Meisler, B.E.; Sershen, H.; and Lajtha, A. Structural

requirements for cocaine congeners to interact with dopamine and

serotonin uptake sites in mouse brain and to induce stereotyped

behavior. Biochem Pharmacol 35:1123-1129, 1986b.

Reith, M.E.A.; Kim, S.S.; and Lajtha, A. Structural requirements for

cocaine congeners to interact with [

3H]bratachotoxinin A20

benzoate binding sites on sodium channels in mouse brain synaptosomes. J Biol Chem 261:7300-7305, 1986c.

Reith, M.E.A.; Benuck, M.; and Lajtha, A. Cocaine disposition in the

brain after continuous or intermittent treatment and locomotor

stimulation in mice. J Pharmacol Exp Ther 243:281-287, 1987.

Ritchie, J.M., and Greene, N.M. Local anesthetics. In: Goodman,

LS., and Gilman, A., eds. The Pharmacological Basis of Therapeutics. 6th ed. New York: MacMillan Publishing Company, 1980.

pp. 300-320.

Ross, S.B. The characteristics of serotonin uptake systems. In:

Osborne, N.N., ed. Biology of Serotonergic Transmission.

New York: Wiley, 1982. pp. 159-195.

41
Ross, S.B., and Renyi, A.L Tricyclic antidepressant agents. I.

Comparison of the inhibition of the uptake of 3H-noradrenaline

and 14C-5-hydroxytryptamine in slices and crude synaptosome

preparations of the midbrain-hypothalamus region of the rat

brain. Acta Pharmacol Toxicol 36:382-394, 1975.

Schoemaker, H., and Langer, S.Z. [3H]-8-OH-DPAT labels the

serotonin transporter in the rat striatum. Eur J Pharmacol

124:371-373, 1986.

Schoemaker, H.; Pimoule, C.; Arbilla, S.; Scatton, B.; Javoy-Agid,

F.; and Langer, S.Z. Sodium dependent [

3H]cocaine binding

associated with dopamine uptake sites in the rat striatum and

human putamen decrease after dopaminergic enervation and in

Parkinson disease. Naunyn-Schmiedebergs Arch Pharmacol

329:227-235, 1985.

Schweri, M.M.; Jacobson, A.E.; Lessor, R.A.; and Rice, K.C.

Metaphit irreversibly inhibits [

3H]threo-(±)-methylphenidate

binding to rat striatal tissue. J Neurochem 48:102-105, 1987.

Sershen, H.; Berger, P.; Jacobson, A.E.; Rice, K.C.; and Reith,

M.E.A. Metaphit prevents locomotor activation induced by various

psychostimulants and interferes with the dopaminergic system in

mice. Neuropharmacol 27:23-30, 1988.

Shuster, L.; Casey, E.; and Welankiwar, S.S. Metabolism of cocaine

and norcocaine to N-hydroxynorcocaine. Biochem Pharmacol

32:3045-3051, 1983.

Stahl, S.M., and Meltzer, H.Y. A kinetic and pharmacologic

analysis of 5-hydroxytryptamine transport by human platelets and

platelet storage granules: Comparison with central serotonergic

neurons. J Pharmacol Exp Ther 205:118-132, 1978.

Whittaker, V.P. The synaptosome. In: Lajtha, A., ed. Handbook

of Neurochemistry. Vol. II. New York: Plenum Press, 1969.

pp. 327-364.

ACKNOWLEDGMENTS

The experimental work was supported by research grant DA 03025 from

the National Institute on Drug Abuse and by a grant for cocaine

research from the New York State Department of Mental Hygiene.

42
AUTHOR

Maarten E.A. Reith, Ph.D.

Center for Neurochemistry

The Nathan S. Kline Institute

for Psychiatric Research

Ward’s Island

New York, NY 10035

43
Modulation of Cocaine Receptors

lngeborg Hanbauer

lNTRODUCTlON

The understanding of the neuronal sites of action of cocaine are of

great importance since cocaine is widely abused and produces, in man,

behavioral abnormalities, as well as vascular and cardiac toxicity.

The main pharmacological actions of cocaine are local anesthetic and

central stimulant. In neuronal tissue, cocaine blocks the uptake of

catecholamines and serotonin. This action was thought to be

important to explain its behavioral effect because it altered central

synaptic transmission. Cocaine has been demonstrated to bind to

specific sites on brain membranes (Reith et al. 1980), where it can be

displaced by compounds that fall in the pharmacological categories of

nonamphetamine stimulants and antidepressants. Although the

presence of saturable binding sites for cocaine was demonstrated in

various types of tissues (Sershen et al. 1982), the physiological role

of these sites is not well understood. At the present time, research

efforts in our laboratory are focusing on several unresolved questions

regarding these binding sites: (1) Does cocaine bind merely to drug

acceptor sites or does it label receptors of physiological significance?

(2) If cocaine binding sites are physiological receptors, are they

linked to specific transducer systems? and (3) Does cocaine bind to

recognition sites for an endogenous ligand?

MODULATION OF COCAlNE BlNDlNG SlTES BY INORGANIC IONS

Reports in the literature indicated that the characteristics of the

specific binding of a number of radioligands changed with the

presence of specific ions (Enna and Snyder 1975; Baudry and Lynch

1981; Briley and Langer 1981). Studies on the effect of various ions

on the specific binding of cocaine on synaptosomal membranes of

44
caudate nucleus are summarized in table 1. The presence of Na+

elicited a half-maximal increase in binding at about 11 mM NaCl and

a maximal increase at about 25 to 60 mM NaCI. In contrast, other

ions, including Li+, K+, and Ca2+ reduced cocaine binding to striatal

Type of Ion

(50 mM) Brain Area 3H-Cocaine Binding

TABLE 1. Modulation of specific 3H-cocaine binding by inorganic

substances

Na+ Caudete Nucleus

(dopaminergic terminals)

Na+ Hippocampus

Frontal Cortex

Cerebellum

Hypothalmus

K

+

Cuadet Nucleus

Ca2+ Hippocampus

Mg2+ Cerebellum

NOTE: Cocaine binding was measured, in the presence of 100 nM 3H-cocaine in 50 mM

tris buffer, pH 7.7. Nonspecific H-cocaine binding was measured in the

presence of 50 μM cocaine.

membrane preparations (Kennedy and Hanbauer 1983). This facilitation of cocaine binding by Na+

 appeared to be limited to only one

region of the brain, namely, the caudate nucleus, while in other

areas, including frontal cortex, hippocampus, and cerebellum, cocaine

binding was inhibited by the presence of Na+. Kinetic analysis of

saturation isotherms for cocaine binding demonstrated that in the

presence of 50 mM NaCl the number of 3H-cocaine binding sites was

increased without altering the affinity of these sites for cocaine

(Kennedy and Hanbauer 1983). Since Na+ increased 3H-cocaine

binding exclusively in caudate nucleus, it was of interest to determine

whether Na+-dependent cocaine binding was linked to a specific

neuronal site in this brain area. The effect of specific neuronal

45
lesions is summarized in table 2. lntrastriatal injection of kainic

acid, a neurotoxin that selectively destroys nerve cell bodies but

spares nerve endings, failed to alter Na+-increased cocaine binding.

In contrast, intraventrlcular infusion of 6-hydroxydopamine, a

TABLE 2. Effect of specific neuronal lesions on Na+-dependent

cocaine binding to striatal membranes

3H-Cocaine

K D (μM) BMax Dopamine

Type of Lesion pmol/mg (pmol/mg

protein) protein)

Sham-Operated 0.34±0.02 13.4±1.6 937±34

6-Hydroxydopamine 0.37±0.06 3.7±0.3 159±61

Kainic Acid 0.34±0.08 14.0±1.5 nm

*p<0.001 when compared with sham-operated group.

KEY: nm=not measured.

NOTE: Rats were injected with desmethylimipramine (25 mg/kg IP) before infusing

6-hydroxydopamine (250 μg / 20 μl) in 0.0 percent saline and 0.1 percent ascorbic

acid. Another group of rats were infused unilaterally in the caudate nucleus

with kainic acid (1 mg/2 1/rat). Cocaine binding was measured in the

presence of 50 mM NaCI, 50 nM tris buffer, pH 7.7, and various concentrations

of H-cocaine (0.01 to 1.0 μM). The values represent the mean ± SE of five

experiments and were obtained by Scatchard analysis of saturation isotherms.

neurotoxin that selectively destroys dopaminergic terminals, reduced

the density of cocaine binding sites to a level that is obtained when

measurements are carried out in the absence of Na+. When cocaine

binding in the presence of 50 mM NaCl was evaluated as a function

of dopamine content, both parameters were strongly correlated

(Kennedy and Hanbauer 1983). This anatomical relationship was

further supported by Pimoule and coworkers (1983), who reported that

the dopamine content and Na+dependent cocaine binding were greatly

reduced in putamen of postmortem brain from Parkinson’s diseased

individuals when compared with putamen from non-Parkinsonians.

The Na+-dependent cocaine binding to striatal membranes was most

46
TABLE 3. Displacement of 3H-cocaine from specific striatel binding

sites in the presence of 50 mM NaCl

Drug IC50(M)

Nomifensine 2x10-7

Benztropine 5x10-7

Phencylidine 8x10-7

+Amphetamine 10-5

SFK 38393 10-5

LY 141865 7x10-5

Dopamine 2x10-5

Fluoxetine 10-5

Tetracaine 8x10-5

Atropine 2x10-4

NOTE: IC50 values were determined by linear reqression analysis of dose response

curves and represent the mean of two to three experiment. Cocaine

binding was measured in presence of 100 μM 3H-cocaine, 50 mM NaCl, 50 mM

tris buffer pH 7.7.

potently inhibited by dopamine uptake blockers (table 3). Interestingly, dopamine itself was a relatively weak competitor for Na+

-

dependent cocaine binding sites. However, the IC50 value for

dopamine was significantly lower in the presence than in the absence

of Na+ (-NaCI: 1.9 x 103M; +NaCI: 2 x 10-5M). Specific blockers

of serotonin uptake were less effective and so also were D1 and D2

dopamine receptor ligands, amphetamine, atropine, or tetracaine.

Recently, Reith and coworkers (1985) obtained evidence that destruction of serotonergic nerve endings in caudate nucleus led to a

decrease of Na+-independent cocaine binding. lf this type of

lesioning absolutely spared dopaminergic terminals, this observation is

of great interest, because one of the explanations given for the

increased radioligand binding elicited by Na+ was that binding to

neurotransmitter uptake sites is facilitated (Baudry and Lynch 1981;

Rehavi et al. 1982). lf this hypothesis were to be correct, cocaine

binding sites may not be functionally linked to serotonin uptake sites.

But, in contrast, Reith and coworkers (1985) showed that serotonin

uptake blockers were highly specific in blocking Na+-independent

cocaine binding. Thus, it appears that cocaine binding sites in

caudate nucleus could be closely associated with the dopamine and

47
serotonin transporter systems, which may differ in their requirement

for Na+.

At the present time, the mechanism whereby Na+ facilitates cocaine

binding to dopaminergic nerve terminals is not known. Some reports

in the literature on other radioligand studies attributed Na+-induced

increases in radioligand binding to stabilization of the ligandrecognition site complex (Usdin et al. 1980) or alteration in the

conformation of the receptor (Pastemak et al. 1975). lf cocaine

binds to a transporter operative in monoamine uptake it is conceivable that Na+

 may facilitate cocaine binding by spatially exposing

more transporter sites.

ENDOGENOUS MODULATORS FOR COCAINE BINDING SITES IN

CAUDATE NUCLEUS

Although ample experimental evidence suggests that cocaine binding

sites may play a role in the regulation of synaptic transmission by

blocking neurotransmitter reuptake, there exists little published

information on the regulation of cocaine binding sites. As pointed

out in the previous section, in caudate nucleus, two types of cocaine

recognition sites are present that differ in ion sensitivity, in anatomical localization, and in their interaction with neurotransmitter

systems. Recently, our laboratory has contributed new evidence for

endogenous regulatory mechanisms for the striatal Na+-dependent

cocaine binding sites. One of these mechanisms involves an

interaction between dopamine and L-glutamate receptors (table 4)

while the other involves a synaptosomal membrane-bound regulatory

protein (Hanbauer et al. 1984) that belongs to the family of

TABLE 4. Regulation of cocaine recognition sites in caudate nucleus

Na+-Dependent Na+-Independent

Anatomical

Localization

Dopaminergic

Terminals

Serotonergic

Terminals

Small Neurons

Receptor-Receptor

Interaction

GABA

L-Glutamate

?

48
membrane-associated basic proteins and was found to be strikingly

similar to GABAmodulin (table 5).

Receptor-Receptior Interaction

The glutamatergic neurons located in the frontal and parietal cerebral

cortex project terminals into caudate nucleus, an area rich in Nmethyl-D-aspartate-sensitive receptor sites (Cotman et al. 1987).

Llgation of the pia mater vessels supplying blood to the frontal and

parietal cortex causes destruction of these cortical layers and results

in a degeneration of glutamatergic terminals in caudate nucleus, as

indicated by a greatly reduced D-aspartate uptake in striatal slices

(Walker and Fonnum 1983). Measurements of the Na+-dependent

cocaine binding carried out 14 days after the ligation of the pia

mater vessels revealed a significant increase in the Bmax of 3Hcocaine binding sites in ipsilateral caudate nucleus, while the affinity

for cocaine was not significantly different when compared with

contralateral caudate nucleus (Sanna and Hanbauer, in preparation).

The outcome of these experiments suggests that L-glutamate may

exert a negative modulation on the cocaine receptors present in

dopaminerglc terminals. lt is of interest to mention at this point

that several reports in the literature (Roberts and Anderson 1979;

Donzanti and Uretsky 1983; Rudolph et al. 1983) brought evidence for

a dopamine receptor-glutamate receptor interaction in caudate nucleus

and nucleus accumbens. In fact, L-glutamate was found to potentiate

the K

+

-elicited release of 3H-dopamine in rat striatal slices. Since

cocaine receptors on dopaminergic terminals appear to be functionally

linked to dopamine reuptake, it is tempting to speculate that the

TABLE 5. Extraction and purification of endogenous displacer of

3H-cocaine binding from caudate nucleus of fats

1. Synaptosomal fraction obtained by sucrose density gradient

centrifugation.

2. Synaptosomal pellet extracted with 1 M acetic acid at 90 °C.

Biogel-P10 filtration of supernatant fraction.

3. HPLC: μBondapak c18 preparative column eluted with linear

gradient of acetonitrile containing 0.1 percent trifluoroacetic acid

(15 to 40 percent acetonitrile in 50 minutes).

49
upregulation of cocaine receptor sites following degeneration of

glutamatergic terminals may also be associated with alterations of the

dopamine uptake system.

Regulation of Cocaine Binding Sites by a Synaptosomal

Basic Protein

The functional and anatomical association of cocaine binding sites

with the dopamine uptake system in caudate nucleus prompted us to

investigate whether an endogenous modulator may exist that would

interact with the binding sites to which cocaine binds with relatively

high affinity. In the search for an endogenous modulator, we

prepared extracts from striatal synaptosomes that were then tested

for the presence of material that showed cocaine-displacing activity.

Table 5 summarizes the purification steps of a cocaine bindinginhibitory protein with a molecular weight of 17,000 dalton. The

final purified material was further examined by isoelectric focusing,

which revealed that the cocaine binding inhibitor was a basic protein

with a pl = 10.4. To obtain information on the mode of interaction

of this inhibitor protein with cocaine binding sites, we measured 3Hcocaine saturation isotherms in absence and presence of the inhibitor

protein. The data obtained from Scatchard analysis of the 3H-cocaine

binding curves are shown in table 6 and clearly demonstrate that the

cocaine binding inhibitor was not directly competing for 3H-cocaine

recognition sites but was allosterically interacting with the cocaine

TABLE 6. Scatchard analysis of 3H-cocaine binding to striatel

membranes in presence or absence of cocaine binding

inhibitor

3H-Cocaine Binding

Bmax

Addition to (pmol/mg

Incubation Mixture (1 ml) KD (mM) protein)

None 0.89 11.7

Cocaine Binding Inhibitor

(30 ug)

0.70 5.6

NOTE: The values represent the mean of two different measurements and were derived

by Scatchard analysis of H-cocaine binding isotherms.

50
binding. To characterize further the molecular nature of the cocaine

binding inhibitor, we determined its amino acid composition after

hydrolysis in constant-boiling HCI at 110 °C for 22 hours. The

protein contained about 126 amino acid residues and had a relative

abundance of Lys and Arg residues. Moreover, the relative number

of the various amino acids residues/mol protein that were obtained

for the cocaine binding inhibitor were very similar to those obtained

for GABAmodulin (Vaccarino et al. 1985), a protein that was shown to

allosterically modulate GABAA receptors (Guidotti et al. 1983). This

similarity prompted us to study the effect of GABAmodulin on Na+-

sensitive 3H-cocaine binding to striatal membranes. The data in table

7 show that GABAmodulin that was isolated from brain synaptosomes

inhibited Na+-dependent 3H-cocaine binding with a potency comparable to that of the cocaine binding inhibitor. In fact, halfmaximal inhibition of 3H-cocaine binding was elicited by 1.7 μM of

TABLE 7. Inhibition of NA+

-dependent 3H-cocaine binding to striatal

membranes by GABAmodulin and a cocaine binding

inhibitory protein

Addition to Incubation 3H-Cocaine Bound

Mixture (1 ml) (pmol/mg protein)

None 0.122

HPLC-Vehicle 0.140

Cocaine Binding

Inhibitor (30 ug) 0.061

GABAmodulin

(30 ug) 0.073

NOTE: The values represent the mean of two experiments in triplicate. GABAmodulin

isolated from rat brain synaptosomes was kindly provided by Dr. A. Guidotti,

FIDIA-Georgetown Institute for Neuroscience, Washington, DC.

GABAmodulin and the same degree of inhibition of 3H-muscimol

binding was achieved by 1.2 μM of the same protein. Considering the

evidence on biochemical and functional similarities and the fact that

51
the isolation procedures were very similar, except that the cocaine

binding inhibitor was extracted from synaptosomes prepared from

caudate nucleus and not whole brain, it appears that GABAmodulin

and the cocaine binding inhibitor are either the same protein or very

similar. Moreover, the noncompetitive nature for the inhibition of

the binding of both radioligands suggests that the synaptosomal basic

protein may act indirectly as an allosteric modulator on recognition

sites for GABA or cocaine. Previous reports in the literature

demonstrated that the reduction in high-affinity blnding of GABA

elicited by this synaptosomal basic protein was abolished if the

protein was phosphoylated in vitro by cyclic AMP-dependent protein

kinase (Wise et al. 1983). These results suggest that phosphorylation

may be a physiological mechanism to regulate the efficacy of the

synaptosomal basic protein. Ongoing work in our laboratory is aimed

at establishing a physiological role of this protein in the regulation

of cocaine receptor sites located at dopaminergic terminals. In

particular, phosphorylation of the synaptosomal basic protein or other

membrane’s proteins may regulate the sensitivity of cocaine recognition sites to ligands.

CONCLUSION

Studies on ion sensitivity revealed the existence of two types of

cocaine binding sites: Na+-sensitive and Na+-insensitive sites. The

Na+-insensitive cocaine binding sites appear to be uniformly distributed in brain, but they occur in low density. The physiological role

of Na+-insensitive cocaine recognition sites and mechanisms by which

such sites are regulated are unknown at the present time. The Na+-

sensitive cocaine binding sites found to be specifically located in

dopaminergic terminals in caudate nucleus were found to be physiologically linked to the dopamine uptake system. The Na+

-sensitive

cocaine binding sites can be allosterically downregulated by a

synaptosomal basic protein, which was found to downregulate also

GABAA binding sites. In addition, the density of Na+-sensitive

cocaine binding sites in caudate nucleus was increased after the

destruction of glutamatergic terminals by cortical ablation. These

results suggest that L-glutamate may function as a negative modulator

of cocaine binding sites located in dopaminergic nerve terminals.

REFERENCES

Baudry, M., and Lynch, G. Characterization of two [

3H]glutamate

binding sites in hippocampal membranes. J Neurochem 36:811-820,

1981.

52
Briley, M., and Langer, S.Z. Sodium dependency of [

3H]imipramine

binding in rat cerebral cortex. Eur J Pharmacol 72:377-380, 1981.

Cotman, C.W.; Monaghan, D.T.; Ottersen, O.P.; and Storm-Mathisen, J.

Anatomical organization of excitatory amino acid receptors and

their pathways. Trends in Neurosciences 10:273-279, 1987.

Donzanti, B.A., and Uretsky, N.Y. Effects of excitatory amino acids

on locomotor activity after bilateral microinjection into the rat

nucleus accumbens: Possible dependence on dopaminergic mechanisms. Neuropharmacology 22:971-981, 1983.

Enna, S.J., and Snyder, S.H. Properties of y-aminobutric acid (GABA)

receptor binding in rat brain synaptic membrane fractions. Brain

Res 100:81-97, 1975.

Guidotti, A.; Konkel, D.R.; Ebstein, B.; Corda, M-G.; Wise, B.C.;

Krutzsch, H.; Meek, J.L; and Costa, E. Isolation, characterization

and purification to homogeneity of a rat brain protein (GABAmodulin). Proc Natl Acad Sci USA 79:6084-6088, 1983.

Hanbauer, I.; Kennedy, LT.; Missale, M.C.; and Bruckwick, E.C.

Cocaine binding sites located in striatel membranes are regulatory

sites for dopaminergic synapses. In: Biggio, G.; Toffano, G.; and

Gessa, G.L., eds. Neuromodulation and Brain Function. Vol. 48.

Oxford: Pergamon Press, 1984.. pp. 41-49.

Kennedy, L.T., and Hanbauer, I. Sodium-sensitive cocaine binding to

striatal membrane: Possible relationship to dopamine uptake sites.

J Neurochem 41:172-178, 1983.

Pasternak, G.W.; Wilson, H.A.; and Snyder, S.H. Differential effects

of protein-modifying regents on receptor binding of opiate agonists

and antagonists. MOl Pharmacol 11:340-351, 1975.

Pimoule, C.; Schoemaker, H.: Javoy-Agid, F.; Scatton, B.; Agid, Y.; and

Langer, S.F. Decrease in 3H-cocaine binding to the dopamine

transporter in Parkinson’s disease. Eur J Pharmacol 95:145-146,

1983.

Rehavi, M.; Skolnick, P.; Brownstein, M.J.; and Paul, S.M. Highaffinity binding of [

3H]desipramine to rat brain: A presynaptic

marker of noradrenergic uptake sites. J Neurochem 38:889-895,

1982.

Reith, M.E.A.; Sershen, H.; and Lajtha, A. Saturable [

3H]cocaine

binding in central nervous system of mouse. Life Sci 27:1055-1062,

1980.

Reith, M.E.A.; Meisler, B.E.; Sershen, H.; and Lajtha, A. Sodiumindependent binding of 3H-cocaine in mouse striatum is serotonin

related. Brain Res 342:145-148, 1985.

Roberts, P.J., and Anderson, S.D. Stimulatory effect of L-glutamate

and related acids on 3H-dopamine release from rat striatum: An in

vitro model for glutamate actions. J Neurochem 32:1539-1545, 1979.

53
Rudolph, M.I.; Argueros, L.; and Bustos, G. L-glutamic acid, a

neuromodulator of dopaminergic transmission in the rat corpus

striatum. Neurochemistry International 5:479-486, 1983.

Sershen, H.; Reith, M.E.A and Lajtha, A. Comparison of the

properties of central and peripheral binding sites for cocaine.

Neuropharmacology 21:469-474, 1982.

Usdin, T.B.; Creese, I.; and Snyder, S.H. Regulation by cations of

[

3H] spiroperidol binding associated with dopamine receptors of rat

brain. J Neurochem 34:669-676, 1980.

Vaccarino, F.; Conti-Tronwni, B.M.; Panula, P.; Guidotti, A.; and

Costa, E. GABA-modulin: A synaptosomal basic protein that

differs from small myelin basic protein of rat brain. J Neurochem

44:278-290, 1985.

Walker, J.E., and Fonnum, F. Effect of regional cortical ablations on

high affinity D-aspartate uptake in striatum olfactory tubercle and

pyriform cortex of the rat. Brain Res 278:283-286, 1983.

Wise, B.C.; Guidotti, A.; and Costa, E. Phosphorylation induces a

decrease in the biological activity of the protein inhibitor (GABAmodulin) of gamma-amino butyric acid binding sites. Proc Natl

Acad Sci USA 80:886-890, 1983.

AUTHOR

lngeborg Hanbauer, Ph.D.

National Heart, Lung, and Blood Institute

National Institutes of Health

Building 10, Room 7N244

Bethesda, MD 20892

54
Sensitization to Cocaine in the

Nigrostriatal Dopamine System

Nancy R. Zahniser, Joanna Peris, Linda P. Dwoskin,

Pamela Curella, Robert P. Yasuda, Laurie O’Keefe, and

Saiiy J. Boyson

INTRODUCTlON

Cocaine is a widely abused drug presumably because of its stimulant

and euphoric properties. Rats given a moderate dose of cocaine show

increased locomotor activity and stereotypic behavior. These

behaviors are thought to be mediated by the mesolimbic and nigrostriatal dopamine (DA) pathways, respectively (Costal and Naylor

1977). Our interest in understanding the mechanisms underlying the

action of cocaine in the brain was sparked by two observations

reported in the literature involving these central dopaminergic

systems. The first observation was that repeated cocaine administration resulted in sensitization of both locomotor activity and

stereotypic behavior (Stripling and Ellenwood 1977; Post and Contel

1983). This is in contrast to repeated administration of other drugs

that result in tolerance, for example, anticholinesterase agents

(Overstreet and Yamamura 1979). Second was the observation that

repeated cocaine administration, like that of other direct or indirect

DA receptor agonists, did not consistently downregulate postsynaptic

striatal DA receptors (Creese and Sibley 1981). This finding is also

in contrast to other receptors such as brain muscarinic cholinergic

receptors that show consistent downregulation in response to chronic

agonist stimulation induced by anticholinesterase drugs (Overstreet

and Yamamura 1979). Thus, repeated activation of DA pathways by

cocaine may actually result in increased activity of dopaminergic

systems leading to behavioral sensitization in response to subsequent

cocaine administration.

A similar hypothesis has been proposed based on the results of

studies demonstrating behavioral sensitization after repeated amphetamine treatment. Robinson and Becker (1986) have suggested that

55
development of behavioral sensitization to amphetamine in the

nigrostriatal DA pathway is not due to an increase in DA synthesis

or postsynaptic DA receptors, but rather to enhanced release of DA

upon reexposure to amphetamine. They also concluded that enhanced

DA release after repeated amphetamine treatment did not appear to

be due to DA autoreceptor subsensitivity. It is possible that a

similar augmentation of DA release underlies cocaine-induced sensitization involving the nigrostriatal DA system. To address this

question, we measured stereotypic behavior and neurochemical

parameters of DA activity in the nigrostriatal pathway in rats treated

with either single or repeated injections of cocaine. The results from

these studies, as well as those in the literature, will be discussed in

this chapter.

When reviewing the literature, it is essential to distinguish between

changes in parameters in both cell body and terminal regions and

changes in pre- and postsynaptic receptors because it is probable that

these are regulated in different ways. For example, the striatum, or

terminal region of the nigrostriatal tract, has been the focus of the

majority of the studies. It has been suggested, however, that regions

containing the cell bodies such as the substantia nigra may be more

important in the development of sensitization than terminal regions.

Behavioral sensitization to cocaine occurs only when repeated

injections are made directly into substantia nigra, as opposed to

striatum (Kalivas and Weber 1987). It is also likely that pre- and

postsynaptic DA receptors would be regulated in a different fashion

by repeated exposure to cocaine. Increased dopaminergic activity

could be explained either by decreased activation of DA autoreceptors

on either cell bodies or terminals or by increased activation of

postsynaptic DA receptors. On the other hand, in order to compensate for higher levels of synaptic DA, autoreceptor activity would be

expected to increase, thereby more efficiently inhibiting firing or

release, and the density, affinity, or coupling of postsynaptic

receptors would be expected to decrease. As mentioned previously,

however, DA receptors, unlike other neurotransmitter receptors, do

not always downregulate in response to chronic agonist exposure.

These points will be considered below.

It is important to consider cocaine-induced changes in DA neurotransmission in the mesolimbic system in addition to those in the

nigrostriatal system. Sensitization occurs to the locomotor-activating

effect of cocaine, and it is known that this behavior is mediated by

the mesolimbic system. However, fewer neurochemical studies have

focused on this system, perhaps because of problems in working with

56
smaller, more diffuse brain areas or because of the more complicated

behavioral changes mediated by this system. The euphoric effects of

cocaine, as well as the locomotor-activating effects, are mediated by

the mesoiimbic system. Tolerance has been reported to occur to the

euphoric effects with repeated administration (Fischman et al. 1985).

Thus, the mesolimbic system represents a more complex model in

which to investigate dopaminergic changes underlying sensitization to

cocaine. For this reason, we have focused our own work and this

chapter on the nigrostriatal system but discuss the literature pertaining to the mesolimblc system as a future direction for cocaine

research.

SINGLE-DOSE ADMINISTRATlON OF COCAINE

Sensitization of DA-mediated behaviors has been shown to occur

after only one injection of either amphetamine (Robinson et al. 1982)

or cocaine (Lin-Chu et al. 1985). Thus, one would expect that

regulation of DA-containing systems would occur in response to a

single dose of cocaine and, additionally, would begin in response to

the first dose of cocaine in a series of repeated exposures. When

investigating the effects of repeated administration of cocaine, we

measured neurochemical parameters 24 hours after the last cocaine

injection. Therefore, initially, we measured the effect of a single

moderate dose of cocaine (10 mg/kg, intraperitoneally) injected 24

hours prior to sacrifice on several parameters of DA transmission.

These parameters included tritium release elicited by either electrical

stimulation or brief exposure to amphetamine from striatal slices

preloaded with 3H-DA, the ability of D-2 DA autoreceptors to

modulate the electrically evoked release, and the number and affinity

of postsynaptic striatal D-2 DA receptors. When a change was found,

the duration of the change following a single injection was

investigated.

Presynaptic Parameters

DA release. As already mentioned, studies of behavioral sensitization

to amphetamine have supported the hypothesis that increased activity

of central DA systems underlies the enhanced responsiveness to

subsequent drug administration. Sensitization of both stereotypic and

locomotor behavior and of amphetamine-induced release of striatai DA

has been shown to develop in response to a single injection of

amphetamine, and these investigators have concluded that sensitization to amphetamine in the nigrostriatal tract is predominantly due to

57
this increased DA release (Robinson and Becker 1986). The augmented behavioral responsiveness and neurotransmitter release are

present 24 hours after acute exposure and have been shown to persist

for at least 2 weeks (Robinson et al. 1982).

Likewise, both behavioral sensitization of the nigrostriatal DA system

(Lin-Chu et al. 1985) and increased amphetamine-induced 3H-DA

release from striatal slices (Peris and Zahniser 1987) occur after a

single dose of cocaine. The increase in amphetamine-stimulated

release of 3H-DA is present from 24 hours through 2 weeks after

cocaine injection but returns to control levels after 1 month.

Tritium overflow evoked by 300 pulses of electrical stimulation of

slices from animals injected once with cocaine is identical in cocaineand saline-injected animals at all of these timepoints (Peris and

Zahniser 1987). lt should be noted that amphetamine causes tritium

ovefflow from tissue preloaded with 3H-DA that consists primarily of

3H-DA (Parker and Cubeddu 1986b), while electrical stimulation of

such tissue in the absence of uptake blockers and monoamine oxidase

inhibitors causes tritium overflow that consists primarily of 3Hdihydroxyphenylacetic acid (

3H-DOPAC) (Parker and Cubeddu 1985).

Thus, we will refer to 3H-DA overflow in response to amphetamine

stimulation but tritium overflow in response to electrical stimulation.

Since amphetamine and electrical stimulation do not evoke release

from the same pools (Niddam et al. 1985; Parker and Cubeddu 1986a;

Parker and Cubeddu 1986b), the results reported by Peris and

Zahniser (1987) suggest a redistribution of DA in the behaviorally

sensitized animals into the pool(s) susceptible to release by amphetamine. Although amphetamine and cocaine may produce their in

vivo effects on DA nerve terminals primarily via different mechanisms, i.e., amphetamine via promotion of release and cocaine via

inhibition of uptake, the resulting increase in synaptic levels of DA

may ultimately result in the same regulatory changes being induced in

the intraneuronal pools of DA and receptor and/or transporter

proteins. Thus, as Robinson and Becker (1986) have suggested for

amphetamine-induced sensitization, the increase in DA-mediated

behaviors induced by previous exposure to cocaine may be related to

increased striatal DA release in these animals.

Cocaine has several different pharmacological actions in the central

nervous system. lt is an inhibitor of DA uptake, an indirect-acting

DA receptor agonist, and a local anesthetic. In order to determine

which of these actions is responsible for the long-lasting cocaineinduced increase in amphetamine-stimulated 3H-DA release previously

observed (Peris and Zahniser 1987). release was measured either 1 day

58
or 1 week after animals received a single injection of various

substances sharing one or more of these properties of cocaine.

Substances tested were mazindol, a DA uptake inhibitor and indirectacting DA receptor agonist with no known local anesthetic properties;

apomotphine, a direct-acting DA receptor agonist; and lidocaine, a

local anesthetic. Additionally, in order to evaluate the contribution

of the indirect receptor agonist activity of cocaine, the effects

produced by a single injection of cocaine were measured after

pretreatment with DA receptor antagonists.

The methods for measuring “H-DA release from rat striatal slices

have been described previously (Peris and Zahniser 1987). A single

injection of cocaine augments amphetamine-stimulated DA release

24 hours after the injection (Peris and Zahniser 1987). We measured

the effects of mazindol(1 or 10 mg/kg), apomorphine (0.5 or 5

mg/kg), or lidocaine (20 mg/kg) on DA release evoked by 2.5-minute

superfusion with 6 μM amphetamine 24 hours after a single injection.

It should be kept in mind that many of these drugs have longer halflives than cocaine; therefore, the results 1 day after injection may

have been confounded. For example, the higher dose (10 mg/kg) of

mazindol decreased amphetamine-stimulated release at 24 hours and

increased electrically stimulated release, indicating that mazindol was

still present in the striatal slices during the release experiments.

Neither lidocaine nor apomorphine was effective in augmenting release

after 24 hours; however, pretreatment with DA receptor antagonists

abolished the effect of cocaine at 24 hours. These data suggest that

the indirect agonist effect of cocaine is necessary for the augmentation of release, although one would have expected apomorphine to

augment release in a manner similar to cocaine. In any case, the

effect of a single injection of cocaine that remains 24 hours later

should be taken into account in interpreting data from repeated

cocaine administration studies in which animals were sacrificed 24

hours after the last dose of cocaine.

The effects of a single injection of cocaine and the drugs mentioned

above were also examined 1 week after the drugs were injected

(figure 1). As previously reported (Peris and Zahniser 1987), in vivo

pretreatment with cocaine produced a long-lasting 25 percent increase

in amphetamine-stimulated release. In comparison, pretreatment with

lidocaine did not increase release significantly, whereas pretreatment

with apomorphine or mazindol increased release to a similar extent as

the pretreatment with cocaine. The results of the experiments

examining the effects of the pretreatment with DA receptor

antagonists are also shown in figure 1. When the nonselective DA

59
In Vivo Drug Treatment

FIGURE 1. D-amphetamine-induced 3H-DA release is augmented

1 week after a single cocaine or apomorphine injection;

this augmentation is blocked by DA receptor antagonists

NOTE: Rats were injected with the drugs indicated and sacrificed 1 week later.

H-DA release was measured from striatal slices in response to exposure to

6 μM amphetamine. n=7 (saline), n=5 (mazindol, apomorphine, lidocaine). n=6

(all other treatments).

receptor antagonist fluphenazine (5 mg/kg) was injected 15 minutes

prior to the cocaine administration, the long-lasting potentiation of

amphetamine-induced 3H-DA release produced by cocaine was

abolished. Pretreatment with either the selective D-1 DA receptor

antagonist SCH 23390 (0.5 mg/kg) or the selective D-2 DA receptor

antagonist sulpirfde (100 mg/kg) was equally effective in blocking the

effects of cocaine on amphetamine-stimulated release. Similar results

were found when 3H-DA release was evoked by superfusion with

either 2 or 20 μM amphetamine (data not shown). None of the

antagonists had an effect on release when injected without cocaine.

Furthermore, pretreatment with any of the drugs did not affect

tritium ovefflow evoked by electrical field stimulation (5 Hz,

1 minute, 300 pulses) from these slices in the same experiments (data

not shown).

60
These results suggest that the indirect agonist effects of cocaine are

involved in producing the enhanced amphetamine-evoked release

observed after a single dose of cocaine. Apomorphine, a direct-acting

DA receptor agonist, mimics the effects of cocaine, and DA receptor

antagonists block the effects of cocaine. As expected, mazindol

produces the same effects as cocaine because mazindol is also an

indirect DA receptor agonist. However, one difference between

cocaine, apomorphine, and mazindol is their duration of action in the

brain. Cocaine and apomorphine are essentially completely cleared

from brain within 60 minutes (Nayak et al. 1975; Schwartz et al.

1978), while mazindol has a much longer half-life in plasma of 30

hours (Shindler et al. 1985). lt is likely that the effects of these

drugs at 24 hours after injection differ because of the different halflives. These drugs also differ in their effects on different amine

neurotransmitters. Cocaine inhibits uptake with the following order

of potencies: serotonin > DA > norepinephrlne, while the order of

potencies for mazindol is norepinephrine > DA = serotonin (Ritz et al.

1987). Apomorphine, on the other hand, is a selective agonist for DA

receptors. These differences will be explored in future experiments.

Additionally, it appears that activation of both D-1 and D-2 DA

receptors is necessary for cocaine-induced sensitization to occur.

Along these same lines, apomorphine activation of locomotor behavior

can be blocked by either D-1 or D-2 receptor antagonists (Schwartz

et al. 1978, Herrera-Marschitz and Ungerstedt 1985; Amalric et al.

1986; Vaccheri et al. 1986). As has been suggested recently for a

number of DA receptor-mediated behaviors (Gershanik et al. 1983;

Braun and Chase 1986; Amt et al. 1987; Carlson et al. 1987; Wachtel

et al. 1987; Walters et al. 1987). D-1 receptors play an additive, a

permissive, or even an obligatory role in D-2 receptor-mediated

events.

lf the indirect-acting agonist effects of cocaine underlie the sensitization to amphetamine-evoked release, then these effects may also be

the basis for the behavioral sensitization seen in response to a single

injection of cocaine. lt would be of interest to measure whether

stereotypic behavioral sensitization develops to cocaine if rats are

pretreated with selective D-1 and D-2 DA receptor antagonists prior

to the initial dose of cocaine. Previous attempts to block behavioral

sensitization to repeated cocaine injections with nonselective DA

receptor antagonists have resulted in either complete (Borison et al.

1979; Beninger and Herz 1988) or partial blockade (Gale 1984).

DA Uptake, Synthesis, and Metabolism. Despite the fact that many

of the effects of cocaine can be explained by its ability to block

61
reuptake of DA into the nerve terminal, we have found no changes in

either the affinity or the maximal velocity of neuronal 3H-DA uptake

into striatal slices 24 hours after pretreatment with a single

10 mg/kg injection of cocaine (Zahniser et al. 1986). 3H-nomifensine

can be used to label specific recognition binding sites for the DA

uptake pump (Dubocovich and Zahniser 1985). Saturation analysis of

specific 3H-nomifensine binding to striatal membranes of rats treated

24 hours earlier with cocaine also revealed no differences when

compared with controls (Zahniser et al. 1986). Thus, no persistent

changes appear to be induced in the neuronal DA uptake pump by a

single injection of cocaine. When neuronal 3H-DA uptake is measured

in vitro 20 minutes after cocaine is administered in vivo, there is, as

expected, a dose-related decrease in striatal DA uptake (Missale et al.

1985). However, by 60 minutes after cocaine administration, uptake

returns to control levels (Missale et al. 1985). These results are

consistent with the 20-minute half-life of cocaine in the brain (Nayak

et al. 1975).

Tissue levels of DA are not changed in the striatum between 3 and

24 hours after a single injection of cocaine (Taylor and Ho 1977;

Zahniser et al. 1986; Hanson et al. 1987), indicating that cocaine is

probably not having a neurotoxic effect. Cocaine increases striatal

tyrosine hydroxylase activity up to 40 minutes after cocaine injection

(Pradhan 1983) but has no effect 3 hours after injection (Hanson

et al. 1987). The level of DOPAC, one of the major metabolites of

DA, decreases in the striatum 60 minutes after an acute injection of

cocaine (Kalivas et al. 1988). The levels of the other major

metabolite, homovanillic acid, or HVA, are not altered in striatum 30

minutes after a single cocaine injection (Taylor and Ho 1977) but

have been reported to increase 3 hours after injection (Hanson et al.

1987). Thus, the short-term effects of a single injection of cocaine

on DA synthesis and metabolism are complex, but there appears to be

an overall decrease in both intraneuronal and extraneuronal DA

metabolism (Bagchi 1986). In general, an increase in synthesis and

decrease in metabolism caused by cocaine should result in an overall

transient increase in synaptic concentrations of DA after a single

injection of cocaine. However, this effect does not persist long

enough to make it a good candidate to explain sensitization to

cocaine.

D-2 DA Autoreceptors D-2 DA receptors localized on DA neurons

are termed autoreceptors. The following three types of DA autoreceptors have been proposed to be associated with dopaminergic

nigrostriatal neurons: impulse-regulating autoreceptors associated

62
with cell bodies and synthesis- and release-modulating autoreceptors

localized on the nerve terminals (Jackisch et al. 1980; Bannon and

Roth 1988). The activity of the first two types of nigrostriatal DA

autoreceptors has not been reported following a single injection of

cocaine. The activity of striatal D-2 release-modulating autoreceptors

can be measured using an in vitro superfusion system, by determining

the ability of the D-2 receptor agonist pergolide to inhibit the

tritium overflow evoked by electrical field stimulation from striatal

slices preloaded with 3H-DA (Dwoskln et al. 1988). These receptors

must be studied with a functional assay because they have not been

detected using radioligand binding assays either with striatal membranes and conventional filtration assays (Zahniser et al. 1986) or

with striatal sections and quantitative autoradiography (Filloux et al.

1988). The activity of these striatal D-2 release-modulating autoreceptors is not different when measured 24 hours after one injection

of cocaine (Dwoskin et al. 1988). Therefore, it is unlikely that

changes in these D-2 release-modulating autoreceptors could explain

the increased behavioral sensitivity to cocaine seen after one

injection (Lin-Chu et al. 1985). Indeed, of all the presynaptic

parameters investigated, only increases in amphetamine-induced

release of DA and possible increases in synthesis of DA coupled with

decreases in metabolism could underlie behavioral sensitization.

Postsynaptic Parameters

D-1 and D-2 DA Receptors. Only a few studies have measured the

effect of one cocaine injection on DA receptor binding properties.

An increased density of D-2 DA receptors in striatum with no effect

on receptor affinity has been observed 60 minutes after a single

injection of cocaine (Memo et al. 1981). However, when striatal D-2

receptors were measured 24 hours after one injection of cocaine

(10 mg/kg), no differences were seen in either receptor density or

affinity (Dwoskin et al. 1988). Thus, once again it appears as if

there is no long-lasting change in postsynaptic striatal D-2 DA

receptors that could explain the persistent behavioral changes seen

after a single exposure to cocaine. To our knowledge, the effects of

a single injection of cocaine on striatal D-1 DA receptors have not

been examined.

Second Messengers. DA acting at postsynaptic DA receptors in the

striatum exhibits dual regulation of adenylate cyclase activity.

Activation of D-1 DA receptors stimulates cAMP production, while

activation of D-2 receptors inhibits CAMP formation (Stoof and

Kebabian 1981; Cooper et al. 1986). DA exhibits a decreased potency

63
in stimulating striatal CAMP formation 60 minutes after one cocaine

injection (Memo et al. 1981). It is interesting to note that acute

administration of amphetamine promotes a rapid, transient desensitization of sttiatal DA-stimulated adenylate cyclase activity (Bamett and

Kuczenskl 1986). These findings may be interpreted as either a

decrease in D-1 receptor-mediated stimulation of adenylate cyclase

activity or an increase in D-2 receptor-mediated inhibition. Longlasting effects of acute exposure to cocaine on striatal adenylate

activity have not yet been measured.

REPEATED ADMINlSlRATlON OF COCAINE

Behavioral sensitization to the locomotor activating and stereotypic

effects of repeated cocaine administration is well-documented in rats

(Stripling and Ellenwood 1977). The degree of sensitization is also

augmented with repeated exposure to cocaine, and this expression of

sensitization may therefore involve mechanisms additional to those

involved following a single injection of cocaine. More studies have

focused on measuring parameters that may explain behavioral

sensitization following repeated, as opposed to single dose, administration of cocaine. Since activation of the nigrostriatal

dopaminergic pathway is implicated in these stereotypic behaviors, it

has again been reasoned, as in the single-dose administration studies,

that there may be a concomitant change in either pre- or postsynaptic parameters of DA neurotransmission in this pathway leading to

greater activation and explaining sensitization.

Presynaptic Parameters

DA Release. Based on our findings of a long-lasting increase in

amphetamine-induced release of 3H-DA from striatal slices of rats

treated with only one dose of cocaine (10 mg/kg), we expected to

find an even greater augmentation following repeated administration

of cocaine. In contrast to the acute injection results, however,

following either 8 or 14 once-daily injections of 10 mg/kg cocaine,

amphetamine-induced 3H-DA release was not different from salinetreated animals (unpublished observations). This finding was puzzling

because our previous results showed that release was increased for 2

weeks following a single injection of cocaine. Thus, in these

experiments, we expected that release would be increased in response

to the last injection of cocaine for the same duration. However,

recent data indicate that if animals are sacrificed 72 hours after the

last dose of cocaine, then amphetamine-stimulated 3H-DA release is

increased twofold over water-injected controls (Johnson and Snell

64
1987). Thus, it appears that a compensatory change that obscures

the increase in amphetamine-induced release occurs after repeated

cocaine injections, but that this compensatory mechanism disappears

within a few days of terminating the cocaine administration. One

possible compensatory change involves the D-2 release-modulating

autoreceptors in the striatum. lt is controversial whether D-2

autoreceptors are capable of modulating amphetamine-induced release

(Kamal et al. 1981; Kelly and Nahorski 1987). We have preliminary

evidence that the D-2 receptor antagonist S-sulpiride increases 3H-DA

release evoked by low concentrations of amphetamine but not that

evoked by high concentrations. If the activity and/or density of

these autoreceptors were increased in response to the repeated

cocaine injections (as discussed below), then these receptors could

inhibit DA release to a greater extent in striatum of cocaine-treated

animals. This hypothesis could be tested by determining whether

blockade of D-2 autoreceptors during measurement of amphetamine

induced 3H-DA release from cocaine-sensitized animals reinstates the

augmented release seen after one injection of cocaine. When

augmented amphetamine-evoked DA release was seen after repeated

amphetamine administration, measurements of release occurred 10 days

after the last amphetamine exposure, thereby allowing some degree of

withdrawal to take place (Robinson and Becker 1986). lt is possible

that if release were measured 24 hours after the last amphetamine

injection, augmentation of release would not have occurred.

Dendrites release DA, which can interact with impulse-regulating

autoreceptors in the cell body region. When rats were injected with

cocaine (15 or 45 mg/kg) for 3 days and then withdrawn for 2 to 3

weeks, a decrease in both amphetamine- and K+-stimulated release of

endogenous DA was observed in ventromedial mesencephalon, the

region containing the cell bodies of the nigrostriatal, mesolimbic, and

mesocortical DA-containing pathways (Kalivas and Duffy 1988). This

decreased somatodendritic release could result in less autoreceptor

inhibition at the cell body, thereby increasing the firing of the

nigrostriatal tract and DA release from the terminals. In fact,

Kalivas and Duffy observed an increased release in striatum only

after the animals had been withdrawn from the higher dose of

cocaine. This alteration is consistent with sensitization and stresses

the importance of cocainsinduced changes that may occur at the cell

body. It is also interesting to note that changes that occur that are

consistent with the development of behavioral sensitization are longlasting and may become apparent only after a certain drug-free

period.

65
DA Uptake, Synthesis, and Metabolism. No changes in the neuronal

uptake of 3H-DA in the striatum were observed 20 minutes after

withdrawal from repeated cocaine administration (Missale et al. 1985).

However, acute cocaine decreases the Vmax of 3H-DA uptake in

striatum; therefore, repeated administration can be said to result in

the development of tolerance to the ability of cocaine to block

uptake (Missale et al. 1985). Consistent with the findings of this

group, we also observed no differences in the properties of either

high-affinity 3H-DA uptake or 3H-nomifensine binding in striatal

tissue from behaviorally sensitized animals that had been treated for

8 or 14 days with cocaine and then withdrawn for 24 hours (Zahniser

et al. 1986). Taken together, these results suggest that in relation to

the uptake blockade produced by cocaine in the striatum, there may

be less of an increase in synaptic DA levels produced in response to

repeated cocaine exposure. This finding is consistent with the

development of tolerance rather than sensitization.

Striatal DA levels are the same as those in control rats after a 24

hour withdrawal from repeated cocaine treatment (Zahniser et al.

1986; Yeh 1987). This indicates that repeated administration of

moderate doses of cocaine does not produce DA neurotoxictty.

Following 9 days of repeated cocaine injections, there is an increase

in DA content in both terminal and cell body regions when DA is

measured 20 minutes, but not 60 minutes, after the last cocaine

injection (Roy et al. 1978). This increase is no longer present on the

18th or 30th day of repeated cocaine injections. Chronic cocaine

decreases tyrosine hydroxylase immunoreactivity in both nigrostriatal

cell body and terminal regions (Trulson et al. 1986). However, there

has also been a report of increased striatal tyrosine hydroxylase

activity after repeated cocaine injections (Taylor and Ho 1977). In

the striatum, a persistent decrease in DA metabolism following

repeated injections of cocaine (Karoum et al. 1987), a slight increase

(Taylor and Ho 1977), or no change (Yeh 1987) have been reported.

In summary, no consensus exists as to long-lasting changes in DA

content, synthesis, or metabolism produced by repeated administration

of cocaine.

D-2 DA Autoreoeptors. As discussed previously, D-2 DA autoreceptors on terminals of the nigrostriatal tract modulate the

stimulation-evoked release of DA. These autoreceptors appear to

comprise an insignificant number of the D-2 receptors measured with

in vitro radioligand binding assays. When these receptors were

measured functionally, they showed a small but consistent increased

sensitivity 24 hours after the 8- or 14-day cocaine treatment

66
(Dwoskin et al. 1988). These data are in contrast to the classical

downregulation or decreased sensitivity observed for many postsynaptic receptors after prolonged agonist stimulation (Creese and

Sibley 1981). An increased sensitivity of these receptors would

inhibit stimulation-evoked release of DA to a greater extent and

would oppose the increased wncentrations of synaptic DA produced in

response to cocaine. This change would also oppose increased DA

release in response to decreased impulse-regulating autoreceptor

activation and the component of release that is represented by

increased amphetamine-induced release. Therefore, this appears to be

a compensatory response consistent with the development of

tolerance.

In contrast with D-2 DA receptors in the terminal fields, those in the

cell body region, i.e., the substantia nigra compacta, are all impulseregulating autoreceptors (Filloux et al. 1988). Therefore, radioligand

binding assays can be used to quantitate D-2 autoreceptors in this

area. We have recently used quantitative autoradiographic analysis to

examine specific binding of 3H-spiperone to D-2 receptors in the

substantia nigra compacta and the substantia nigra reticulata of rats

treated with cocaine for 8 days and withdrawn for 24 hours. The

binding to a single high concentration of 3H-spiperone was significantly increased in the caudal half of the substantia nigra

compacta but was not changed in the substantia nigra reticulata (data

not shown). lf this increase in D-2 autoreceptors in the substantia

nigra is functionally significant, then this cocaine-induced change,

like that of the release-modulating autoreceptors in the striatum,

would also be consistent with dampened dopaminergic activity and the

development of tolerance to the stimulating effects of cocaine. lt

should be noted that the decreased somatodendritic release reported

by Kalivas and Duffy (1988) could be the result of the increased D-2

autoreceptors in this area. When the data are considered in this

way, they are more consistent with tolerance rather than sensitization.

Postsynaptic Parameters

D-1 and D-2 DA Receptors. Most of the studies concerned with DA

receptor regulation after repeated cocaine administration have

measured parameters of D-2 receptor binding. Until recently, it was

generally accepted that D-2 receptors were functionally more

important than D-1 receptors (Seeman 1980). Despite several studies,

however, no consensus has been reached as to the modulation of D-2

67
receptors by repeated cocaine administration. Initial studies reported

dose-related increases in striatal D-2 DA receptor number (Borison

et al. 1979; Taylor et al. 1979). More recently, Goeders and Kuhar

(1987), using quantitative autoradiography, found that repeated

cocaine administration for 15 days resulted in a decrease in the

density of striatal D-2 receptors. Using either membrane binding

assays (Dwoskin et al. 1988) or quantitative autoradiography (unpub

lished observations), we found no differences in the affinlty or

density of these receptors in the striatum of rats treated for 8 or 14

days with once-daily injections of cocaine. Goeders and Kuhar (1987)

sacrificed the rats 20 minutes after the last dose as compared to 24

hours or more in all the other studies. At 20 minutes, the tissue

concentration of cocaine would still be relatively high, whereas by 24

hours it would be negligible. Thus, the difference in the time when

the tissue was taken may explain some of the differences in the

results of these studies.

The fact that we found no significant change in the density of

striatal D-2 receptors may seem at odds with our functional data

showing increased inhibition by D-2 autoreceptors in striatum after

repeated cocaine treatment. The discrepant results are not explained

by the fact that the binding studies were carried out using an antagonist, while the functional studies were carried out using an agonist.

Competition curves for the agonist pergolide revealed similar binding

affinities in the control and cocaine-treated rat striatal membranes

(Dwoskin et al. 1988). Instead, these data support the idea that the

number of autoreceptors may be a relatively small proportion of the

total D-2 receptors in striatum and that for this reason the autoreceptors may not be detected by radioligand binding assays even

with the added sensitivity of quantitative autoradiography.

In adjacent tissue sections in the quantitative autoradiographic

studies, we used 3H-SCH 23390 to label D-1 DA receptors. In the

striatum, the D-1 receptors are thought to be localized postsynaptically, whereas in the substantia nigra reticulata the D-1 receptors

appear to be localized on the striatonigral terminals (Altar and

Hauser 1987). No changes in D-1 receptors were found in either area

when the values from the control and cocaine-treated rats were

compared (data not shown).

Second Messengers. It has recently been reported that the potency

of DA to stimulate adenylate cyclase activity in the striatum

decreases after repeated amphetamine exposure; this desensitization

occurs concomitantly with behavioral sensitization (Barnett et al.

68
1987). Repeated treatment with the DA uptake blocker nomifensine

also decreases DA-stimulated CAMP formation (Algeri et al. 1980).

Chronic treatment with DA receptor agonists L-DOPA, pinbedil, or

bromocriptine also abolishes DA-stimulated cAMP formation in striatal

slices (Mishra et al. 1978). These effects on adenylate cyclase are

presumably mediated via postsynaptic D-1 DA receptors and are

consistent with the development of tolerance. As yet, no one has

measured the effect of repeated cocaine administration on the

regulation of adenylate cyclase by either D-1 or D-2 receptors in the

striatum.

FUTURE DIRECTIONS

As mentioned in the introduction, sensitization also occurs to the

locomotor-activating effect of cocaine, a behavior that is mediated by

the mesolimbic DA pathway. lt is therefore possible that similar

changes in DA function occur both in this pathway and in the nigrostriatal pathway, which mediates behavioral sensitization to the

stereotypic effects of cocaine. However, changes in the mesolimbic

system may be more complex because the mesolimbic, as well as the

mesocortical, dopaminergic projections are implicated in the

reinforcing or euphoric effects of cocaine (Wise 1981; Wise 1984) and

tolerance, rather than sensitization, occurs to the euphoric effect of

cocaine (Fischman et al. 1985). We will now discuss the observations

made concerning cocaine-induced changes in the mesolimbic system as

they pertain to the future directions of research in the cocaine field.

lt is clear that at least some of the effects of cocaine differ

depending on whether measurements are made in the mesolimbic or

nigrostriatal pathways. Application of cocaine inhibits spontaneous

firing of DA neurons in the ventral tegmental area both more

potently and more effectively than it does in substantia nigra

compacta (Pitts and Marwah 1987), supporting the hypothesis that cell

bodies in mesolimbic and nigrostriatal DA systems are affected

differently by cocaine exposure. In vitro cocaine inhibits DA uptake

to a greater extent in striatum than in other DA terminal areas such

as prefrontal cortex (Hadfield and Nugent 1983) and nucleus accumbens (Missale et al. 1985). Similar effects are observed transiently in

the striatum and prefrontal cortex after in vivo administration of

cocaine; paradoxically, in vivo pretreatment with cocaine transiently

increases uptake into nucleus accumbens (Missale et al. 1985).

Chronic cocaine administration increases the maximal velocity of

3H-DA uptake into nucleus accumbens, whereas no change occurs in

striatum (Missale et al. 1985). These same investigators also found

69
that 3H-cocaine binding in nucleus accumbens is sodium ionindependent unlike that found in striatum, which is sodium iondependent. Thus, cocaine binding sites and/or coupling of these sites

to the DA transporter may differ between the mesolimbic and nigro

striatal pathways.

Differences are also evident in the receptor regulation induced by

cocaine. Unlike the nigrostriatal system, in which both the cell body

and terminal D2 autoreceptors change in a manner consistent with

increased inhibitory activity and, therefore, tolerance following

repeated cocaine administration, D-2 autoreceptors in the mesolimbic

system change in the opposite direction. Consistent with sensitization, D-2 autoreceptors in the ventral tegmental area are subsensitive

after chronic cocaine treatment (Greene and White 1986; Henry et al.

1987). Furthermore, postsynaptic D-2 receptors in nucleus accumbens

are more sensitive to agonists after repeated cocaine exposure (Henry

et al. 1987). This may be explained by the fact that after repeated

cocaine injections, an increased density of D-2 DA receptors in

nucleus accumbens has been found in two studies using quantitative

autoradiographic analysis (Goeders and Kuhar 1987; unpublished

observations). Thus, both pre- and postsynaptic D-2 receptors

associated with the mesolimbic tract appear to be regulated differently than those in the nigrostriatal tract. Another difference

between these two systems that has recently been reported suggests

that cocaine might not be expected to have the same postsynaptic

effects in the mesolimbic and nigrostriatal pathways. D-2 DA

receptor-mediated inhibition of adenylate cyclase activity occurs in

striatum and not in nucleus accumbens (Stoof and Verheijden 1986).

On the other hand, some of the effects of exposure to cocaine are

similar in the nigrostrfatal and mesolimbic pathways. For example,

after a single injection of cocaine, DA levels remain stable in both

striatum and nucleus accumbens (Henry et al. 1987). Chronic cocaine

also decreases tyrosine hydroxylase immunoreactivity in terminal

regions of the mesolimbic system (Babb et al. 1986; Trulson et al.

1986). Similar to the nigrostriatal pathway, cell body regions appear

more important in the action of cocaine because behavioral sensitization occurs only if cocaine or amphetamine is injected into the

ventral tegmental area, as opposed to the nucleus accumbens (Kalivas

and Weber 1987). One other similarity is that following repeated

administration of cocaine, D-1 receptors are not regulated in either

the nigrostriatal or mesolimbic systems (unpublished observations).

70
All of these observations considered together suggest that activity in

both the cell body and terminal regions of the mesolimbic and

nigrostriatal pathways are modulated somewhat differently by repeated

cocaine administration. Perhaps most striking are the differences in

the regulation of D-2 receptors in the two pathways. In general, the

receptor changes in the nigrostriatal pathway are consistent with

tolerance, whereas those in the mesolimbic pathway are consistent

with sensitization. lt is intriguing that the majority of the changes

observed in the mesolimbic pathway could be the basis for behavioral

sensitization, since both sensitization and tolerance have been

reported to occur for behaviors mediated by this pathway.

SUMMARY

Behavioral sensitization involving the nigrostriatal dopaminergic tract

is manifested after treatment with only a single dose of cocaine and

is augmented following repeated treatment. One neurochemical

change observed that is consistent with behavioral sensitization is the

increase in amphetamine-induced 3H-DA release from striatal slices

seen after one injection of cocaine. One day after repeated administration of cocaine, however, the increase is no longer evident. lt is

possible that transient compensatory changes, such as increased D-2

autoreceptor inhibition, may obscure this effect when it is measured

at relatively short times after the repeated administration has been

terminated. One day after cessation of repeated cocaine administration, D-2 autoreceptors in both striatum and substantia nigra

compacta were upregulated consistent with a compensatory mechanism

and the development of behavioral tolerance rather than sensitization.

In contrast, DA content, neuronal DA uptake, and postsynaptic D-2

DA receptors in striatum were not regulated by this treatment.

Likewise, D-1 DA receptors in striatum and substantia nigra were

unaffected. In the mesolimbic system, both the pre- and postsynaptic

receptor changes are consistent with sensitization. Amphetaminestimulated release from nucleus accumbens has not yet been measured

in cocaine-sensitized animals. lt is possible that changes similar to

those seen in striatum may occur in this area. It is interesting that,

in general, presynaptic parameters associated with the DA neuron,

with the notable exception of the uptake pump, appear to be more

sensitive to regulation by cocaine administration than do postsynaptic

parameters. The long-lasting effects of a single moderate dose of

cocaine are also surprising. It will be important to determine the

molecular mechanisms underlying this regulation and whether or not

similar changes are induced in mesolimbic dopaminergic systems by

single and repeated administration of cocaine.

71
REFERENCES

Algeri, S.; Brunello, N.; and Vantini, G. Different adaptive responses

by rat striatal dopamine synthetic and receptor mechanisms after

repeated treatment with d-amphetamine, methylphenidate and

nomifensine. Pharmacol Res Commun 12:675-681,1980.

Altar, C.A., and Hauser, K. Topography of substantia nigra innervation by D1

 receptor-containing striatal neurons. Brain Res 410:1-

11, 1987.

Amalric, M.; Koob, G.F.; Creese, I.; and Swerdlow, N.R. “Selective”

D-1 and D-2 receptor antagonists fail to differentially alter

supersensitive locomotor behavior in the rat. Life Sci 39:1985-

1993, 1986.

Art, J.; Hyttel, J.; and Perregaard, J. Dopamine D-1 receptor

agonists combined with the selective D-2 agonist quinpirole

facilitate the expression of oral stereotyped behaviour in rats. Eur

J Pharmacol 33:137-145, 1987.

Babb, S.; Joe, J.C.; Raese, J.D.; and Trulson, M.E. Chronic cocaine

administration depletes tyrosine hydroxylase immunoreactivity in the

nigrostriatal and mesolimbic dopamine systems in the rat brain.

Abstr Soc Neurosci 12:141, 1986.

Bagchi, S.P. Multiple actions of cocaine with opposing effects on

brain 3, 4-dihydroxyphenylacetic acid level. Abstr Soc Neurosci

12:141, 1986.

Bannon, M.J., and Roth, R.H. Pharmacology of mesocortical dopamine

neurons. Pharmacol Rev 35:53-68, 1983.

Bamett, J.V., and Kuczenski, R. Desensitization of rat striatal

dopamine-stimulated adenylate cyclase after acute amphetamine

administration. J Pharmecol Exp Ther 237:820-825, 1986.

Barnett, J.V.; Segal, D.S.; and Kuczenski, R. Repeated amphetamine

pretreatment alters the responsiveness of striatal dopaminestimulated adenylate cyclase to amphetamine-induced desensitization.

J Pharmacol Exp Ther 242:40-47, 1987.

Beninger, R.J., and Hen, R.S. Pimozide blocks establishment but not

expression of cocaine-produced environment-specific conditioning.

Life Sci 38:1425-1431, 1986.

Borison; R.L; Hitri, A.; Klawans, H.L; and Diamond, B.I. A new

model for schizophrenia: Behavioral and receptor binding studies.

In: Usdin, E.; Kopin, I.J.; and Barchas, J., eds. Catecholamines:

Basic and Clinical Frontiers. New York: Pergamon Press, 1979.

pp. 719-721.

Braun, A.R., and Chase, T.N. Obligatory D-1/D-2 receptor interaction

in the generation of dopamine agonist related behaviors. Eur J

Pharmacol 131:301-306, 1986.

72
Carlson, J.H.; Bergstrom, D.A.; and Walters, J.R. Stimulation of both

D1 and D2 dopamine receptors appears necessary for full expression

of postsynaptic effects of dopamine agonists: A neurophysiological

study. Brain Res 400:205-218, 1987.

Cooper, D.M.F.; Bier-Laning, C.M.; Halford, M.K.; Ahlijanian, M.K.;

and Zahniser, N.R. Dopamine, acting through D-2 receptors,

inhibits rat striatal adenylate cyclase by a GTP-dependent process.

Mol Pharmacol 29:113-119, 1986.

Costall, B., and Naylor, R.J. Mesolimbic and extrapyramidal sites for

the mediation of stereotyped behavior patterns and hyperactivity by

amphetamine and apomorphine in the rat. In: Ellinwood, E.H., Jr.,

and Kilbey, M.M., eds. Advances in Behavioral Biology, Volume 21,

Cocaine and Other Stimulants. New York: Plenum Press, 1977.

pp. 47-76.

Creese, I., and Sibley, D.R. Receptor adaptations to centrally-acting

drugs. Annu Rev Pharmacol Toxicol 21:357-391, 1981.

Dubocovich, M.L, and Zahniser, N.R. Binding of the dopamine uptake

inhibitor 3H-nomifensine to striatal membranes. Biochem Pharmacol

34:1137-1144, 1985.

Dwoskin, L.P.; Peris, J.; Yasuda, R.P.; Philpott, K.; and Zahniser, N.R.

Repeated cocaine administration results in supersensitivity of

striatal D-2 dopamine autoreceptors to pergolide. Life Sci 42:255-

262, 1988.

Filloux, F.; Dawson, T.M.; and Wamsley, J.K. Localization of nigrostriatal dopamine receptor subtypes and adenylate cyclase. Brain

Res Bull 20:447-459, 1988.

Fischman, M.W.; Schuster, C.R.; Javaid, J.; Hatano, Y.; and Davis, J.

Acute tolerance development to the cardiovascular and subjective

effects of cocaine. J Pharmacol Exp Ther 235:677-682, 1985.

Gale, K. Catecholamine-independent behavioral and neurochemical

effects of cocaine in rats. In: Sharp, C.W., ed. Mechanisms of

Tolerance and Dependence: 1984. National Institute on Drug Abuse

Research Monograph 54. DHEW Pub. No. (ADM) 84-1330.

Washington, DC: Supt. of Docs., U.S. Govt. Print. Off., 1984. pp.

323-332.

Gershanik, 0.; Heikkila, R.E.; and Duvoisin, R.C. Behavioral

correlations of dopamine receptor activation. Neurology 33:1489

1492, 1983.

Goeders, N.E., and Kuhar, M.J. Chronic cocaine administration

induces opposite changes in dopamine receptors in the striatum and

nucleus accumbens. Alcohol Drug Res 7:207-216, 1987.

Greene, M.A., and White, F.J. Effects of chronic cocaine administration on A10 dopamine neurons in the rat: Electrophysiological

studies. Fed Proc 45:1061, 1986.

73
Hadfield, M.G., and Nugent, E.A. Cocaine: Comparative effect on

dopamine uptake in extrapyramidal and limbic systems. J

Neurochem 32:744-746, 1983.

Hanson, G.R.; Matsuda, LA.; and Gibbs, J.W. Effects of cocaine on

methamphetamine-induced neurochemicai changes: Characterization

of cocaine as a monoamine uptake blocker. J Pharmacol Exp Ther

242:507-513, 1987.

Henry, D.J.; Greene, M.A.; Chen, S-Y.; and White, F.J. Electrophysiological effects of repeated cocaine administration on the

mesoaccumbens dopamine system. Abstr Soc Neurosci 13:911, 1987.

Herrera-Marschitz, M., and Ungerstedt, U. Effect of the dopamine

D-1 antagonist SCH 23390 on rotational behavior induced by

apomorphine and pergolide in 6-hydroxy-dopamine denenrvated rats.

Eur J Pharmacol 109:349-354, 1985.

Jackisch, R.; Zumstein, A.; Hertting, G.; and Starke, K. lnterneurones

are probably not involved in the presynaptic dopaminergic control

of dopamine release in rabbit caudate nucleus. NaunynSchmiedebergs Arch Pharmacol 314:129-133, 1980.

Johnson, K.M., and Snell, L.D. Sensitization to the behavioral effects

of cocaine is associated with altered dopamine metabolism and

release in rat brain. Abstr Soc Neurosci 13:1718, 1987.

Kalivas, P.W., and Duffy, P. Effects of daily cocaine and morphine

treatment on somatodendritic and terminal field dopamine release.

J Neurochem 50:1498-1504, 1988.

Kalivas, P.W.; Duffy, P.; DuMars, L.A.; and Skinner, C. The

behavioral and neurochemical effects of acute and daily cocaine

administration in rats. J Pharmacol Exp Ther 245:485-492, 1988.

Kalivas, P.W., and Weber, B. Role of the Al 0 dopamine neurons to

cocaine and amphetamine. Abstr Soc Neurosci 13:600, 1987.

Kamal, L.A.; Arbilla, S.; Galzin, A.-M.; and Langer, S.Z. Amphetamine

inhibits the electrically evoked release of [

3H]dopamine from slices

of rabbi caudate. J Pharmacol Exp Ther 227:446-457, 1981.

Karoum, F.; Fawcett, R.; and Wyatt, R.J. The effect of chronic

cocaine treatment on catecholamine disposition in different brain

regions in rat: A specific target effect on frontal cortex dopamine.

Abstr Soc Neurosci 13: 1460, 1987.

Kelly, E., and Nahorski, S.R. Endogenous dopamine functionally

activates D-1 and D-2 receptors in striatum. J Neurochem 49:115-

120, 1987.

Lin-Chu, G.; Robinson, T.E.; and Becker, J.B. Sensitization of

rotational behavior produced by a single exposure to cocaine.

Pharmacol Biochem Behav 22:901-903, 1985.

74
Memo, M.; Pradhan, S.; and Hanbauer, I. Cocaine-induced supersensitivity of striatal dopamine receptors: Role of endogenous

calmodulin. Neuropharmacology 20:1145-1150, 1981.

Mishra, R.K.; Wong, Y.-W.; Varmuta, S.L; and Tuff, L. Chemical

lesion and drug induced sensitivity and subsensitivity of caudate

dopamine receptors. Life Sci 23:443-446, 1978.

Missale, C.; Castelletti, L.; Govoni, S.; Spano, P.F.; Trabucchi, M.; and

Hanbauer, I. Dopamine uptake is differentially regulated in rat

striatum and nucleus accumbens. J Neurochem 45:51-56, 1985.

Nayak, P.K.; Misra, A.L, and Mule, S.J. Physiological disposition and

biotransformation of 3H-cocaine in acute and chronically-treated

rats. J Pharmacol Exp Ther 196:556-569, 1975.

Niddam, R.; Arbilla, S.; Scatton, B.; Dennis, T.; and Langer, S.Z.

Amphetamine induced release of endogenous dopamine in vitro is

not reduced following pretreatment with reserpine. NaunynSchmeidebergs Arch Pharmacol 329:123-127, 1985.

Overstreet, D.H., and Yamamura, H.I. Receptor alterations and drug

tolerance. Life Sci 25:1865-1877, 1979.

Parker, E.M., and Cubeddu, L.X. Evidence for autoreceptor modulation of endogenous dopamine release from rabbit caudate nucleus in

vitro. J Pharmacol Exp Ther 232:492-500, 1985.

Parker, E.M., and Cubeddu, L.X. Effects of d-amphetamine and

dopamine synthesis inhibitors on dopamine and acetylcholine

neurotransmission in the striatum. I. Release in the absence of

vesicular transmitter stores. J Pharmacol Exp Ther 237:179-192,

1986a.

Parker, E.M., and Cubeddu, L.X. Effects of d-amphetamine and

dopamine synthesis inhibitors on dopamine and acetylcholine

neurotransmission in the striatum. II. Release in the presence of

vesicular transmitter stores. J Pharmacol Exp Ther 237:193-203,

1986b.

Peris, J., and Zahniser, N.R. One injection of cocaine produces a

long-lasting increase in [

3H]-dopamine release. Pharmacol Biochem

Behav 27:533-535, 1987.

Pitts, D.K., and Marwah, J. Cocaine modulation of central monoaminergic neurotransmission. Pharmacol Biochem Behav 26:453-461,

1987.

Post, R.M., and Contel, N.R. Human and animal studies of cocaine:

Implications for development of behavioral pathology. In: Creese,

I., ed. Stimulants: Neurochemical, Behavioral, and Clinical

Perspectives. Vol. I. New York: Raven Press, 1983. pp. 169-203.

Pradhan, S. Effect of cocaine on rat brain enzymes. Arch Int

Pharmacodyn Ther 266:221-228, 1983.

75
Ritz, M.C.; Lamb, R.J.; Goldberg, S.R.; and Kuhar, M.J. Cocaine

receptors on dopamine transporters are related to selfadministration of cocaine. Science 237:1219-1224, 1987.

Robinson, T.E., and Becker, J.B. Enduring changes in brain and

behavior produced by chronic amphetamine administration: A

review and evaluation of animal models of amphetamine psychosis.

Brain Res Rev 11:157-198, 1986.

Robinson, T.E.; Becker, J.B.; and Priesty, S.K. Long-term facilitation

of amphetamine-induced rotational behavior and striatal dopamine

release produced by a single exposure to amphetamine: Sex

differences. Brain Res 253:231-241, 1982.

Roy, S.N.; Bhattacharyya, A.K.; Pradhan, S.; and Pradhan, S.N.

Behavioral and neurochemical effects of repeated administration of

cocaine in rats. Neuropharmacol 7:559-564, 1978.

Schwartz, J.C.; Costenin, J.; Martres, M.P.; Protais, P.; and Baudry, M.

Modulation of receptor mechanisms in the CNS: Hyper- and hyposensitivity to catecholamines. Neuropharmacology 17:665-685, 1978.

Shindler, J.; Schachter, M.; Brincat, S.; and Parkes, J.D. Amphetamine, mazindol and fencamfamin in narcolepsy. Brit Med J Clin

Res 290:1167-1170, 1985.

Seeman, P. Brain dopamine receptors. PharmacoI Rev 32:229-313,

1980.

Stoof, J.C., and Kebabian, J.W. Opposing roles for D-1 and D-2

dopamine receptors in efflux of cyclic AMP from rat neostriatum.

Nature 294:366-368, 1981.

Stoof, J.C., and Verheijden, P.F.H.M. D-2 receptor stimulation

inhibits cyclic AMP formation brought about by D-1 receptor

stimulation in rat neostriatum but not nucleus accumbens. Eur J

Pharmacol 129:205-206, 1986.

Stripling, J.S., and Ellinwood, E.H., Jr. Sensitization to cocaine

following chronic administration in the rat. In: Ellinwood, E.H.,

Jr., and Kilbey, M.M., eds. Advances in Behavioral Biology, Volume

21, Cocaine and Other Stimulants. New York: Plenum Press, 1977.

pp. 327-351.

Taylor, D., and Ho, B.T. Neurochemical effects of cocaine following

acute and repeated injection. J Neurosci Res 3:95-101, 1977.

Taylor, D.; Ho, B.T.; and Fagan, J.D. Increased dopamine receptor

binding in rat brain by repeated cocaine injection. Commun

Psychopharmacol 3:137-142, 1979.

Trulson, M.E.; Babb, S.; Joe, J.C.; and Raese, J.D. Chronic cocaine

administration depletes tyrosine hydroxylase immunoreactivity in the

rat brain nigral striatal system: Quantitative light microscopic

studies. Exp Neurol 94:744-756, 1986.

76
Vaccheri, A.; Dall’Olio, R.; Gandolfi, O.; and Montanaro, N. Involvement of different dopamine receptors in rat diphasic motility

response to apomorphine. Psychopharmacology 89:265-268, 1986.

Wachtel, S.R.; Bednarz, L.M.; Brooderson, R.J.; Hjorth, S.; and White,

F.J. D-1 dopamine receptor stimulation enables functional responses

to D-2 dopamine receptor agonists. Abstr Soc Neurosci 13:910,

1987.

Walters, J.R.; Bergstrom, D.A.; Carlson, J.H.; Chase, T.N.; and Braun,

A.R. D1 dopamine receptor activation required for postsynaptic

expression of D2 agonist effects. Science 236:719-722, 1987.

Wise, R.A. Brain dopamine and reward. In: Cooper, S.J., ed.

Progress in Psychopharmacology. New York: Academic Press, 1981.

pp. 165-196.

Wise, R.A. Neural mechanisms of the reinforcing action of cocaine.

In: Grabowski, J., ed. Cocaine: Pharmacology, Effects and

Treatment of Abuse: 1984. National Institute on Drug Abuse

Research Monograph 43. DHEW Pub. No. (ADM) 84-1330.

Washington, DC: Supt. of Docs., US. Govt. Print. Off., 1984.

pp. 16-34.

Yeh, S.Y. Effects of repeated cocaine administration on brain

monoamines in rats. Fed Proc 46:404, 1987.

Zahniser, N.R.; Yasuda, R.P.; Dwoskin, L.P.; Philpott, K.; and

Dunwiddie, T.V. Repeated cocaine administration: Comparison

between central dopaminergic and noradrenergic systems. Abstr Soc

Neurosci 12:142, 1986.

ACKNOWLEDGEMENTS

This work was supported by U.S. Public Health Service grants

DA 04216, NS 09199, NS 01195, AM 07391, and MH 09387.

AUTHORS

Nancy R. Zahniser, Ph.D., Department of Pharmacology

Joanna Peris, Ph.D., Department of Pharmacology

Linda P. Dwoskin, Ph.D., Department of Pharmacology

Pamela Curella, B.S., Department of Pharmacology

Robert P. Yasuda, Ph.D., Department of Pharmacology

Laurie O’Keefe, M.S., Department of Neurology

Sally J. Boyson, M.D., Departments of Pharmacology and Neurology

University of Colorado Health Sciences Center

4200 East Ninth Avenue

Denver, CO 80262

77
The Interaction of Cocaine With

Central Serotonergic Neuronal

Systems: Cellular Electrophysiologic

Approaches

Joan M. Lakoski and Kathryn A. Cunningham

The numerous behavioral and physiological effects elicited by cocaine

in humans range from affective changes (euphoria and mood elevation) at low doses, to psychiatric disorders (anxiety, depression, and

paranoid psychosis) at high or chronic doses (Fischman 1987). Many

of these effects of cocaine have been related to the ability of

cocaine to stimulate catecholamine neurotransmission in the central

nervous system (CNS) (Seiden and Kleven, this volume). However,

cocaine has also been demonstrated to significantly alter central

serotonin (5-hydroxytryptamine (5HT) function (Blackburn et al.

1967; Friedman et al. 1975; Ross and Renyi 1967; Ross and Renyi

1969; Taylor and Ho 1978). Since brain SHT-containing neuronal

systems have been demonstrated to be involved in psychiatric disorders (Ceulemans et al. 1985; lversen 1984; King et al. 1985; Van Praag

1982), including those evidenced by abusers of this stimulant, a possible role for the interaction of cocaine with serotonergic neuronal

systems warrants further investigation.

The classic pharmacological profile of cocaine includes both its

actions as a local anesthetic (Ritchie and Greene 1985) and as an

inhibitor of monoamine neurotransmitter reuptake. While cocaine has

been recognized to potently inhibit the uptake of the catecholamines

dopamine (DA) (Bagchi and Reilly 1988; Heikkila et al. 1975; Komisky

et al. 1977; Nielsen et al. 1988; Taylor and Ho 1978) and norepinephrine (NE) (Hadfield et al. 1980; Komisky et al. 1977; Taube et al.

1975; Taylor and Ho 1978), a similar inhibition of reuptake also

occurs for the indoleamine 5-HT (Blackbum et al. 1967; Koe 1976;

Ross and Renyi 1967; Ross and Renyi 1969; Taylor and Ho 1978).

Although equipotent as a reuptake inhibitor of these monoamine

neurotransmitters, cocaine is actually more efficacious as an inhibitor

of 5-HT uptake as compared to DA or NE uptake processes. These

78
data indicate that interactions with central serotonergic function may

underlie some of the observed psychological and physiological effects

of cocaine.

Studies of the content and turnover of 54-HT after acute or chronic

cocaine administration also suggest the importance of cocaine-5-HT

interactions. Several groups have reported brain levels of 5-HT to be

unchanged following administration of cocaine (Friedman et al. 1975;

Schubert et al. 1970). However, Pradhan and colleagues (Pradhan et

al. 1978a; Pradhan et al. 1978b; Pradhan et al. 1981; Roy et al. 1978)

have also identified significant decreases in the levels of 5-HT in

several brain regions following either acute or chronic cocaine

administration. In addition, several laboratories have identified

decreases in 5-HT synthesis produced by this psychostimulant

(Friedman et al. 1975; Galloway and Novak 1986; Post et al. 1976;

Schubert et al. 1970). Furthermore, cocaine has been shown to

decrease both the uptake of the 54-HT precursor tryptophan and

increase the activity of tryptophan hydroxylase, the rate-limiting

enzyme in 5-HT synthesis (Knapp and Mandell 1972; Knapp and

Mandell 1976). Likewise, studies of 3H-cocaine binding have revealed

both a differential distribution of binding sites in the brain (Kennedy

and Hanbauer 1983; Reith et al. 1988; Shah et al. 1982) as well as a

presynaptic labeling of DA or 5-HT reuptake sites both in the

striatum (Kennedy and Hanbauer 1988; Reith et al. 1988; Reith et al.

1985; Schoemaker et al. 1985) and the cortex (Reith et al. 1988).

Behavioral studies of cocaine’s actions have also implicated a role for

5-HT neuronal systems in the mediation of specific drug-induced

behaviors. Inhibition of 5-HT synthesis and selective 5-HT receptor

blockade have been reported to potentiate cocaine-induced locomotor

activity (Scheel-Kruger et al. 1977). Conversely, administration of

the precursor 5-hydroxytryptophan has been demonstrated to attenuate the locomotor activity produced by cocaine administration

(Pradhan et al. 1978a). It is relevant to note that although a role

for DA in mediating the reinforcing and discriminative stimulus prop

erties of cocaine is supported by much of the behavioral research

(Koob and Hubner, this volume; Woolverton and Kleven, this volume),

not all of the behavioral effects of cocaine are consistently inhibited

by DA antagonists (Colpaert et al. 1976; Cunningham and Appel 1982;

Friedman et al. 1975; Spyraki et al. 1982). Thus, the behavioral

pharmacology of cocaine suggests that interactions with other neurotransmitters are probably important in mediating the effects of

cocaine.

79
In assessing the neurophannacological ramifications of the interaction

of cocaine with central 5-HT neuronal systems, we have begun to

study the effects of cocaine administration on the spontaneous activity of serotonin neurons recorded extracellularly in the dorsal raphe

nucleus (DRN) of the anesthetized rat. Such a single-unit electrophysiologic approach can help to identify the precise cellular and

molecular physiological events by which cocaine may act on specific,

identified neuronal populations in the brain. These serotonincontaining neurons provide a useful neurobiological model system for

studies of the cellular actions of cocaine, as these DRN neurons are

well characterized, both with respect to their pharmacological and

physiological properties. Not only do these midbrain 5-HT cells have

an identified transmitter substance (Dahlstrom and Fuxe 1964), but

they are readily identified by standard electrophysiologic recording

techniques and discharge spontaneously with a uniform, steady firing

rate (Aghajanian 1976; Haigler and Aghajanian 1974). The synaptic

projections of these 5-HT neurons have also been well characterized

and include extensive innervation of all forebrain regions (Azmitia

and Segal 1978; Conrad et al. 1976; Moore et al. 1978; Parent et al.

1981; Steinbusch 1981; Ungerstedt 1971). A wealth of information has

accumulated with respect to the direct inhibitory effects of microiontophoretically applied 5-HT (Aghajanian 1972; Aghajanlan et al.

1987; Trulson and Jacobs 1976; VanderMaelen 1985), histamine

(Lakoski and Aghajanian 1983; Lakoski et al. 1984), and GABA

(Gallager and Aghajanian 1976b; Wang and Aghajanian 1977) as well

as excitatory effects of the microiontophoretic application of NE

(Baraban and Aghajanian 1980; Gallager and Aghajanian 1976a; Menkes

et al. 1981; Rogawski and Aghajanian 1982) on these neurons. The

inhibitory and excitatory inputs for this presynaptic 5-HT autoreceptor located in the DRN are diagrammed in figure 1.

Systemic administration of cocaine has recently been demonstrated to

inhibit the spontaneous activity of 5-HT neurons recorded in the DRN

(Cunningham and Lakoski 1986; Cunningham and Lakoski 1988; Pitts

and Marwah 1986b Pitts and Marwah 1987a). A complete, dosedependent, and reversible inhibition of the spontaneous firing of

these neurons is observed following intravenous administration of

cocaine (cumulative dose to 50 percent inhibition = 0.66 ± 0.11 mg/kg)

(figure 2). While the central effects of cocaine may be mediated, in

part, via local anesthetic properties of the drug, this is not apparent

with respect to the modulation of 5-HT neuronal activity. As recorded in the DRN, intravenous administration of the local anesthetics

procaine or lidocaine (Cunningham and Lakoski 1988; Pitts and

Marwah 1986b) (figure 2) failed to alter spontaneous cell firing

80
FIGURE 1. A summary of identified physiological inputs to serotonin

(5-HT)-containing dorsal raphe neurons

KEY: NE, norspinephrine; 5-HT, serotonin; GABA, aminobutyric acid; SP, substance

P; EXCIT A.A., excitatory amino acids; HIS, histamine. The plus symbol (+)

indicates a given substance enhances spontaneous cell tiring, while a minus

symbol (-) indicates a corresponding inhibition.

in a manner similar to that observed with cocaine; indeed, several

neurons found to be unresponsive to procaine administration were

inhibited by subsequent systemic cocaine application (Cunningham and

Lakoski 1988).

Since cocaine is a potent inhibitor of 5-HT reuptake processes in

brain tissues, pharmacological characterization of the effects of

cocaine on DRN cell firing has involved comparison of the effects of

cocaine with uptake inhibitors that are selective for DA, NE, or

5-HT. Fluoxetine, a selective 5-HT uptake inhibitor, but not the

selective uptake inhibitors for NE (desipramine) or DA (GBR 12909),

inhibited 5-HT cell firing in a dose-response manner (Cunningham

et al. 1987b); in addition, pretreatment with the 5-HT synthesis

inhibitor p-chlorophenylalanine reduced the sensitivity of 5-HT DRN

neurons to the systemic application of this psychostimulant). These

data are consistent with the idea that depressant effects of cocaine

on serotonergic cellular physiology may result from a direct inhibition

of 5-HT neuronal activity as a consequence of the selective inhibition

of monoamine reuptake (Lakoski and Cunningham 1988). In support

81
CUMULATIVE DOSE (mg/kg,i.v.)

FIGURE 2. Comparison of the effects of intravenous cocaine and

local anesthetics administration on spontaneous

serotonergic cell firing recorded in the dorsal raphe

nucleus

NOTE: Data are presented for the cumulative dose-response curve to cocaine (filled

circles: n=17), lidocaine (filled squares: n=5), and procaine (open circles; n=10).

Each 5-HT neuron was recorded for at least 3 minutes prior to drug injection

to establish a stable baseline firing rate, and then drugs were administered

intravenously at 2-minute intervals in a cumulative dose manner; the percent

change from the baseline firing rate was calculated for each dose by using the

mean firing rate during the 2-minute period between doses (ordinate). Only

one drug was tested per cell and per animal.

of this hypothesis are the observations by Pitts and Marwah (1987a)

that pretreatment with reserpine, which depletes neuronal stores of

monoamines including 5-HT, significantly attenuates the ability of

systemically administered cocaine to suppress midbrain 5-HT activity.

82
Microiontophoretic drug application techniques have also been utilized

in combination with extracellular recording techniques to assess

whether cocaine has direct effects on 5-HT cell firing in the DRN

(Cunningham and Lakoski 1988); this approach is particularly advantageous as it facilitates the direct investigation of synaptic

mechanisms that may mediate the central effects of cocaine. Microiontophoretic application of cocaine produces a current-dependent

suppression of spontaneous 5-HT cell firing without concomitant evidence of any apparent local anesthetic effects (figure 3). This

depression of neuronal activity by cocaine is characterized by a rate

of onset and offset similar to that observed with 5-HT as recorded in

the DRN. Cocaine application, at ejection currents that did not

initially alter spontaneous baseline firing rates, has been found to

potentiate the inhibitory effects of 5-HT when microiontophoretically

applied together (Cunningham and Lakoski 1988). In contrast, when

applied at currents where no local anesthetic effects were apparent,

the microiontophoretic application of procaine consistently failed to

inhibit 5-HT neurons.

FlGURE 3. Effects of the microiontophoretic application of cocaine

and serotonin (5-HT) on the spontaneous firing rate of

5-HT neurons recorded in the dorsal raphe nucleus

NOTE: In this frequency histogram, the duration of drug ejection is indicated by the

bar above the record; numbers refer to iontophoretic current in nanoamperes

used for drug ejection. Cocaine (C; 0.01 M; dots) and 5-HT (5-HT; 0.04 M;

solid bars) depressed cell firing at low microiontophoretic currents.

Acute cocaine administration has been characterized to significantly

modify the electrophysiologic activity of 5-HT neurons. Acknowledging that repetitive, long-term self-administration of this psychostimulant is not uncommon in the user profile of cocaine abusers

83
(Fischman 1987), the pharmacological consequences of prolonged

exposure to this psychostimulant also merit consideration. Preliminary studies suggest that significant modification of both the

cellular pharmacology and physiology of 5-HT neurons is observed in

the midbrain raphe nuclei following repeated cocaine exposure

(Cunningham et al. 1987a). Clearly, the relevance of this change in

5-HT neuronal function observed with chronic cocaine exposure to

psychiatric disturbances observed in abusers remains unknown and

merits further investigation. Likewise, an analysis of the ionic

mechanisms that may underlie both acute and chronic effects of

cocaine on 5-HT function are important areas for future

investigations.

While studies utilizing single-unit electrophysiologic approaches to

characterize the central effects of cocaine on identified monoamine

neurons have only recently been undertaken, several important

aspects of cocaine-induced modulation of 5-HT function reveal a similar profile for the monoamines DA and NE. Inhibitory effects of

acute systemic cocaine have been identified on the spontaneous activity of DA-containing neurons recorded in the ventral tegmental area

(Einhorn et al. 1988; Pitts and Marwah 1987a) and in a postsynaptic

terminal region of these cells, the nucleus accumbens (White 1986;

White et al. 1987). Analogous to observations regarding 5-HT neuronal function in the DRN, mesoaccumbens, but not mesocortical,

projecting DA neurons are inhibited in a rapid, dose-related, and

reversible manner (cumulative dose to 50 percent inhibition = 2.2 ±

0.3 mg/kg) following intravenous cocaine administration (Einhom

et al. 1988; Pitts and Marwah 1987a; White 1985); however, complete

suppression of baseline firing is not achieved in any of the DA cell

groups with systemic cocaine infusion. Similar to responses repotted

in the DRN, pretreatment with reserpine effectively attenuates

cocaine-induced inhibition of DA cell firing recorded in the ventral

tegmental area (Einhom et al. 1988; Pitts and Marwah 1987a); the

selective DA antagonist sulpiride blocks this cocaine-mediated suppression of cell firing recorded both in vivo and in vitro (Brodie and

Dunwiddie 1986; Einhom et al. 1988; White 1986). In studies of the

NE-containing cells recorded in the locus coeruleus, the same pattern

of cocaine-induced responses emerges as that shown for the 5-HT and

DA neuronal systems. Intravenous administration of cocaine rapidly,

dose-dependently, and reversibly suppresses NE neurons in this brain

region to a maximal 80 percent inhibition of baseline firing rates

(Pitts and Marwah 1986a; Pitts and Marwah 1986b; Pitts and Matwah

1987b); likewise, pretreatment with reserpine or administration of the

84
adrenergic antagonist piperoxane effectively attenuates the stimulantinduced changes in locus coeruleus neuronal activity. The observed

differences in the potency of cocaine to suppress the spontaneous

activity of monoamine-containing neurons (i.e., 5-HT>NE>DA) suggest

possible differential contributions of these neuronal systems to the

diverse behavioral effects of cocaine.

Just as cocaine potentiates the inhibitory responses of DRN neurons

to 5-HT, similar modulation has also been revealed for DA- and NEmediated responses. Microiontophoretic application of cocaine significantly potentiates DA-induced depression of cell firing in the ventral

tegmental area and nucleus accumbens (Brodie and Dunwiddie 1986;

Einhom and White 1986; Einhorn et al. 1988; White et al. 1987).

While the direct microiontophoretic effects of cocaine on identified

NE-containing neurons remain, as yet, uncharacterized, pronounced

enhancement of NE-mediated responses has been identified utilizing in

vitro hippocampal (Yasuda et al. 1984) and locus coeruleus

(Surprenant and Williams 1987) preparations. These observations,

which demonstrate potentiation of 5-HT, DA, and NE responses by

cocaine, are in agreement with the neurochemical profile of cocaine

as an inhibitor of monoamine reuptake. Clearly, additional studies

are needed to further assess the site-specific importance of qcocaineinduced modulation of central monoamine function. Single-unitrecording electrophysiologic approaches will continue to provide

needed information regarding the central effects of cocaine on these

neuronal systems.

REFERENCES

Aghajanian, G.K. LSD and CNS transmission. Annu Rev Pharmacol

12:157-168, 1972.

Aghajanian, G.K. LSD and 2-Bromo LSD: Comparison of effects on

serotonergic neurones and neurones in two serotonergic projection

areas, the ventral lateral geniculate and amygdala. Neuropharmacology 15:521-528, 1976.

Aghajanian, G.K.; Sprouse, J.S.; and Rasmussen, K. Physiology of the

midbrain serotonin system. In: Meltzer, H.Y., ed. Psychopharmacology: The Third Generation of Progress. New York: Raven

Press, 1987. pp. 141-149.

Azmitia, E.C., and Segal, M. An autoradiographic analysis of the

differential ascending projections of the dorsal and median raphe

nuclei in the rat. J Comp Neurol 179:641-668, 1978.

85
Bagchi, S.P., and Reilly, M.A. lntraneuronal dopaminergic action of

cocaine and some of its metabolites and analogs. Neuropharmacology 22:1289-1295, 1983.

Baraban, J.M., and Aghajanian, G.K. Suppression of firing activity of

5-HT neurons in the dorsal raphe by alpha-adrenoceptor antagonists. Neuropharmacology 19:355-363, 1980.

Blackburn, K.J.; French, P.C.; and Merrills, R.J. 5-Hydroxytryptamine

uptake by rat brain in vitro. Life Sci 6:1653-1663, 1967.

Brodie, M.S., and Dunwiddie, T.V. The effects of cocaine on ventral

tegmental area spontaneous activity in vitro: Interactions with

dopamine, sulpiride and cholecystokinin. Abstr Soc Neurosci

12(1):233, 1986.

Ceulemans, D.L.S.; Gelders, Y.G.; Hoppenbrouwers, M.-L.J.A.;

Reyntjens, A.J.M.; and Janssen, P.A.J. Effect of serotonin

antagonism in schizophrenia: A pilot study with setoperone.

Psychopharmacology (Berlin) 85:329-332, 1985.

Colpaert, F.C.; Niemegeers, C.J.E.; and Janssen, P.A.J. Cocaine cue in

rats as it relates to subjective drug effects. Eur J Pharmacol

40:195-199, 1976.

Conrad, L.C.A.; Leonard, C.M.; and Pfaff, D.W. Connections of the

medial and dorsal raphe nuclei in the rat: An autoradiographic and

degeneration study. J Comp Neural 156:179-206, 1976.

Cunningham, K.A., and Appel, J.B. Discriminative stimulus properties

of cocaine and phencyclidine: Similarities in mechanism of action?

In: Colpaert, C., and Slangen, J.L., eds. Drug Discrimination:

Applications in CNS Pharmacology. Janssen Research Foundation

Series. Vol. 6. Amsterdam: Elsevier Biomedical Press, 1982.

pp. 181-192.

Cunningham, K.A.; Asprodini, E.K.; Bernau, N.A.; Richard, C.A.; and

Lakoski, J.M. Enhanced inhibitory responses of serotonin neurons

in the dorsal raphe nucleus (DRN) after repeated cocaine exposure.

Abstr Soc Neurosci 13(3):1651, 1987a.

Cunningham, K.A.; Bernau, N.A.; and Lakoski, J.M. Comparison of

cocaine congeners and monoamine uptake inhibitors: Effects on

serotonin (5-HT) neurons of the dorsal raphe nucleus (DRN).

Presented at the 10th International Congress of Pharmacology,

Sydney, Australia, August 1987b. p. 261.

Cunningham, K.A., and Lakoski, J.M. Cocaine: Inhibition of serotonergic cell firing in the dorsal raphe nucleus (DRN). Fed Proc

45:1060, 1986.

Cunningham, K.A., and Lakoski, J.M. Electrophysiological effects of

cocaine and procaine on dorsal raphe serotonin neurons. Eur J

Pharmacol 148:457-462, 1988.

86
Dahlstrom, A., and Fuxe, K. Evidence for the existence of monoamine neurons in the central nervous system. I. Demonstration of

monoamines in the cell bodies of brainstorm neurons. Acta Physiol

Scand 62 [Suppl] 232:1-55, 1964.

Einhom, L.C.; Johansen, PA.; and White, F.J. Electrophysiological

effects of cocaine in the mesoaccumbens dopamine system: Studies

in the ventral tegmental area. J Neurosci 8:100-112, 1988.

Einhom, L.C., and white, F.J. Electrophysiological effects of cocaine

in the mesoaccumbens dopamine system: Studies in the ventral

tegmental area. Abstr Soc Neurosci 12(2):1516, 1986.

Fischman, M.W. Cocaine and the amphetamines. In: Meltzer, H.Y.,

ed. Psychopharmacology: The Third Generation of Progress. New

York: Raven Press, 1987. pp. 1543-1553.

Friedman, E.; Gershon, S.; and Rotrosen, J. Effects of acute cocaine

treatment on the turnover of 5-hydroxytryptamine in the rat brain.

Br J Pharmacol 54:61-64, 1975.

Gallager, D.W., and Aghajanian, G.K Effects of antipsychotic drugs

on the firing of dorsal raphe cells. I. Role of adrenergic systems.

Eur J Pharmacol 39:341-355, 1976a.

Gallager, D.W., and Aghajanian, G.K Effects of antipsychotic drugs

on the firing of dorsal raphe cells. II. Reversal by picrotoxin.

Eur J Pharmacol 39:357-364, 1976b.

Galloway, M.P., and Novak, E.A. Effects of cocaine on monoamine

synthesis in mesocortical neurons. Fed Proc 45:1061, 1986.

Hadfield, M.G.; Mott D.E.W.; and Ismay, J.A. Cocaine: Effect on in

vivo administration of synaptosomal uptake of norepinephrine.

Biochem Pharmacol 29:1861-1863, 1980.

Haigler, H.J., and Aghajanian, G.K Peripheral serotonin antagonists:

Failure to antagonize serotonin brain areas receiving a prominent

serotonergic input. J Neural Transm 35:257-273, 1974.

Heikkila, R.E.; Orlansky, H.; and Cohen, G. Studies on the distinction

between uptake inhibition and release of 3H-dopamine in rat brain

tissue slices. Biochem Pharmacol 24:847-852, 1975.

lversen, S.D. Serotonin and anxiety. Neuropharmacology 23:1553-

1560, 1984.

Kennedy, L.T., and Hanbauer, I. Sodium-sensitive cocaine binding to

rat striatal membrane: Possible relationship to dopamine uptake

sites. J Neurochem 41:172-178, 1983.

King, R.; Faull, K.F.; Stahl, S.M.; Mefford, I.N.; Thiemann, S.:

Barchas, J.D.; and Berger, P.A. Serotonin and schizophrenia:

Correlation between serotonergic activity and schizomotor behavior.

Psychiatry Res 14:235-240, 1985.

87
Knapp, S., and Mandell, A.J. Narcotic drugs: Effects on the

serotonin biosynthetic systems of the brain. Science 177:1209-1211,

1972.

Knapp, S., and Mandell, A.J. Cocaine and lithium: Neurobiological

antagonism in the serotonin biosynthetic system in rat brain. Life

Sci 18:679-684, 1976.

Koe, B.K Molecular geometry of inhibitors of the uptake of

catecholamines and serotonin in synaptosomal preparations of rat

brain. J Pharmacol Exp Ther 199:649-661, 1976.

Komisky, H.L; Miller, D.D.; La Pidus, J.B.; and Patil, P.N. The

isomers of cocaine and tropacocaine: Effect of 3H-catecholeamine

uptake by rat brain synaptosomes. Life Sci 21:1117-1122, 1977.

Lakoski, J.M., and Aghajanian, G.K Effects of histamine, H1- and

Hz-receptor antagonists on the activii of serotonergic neurons in

the dorsal raphe nucleus. J Pharmacol Exp Ther 227:517-523, 1983.

Lakoski, J.M., and Cunningham, K.A. Cocaine interaction with central

monoaminergic systems: Electrophysiologic approaches. Trends

Pharmocol Sci 9(5):177-180, 1988.

Lakoski, J.M.; Gallager, D.W.; and Aghajanian, G.K Histamine-induced

depression of serotonergic dorsal raphe neurons. Antagonism by

cimetidine, a reevaluation. Eur J Pharmacol 103:153-156, 1984.

Menkes, D.B.; Baraban, J.M.; and Aghajanian, G.K Prazosin

selectively antagonizes neuronal responses mediated by alpha,-

adrenoceptors in brain. Naunyn Schmiedebergs Arch Pharmacol

317:273-275, 1981.

Moore, R.Y.; Halaris, A.E.; and Jones, B.E. Serotonin neurons in the

midbrain raphe: Ascending projections. J Comp Neurol 180:417-438,

1978.

Nielsen, J.A.; Chapin, D.S.; and Moore, K.E. Differential effects of

d-amphetamine, phenylethylamine, cocaine and methylphenidate

on the rate of dopamine synthesis in terminals of nigrostriatal and

mesolimbic neurons and on the efflux of dopamine metabolites into

cerebroventricular perfusates of rats. Life Sci 33:1899-1907, 1983.

Parent, A.; Descarries, C.; and Beaudet, A. Organization of ascending

serotonin systems in the aduit rat brain. A radioautographic study

after intraventricular administration of 3H 5-hydroxytryptamine.

Neuroscience 6:115-138, 1981.

Pitts, D.K, and Marwah, J. Effects of cocaine on the electrical

activity of single noradrenergic neurons from locus coeruleus. Life

Sci 33:1229-1234, 1986a.

Pitts, D.K, and Marwah, J. Electrophysiological effects of cocaine on

central monoaminergic neurons. Eur J Pharmacol 131:95-98, 1986b.

88
Pitts D.K., and Marwah, J. Cocaine modulation of central monoaminergic neurotransmission. Pharmacol Biochem Behav 26:453-461,

1987a.

Pitts, D.K., and Marwah, J. Electrophysiological actions of cocaine

on noradrenergic neurons in rat locus coeruleus. J Pharmacol Exp,

Ther 240:345-351, 1987b.

Post, R.M.; Kopanda, R.T.; and Black, K.E. Progressive effects of

cocaine on behavior and central amine metabolism in rhesus

monkeys: Relationship to kindling and psychosis. Biol Psychiatry

11:403-419, 1976.

Pradhan, S.N.; Bhattacharyya, A.K.; and Pradhan, S. Serotonergic

manipulation of the behavioral effects of cocaine in rats.

Communications in Psychopharmacology 2:481-486, 1978a.

Pradhan, S.; Hanson, G.; and Lovenberg, W. Inverse relation of

substance P-like immunoreactivity in dorsal raphe nucleus to

serotonin levels in pons-medulla following administration of cocaine

and 5-hydroxytryptophan. Biochem Pharmacol 30:1071-1076, 1981.

Pradhan, S.; Roy, S.N.; and Pradhan, S.N. Correlation of behavioral

and neurochemical effects of acute administration of cocaine in

rats. Life Sci 22:1737-1744, 1978b.

Reith, M.E.; Meisler, B.E.; Sershen, H.; and Lajtha, A. Sodiumindependent binding of 3H-cocaine in mouse striatum is serotonin

related. Brain Res 342:145-148, 1985.

Reith, M.E.A.; Sershen, H.; Allen, D.L; and Lajtha, A. A portion of

3H-cocaine binding in brain is associated with serotonergic neurons.

Mol Pharmacol 23:600-606, 1983.

Ritchie, J.M., and Greene, N.M. Local anesthetics, In: Gilman, A.G.;

Goodman, L.S.; Rall, T.W.; and Murad, F., eds. The Pharmacological

Basis of Therapeutics. 7th ed. New York: MacMillan, 1985.

pp. 302-321.

Rogawski, M.A., and Aghajanian, G.K. Activation of lateral geniculate

neurons by locus coeruleus or dorsal noradrenergic bundle stimulation: Selective blockade by the alpha1

-adrenoceptor antagonist

prazosin. Brain Res 250:31-39, 1982.

Ross, S.B., and Renyi, A.L Inhibition of the uptake of tritiated

catecholamines by antidepressants and related agents. Eur J

Pharmacol 2:181-186, 1967.

Ross, S.B., and Renyi, A.L Inhibition of the uptake of tritiated

5-hydroxytryptamine in brain tissue. Eur J Pharmacol 7:270-277,

1969.

Roy, S.N.; Bhattacharyya, A.K.; Pradhan, S.; and Pradhan, S.N.

Behavioral and neurochemical effects of repeated administration of

cocaine in rats. Neuropharmacology 17:559-564, 1978.

89
Scheel-Kruger, J.; Braestrup, C.; Nielson, M.; Golembrowska, K.; and

Mogilnicka, E. Cocaine: Discussion on the role of DA in the

biochemical mechanism of action. In: Ellinwood, E.H., and Kilbey,

M.M., eds. Cocaine and Other Stimulants. New York: Plenum

Press, 1977. pp. 373-407.

Schoemaker, H.; Pimoule, C.; Arbilla, S.; Scatton, B.; Tavoy-Agid, F.;

and Langer, S.Z. Sodium-dependent 3H-cocaine binding associated

with dopamine uptake sites in the rat striatum and human putamen

decrease after dopaminergic denervation and in Parkinsons disease.

Naunyn Schmiedebergs Arch Pharmacol 329:227-235, 1985.

Schubert, J.; Fyro, B.; Nyback, H.; and Sedvall, G. Effects of cocaine

and amphetamine on the metabolism of tryptophan and 5-hydroxytryptamine in mouse brain in vivo. J Pharm Pharmacol 22:860-862,

1970.

Shah, N.S.; Powell, D.B.; Shah, A.B.; Wiscovitch, R.; and McAmis, W.

Distribution of 3H-levo-cocaine in brain regions of rabbit after

intracerebroventricular administration. Research Communications in

Substances of Abuse 3:433-441, 1982.

Spyraki, C.; Fibiger, H.C.; and Phillips, A.G. Cocaine-induced place

preference conditioning: Lack of effects of neuroleptics and

6-hydroxydopamine lesions. Brain Res 253: 195-203, 1982.

Steinbusch, H.W.M. Distribution of serotonin immunoreactivity in the

central nervous system of the rat-cell bodies and terminals.

Neuroscience 6:557-618, 1981.

Surprenant, A., and Williams, J.T. Inhibitory synaptic potentials

recorded from mammalian neurones prolonged by blockade of

noradrenaline uptake. J Physiol (Lond) 382:87-103, 1987.

Taube, H.D.; Montel, H.; and Starke, K. Phencyclidine and ketamine:

Comparison with the effect of cocaine on the noradrenergic neurons

of the rat brain cortex. Naunyn Schmiedebergs Arch Pharmacol

291:47-54, 1975.

Taylor, D., and Ho, B.T. Comparison of inhibition of monoamine

uptake by cocaine, methylphenidate and amphetamine. Res Commun

Chem Pathol Pharmacol 21:67-75, 1978.

Trulson, M.E., and Jacobs, B.L Dose-response relationships between

systemically administered L-tryptophan or L-5-hydroxytryptophan

and raphe unit activity in the rat. Neuropharmacology 15:339-344,

1976.

Ungerstedt, U. Stereotaxic mapping of monoamine pathways in rat

brain. Acta Physiol Stand 367: 1-48, 1971.

VanderMaelen, C.P. Serotonin. In: Rogawski, M.A., and Barker,

J.L, eds. Neurotransmitter Actions in the Vertebrate Nervous

System. New York: Plenum Press, 1985. pp. 201-240.

90
Van Praag, H.M. Neurotransmitters and depression. Part A.:

lndoleamines and depression. In: Beumont, P.J.V., and Burrows,

G.D., eds. Handbook of Psychiatry and Endocrinology. Amsterdam:

Elsevier Biomedical Press, 1982. pp. 267-290.

Wang, R.Y., and Aghajanian, G.K. Physiological evidence for habenula

as a major link between forebrain and midbrain raphe. Science

197:89-91, 1977.

White, F.J. Electrophysiological effects of cocaine in the mesoaccumbens and mesocortical dopamine systems. Abstr Soc Neurosci

11(2):828, 1985.

White, F.J. Electrophysiological effects of cocaine in the mesoaccumbens dopamine system: Studies in the nucleus accumbens.

Abstr Soc Neurosci 12(2):1516, 1986.

White, F.J.; Whatcel, S.R.; Johansen, P.A.; and Einhom, LC. Electrophysiological studies of the rat mesoaccumbens dopamine system:

Focus on dopamine receptor subtypes, interactions, and the effects

of cocaine. In: Chiodo, LA., and Freeman, A.S., eds. Neurophysiology of Dopaminergic Systems-Current Status and Clinical

Perspectives. Grosse Pointe, Ml: Lakeshore Publishing Company,

1987. pp. 317-365.

Yasuda, R.P.; Zahniser, N.R. ; and Dunwiddie, T.V. Electrophysiological effects of cocaine in the rat hippocampus in vitro. Neurosci

Lett 45:199-204, 1984.

ACKNOWLEDGMENTS

Efthihia Asprodini and Nicola Bernau of the Department of Pharmacology and Toxicology at the University of Texas Medical Branch

provided expert assistance with electrophysiological recording

studies. The preparation of this manuscript was supported by U.S.

Public Health Service grant 5 R01 DA 04296 from the National

Institute on Drug Abuse.

AUTHORS

Joan M. Lakoski, Ph.D., Assistant Professor

Kathryn A. Cunningham, Ph.D., Assistant Professor

Department of Pharmacology and Toxicology

University of Texas Medical Branch

Galveston, TX 77550

91
The Effects of Cocaine on Local

Cerebral Metabolic Activity

Linda J. Porrino and Conan Kornetsky

INTRODUCTlON

The development of the 2-[14C]deoxyglucose (2-DG) method by Louis

Sokoloff and his colleagues (1977) has provided a powerful tool with

which neuroscientists can investigate the functional events in the

brain related to various physiological, pharmacological, and behavioral

states. lt has been used extensively in neuropharmacology to identify

the neural circuits that mediate the effects of a wide variety of

pharmacological agents (McCulloch 1982).

The quantitative 2-DG autoradiographic method measures local rates

of cerebral glucose utilization in all regions of the central nervous

system simultaneously. In the brain, as in all tissues that do

physicochemical work, the amount of energy used reflects the amount

of work done. Glucose utilization as a measure of energy use under

normal physiological conditions (in the brain, glucose is virtually the

exclusive substrate for energy metabolism) is coupled to functional

activity. Because of this close relationship between energy metabolism and functional activity, it is possible to identify regions of the

brain with altered functional activity in various conditions.

The 2-DG method, then, does not measure functional activity directly,

but measures glucose utilization, and changes in functional activity

are inferred from the changes in glucose utilization. The validity of

this assumption has been demonstrated in numerous experiments in

which evoked changes in the functional activity of a variety of

sensory and motor pathways have been shown by the 2-DG method to

be associated with corresponding changes in rates of glucose utilization in the components of the activated or inhibited system (Sokoloff

1982).

92
Although a number of energy-requiring processes such as synthesis of

neurotransmitters and amino acids contribute to the basal rates of

glucose utilization, the alterations in glucose utilization evoked by a

pharmacological, physiological, or behavioral manipulation are thought

to result mainly from increases or decreases in electrical activity,

i.e., spike frequency in the central nervous system. These changes in

electrical activity in turn produce corresponding increases or

decreases in the activity of Na+, K+-ATPase, the energy-consuming

enzyme involved in the restoration of neuronal ionic gradients to

their resting state. Experiments with deoxyglucose have shown that

the coupling of glucose utilization to functional activity is dependent

on Na+, K+-ATPase activity, in that ouabain, an ATPase inhibitor, can

block the increases in glucose utilization that accompany electrical

stimulation (Mata et al. 1980). Another significant issue in the

interpretation of changes in glucose utilization is the site of these

changes. A number of studies (Schwartz et al. 1979; Kadekaro et al.

1985) have now clearly demonstrated that functional activation of

glucose utilization occurs mainly in nerve terminals rather than in

cell bodies. This means that alterations in energy metabolism in a

specific brain area are due to changes in the afferent inputs to that

structure and not in the cell bodies contained within the structure.

To measure rates of local cerebral glucose utilization, a radioactively

labeled analog of glucose, 2-DG, is used, which, like glucose, is

transported into cerebral tissue and phosphorylated by hexokinase

but, unlike glucose, is trapped within cells. It is this trapping that

allows quantitative autoradiography to be used to measure actual

rates of glucose utilization in individual brain regions. Calculations

are made from the levels of glucose and 2-DG in plasma measured

during the experimental period and from the concentration of

radioactivity in the tissue measured autoradiographically. The details

of the experimental procedures and an extensive discussion of the

theoretical basis and mathematical derivation of the 2-DG method is

beyond the scope of this chapter, but for further discussion see

Sokoloff et al. (1977) and Sokoloff and Porrino (1986).

The most significant advantage of the 2-DG method is that it affords

a means to measure quantitatively a dynamic biochemical process in

conscious, freely moving animals whose behavior is unrestricted.

Furthermore, the 2-DG method makes use of autoradiography, which

permits both high spatial resolution so that small anatomical regions

and subregions can be identified, and the entire central nervous

system can be examined simultaneously. There are, however, several

important limitations of the method. First, it is not possible to

93
identify the specific neurotransmitters, neuromodulators, or receptor

types that are responsible for changes in brain electrical activity.

The 2-DG method measures only glucose flux. Second, it is not

possible to distinguish between inhibitory or excitatory processes

since both are energy-requiring. Third, the 2-DG method is unable to

differentiate between direct and indirect effects of given stimulus.

An entire pathway or circuit may be metabolically activated even

though the direct action of the stimulus may occur only at the origin

of the pathway. Although this may be a limitation in some instances,

as in the case of determining the primary site of action of a drug, it

is an advantage when the goal is to identify the neural circuitry that

mediates a specific behavioral response.

EFFECTS OF COCAINE ON LOCAL CEREBRAL METABOLIC

ACTIVITY

The first report characterizing the functional consequences of cocaine

administration in the brain was that by London and her colleagues

(1986). In this experiment, three doses of cocaine, 1, 10, and 30

mg/kg injected intrapetitoneally, were tested in partially restrained

rats. Restricted changes in rates of brain energy metabolism were

observed even at the highest dose studied. Effects occurred mainly

in the extrapyramidal motor system, generally considered to be

important for the production of stereotyped behavior, which was

elicited at this dose. An interesting finding in this study was the

decreased glucose utilization in the lateral habenula at the 10 and 30

mg/kg doses. Energy metabolism in the lateral habenula is decreased

following the administration of other psychostimulants such as

amphetamine (Wechsler et al. 1979; Porrino et al. 1984) and dopaminergic agonists such as apomorphine (McCulloch et al. 1982). In

addition to the habenula, the pattern of metabolic changes in other

brain areas that resulted from cocaine administration in this study

was generally similar to those patterns observed with stereotypyinducing doses of other direct and indirect dopaminergic agonists.

We took a somewhat different experimental approach in our studies of

the effects of cocaine on cerebral metabolic activity (Porrino et al.

1988). In designing our experiments, two specific factors were

considered. First, the intravenous (IV) route of administration was

chosen because the bioavailability that results from the intravenous

administration best approximates the bioavailability that results from

the routes of administration most often used in humans (Jones 1977).

Rates of local cerebral glucose utilization were measured in rats

following IV administration of three doses of cocaine (0.5, 1.0, and

94
5.0 mg/kg). The second factor was the behavioral response to IV

cocaine administration. We wanted to relate the alterations in

functional activity inferred from changes in glucose utilization to

increases in locomotor activity elicited by cocaine. Measurements of

locomotor activity were made in freely moving rats during the period

of incorporation of the radiolabeled tracer 2-DG. The time of

maximal behavioral response to cocaine was matched to the time of

maximal incorporation to insure the greatest sensitivity of the

method.

We found a highly selective pattern of distribution of alterations in

cerebral energy metabolism at the two lowest doses of cocaine tested,

0.5 and 1.0 mg/kg (figure 1). Significant increments in glucose

utilization were seen only in the medial prefrontal cortex and the

nucleus accumbens. At 1.0 mg/kg, glucose utilization was also

increased in the substantia nigra reticulata and decreased in the

lateral habenula. Metabolic activity in the rest of the 38 structures

examined was not significantly affected by cocaine administration.

Behaviorally, these doses produced small, nonsignificant increases in

locomotor activity as compared with vehicle controls.

The selectivity of effects at low doses was in striking contrast to the

effects at the highest dose tested. Following the IV administration

of 5.0 mg/kg, widespread elevations in glucose utilization were

present throughout the brain, and these were most prominent in the

extrapyramidal motor system (figure 1). Additionally, glucose use was

increased in a number of limbic structures including the hippocampus

and olfactory tubercle. In all, significant alterations in cerebral

energy metabolism were found in 20 of the 38 structures examined.

Behaviorally, animals were significantly more active following the

administration of the 5.0 mg/kg dose of cocaine than after either

vehicle or low-dose cocaine administration.

The significant correlations between locomotor activity and rates of

glucose utilization that were found in portions of the extrapyramidal

motor system, including the globus pallidus and substantia nigra

reticulata, suggest that the changes in functional activity in these

areas are a reflection of the alterations in motor function elicited by

cocaine administration. In contrast, the pattern of alterations in

local cerebral energy metabolism in the medial prefrontal cortex and

nucleus accumbens are more in concert with the role that these

portions of the mesocorticolimbic dopaminergic system are thought to

have in the mediation of cocaine’s reinforcing effects than its effects

on locomotion. The significance of the medial prefrontal cortex and

95
FIGURE 1. Diagrams illustrating the distribution of alterations in

local cerebral glucose utilization associated with the

administration of 1.0 mg/kg and 5.0 mg/kg IV, as

depicted in six frontal planes (A-F)

KEY: AC, Anterior cingulate cortex; ACC, nudeus accumbens; CP, caudate-putamen;

GP, globus pallidus; LS, lateral septum; LT, lateral thalamus; MPFC, medial

prefrontal cortex; SM, somatomotor cortex; SNC, substantia nigra pars compacta;

SNR, substantia nigra pars reticulata; VT, ventral thalamus.

SOURCE: Adapted from Paxinos and Watson (1982), Copyright 1982, Academic Press,

Australia, Ltd.

96
nucleus accumbens has been demonstrated in a number of studies.

Rats, for example, will self-administer cocaine directly into the

prefrontal cortex (Goeders and Smith 1988; Goeders, this volume), and

lesions of the nucleus accumbens can disrupt responding for the

intravenous self-administration of cocaine (Roberts et al. 1977;

Roberts et al. 1980; Koob and Hubner, this volume). The fact that

the selective metabolic effects of cocaine in these areas occurred

only at doses that are known to be reliably self-administered

intravenously by rats is further support for the nucleus accumbens

and medial prefrontal cortex as neural substrates underlying cocaine

reinforcement.

The differences in the distribution of changes in functional activity

found in the present study and those reported earlier by London and

her colleagues (1986) were perplexing. A number of factors including

rat strain, use of restrained vs. freely moving preparations, and the

route of administration may account for the discrepancies. We tested

one of these possibilities, route of administration, by comparing the

effects on local cerebral glucose utilization of an intraperitoneal dose

(10 mg/kg) of cocaine to those of an intravenous dose (1.0 mg/kg),

both of which produced similar increases in locomotor activity. As in

previous studies, rates of glucose utilization did not differ from

control values in most of the structures examined. Rates were

similarly altered in the substantia nigra reticulata (figure 2) and the

lateral habenula following either type of cocaine administration.

Intravenously administered cocaine, however, resulted in significant

changes in rates of glucose use in the nucleus accumbens (figure 2)

and the medial prefrontal cortex. These differences in the distribution of alterations in local cerebral glucose utilization are interesting

in light of recent findings by Spyraki and her coworkers (Nomikos

and Spyraki 1988; Spyraki et al. 1987), who have shown that route of

administration is a significant variable in the establishment of a

conditioned place preference in rats. Brain concentrations and the

rate at which these concentrations are achieved vary with the route

of administration and, therefore, may in turn be significant factors in

determining the local metabolic effects as well as the behavioral

consequences of cocaine administration (Balster, this volume).

METABOLIC MAPPING OF THE EFFECTS OF COCAINE ON BRAIN

STIMULATlON REWARD

lt has been suggested that abused drugs such as cocaine achieve their

reinforcing properties by activation of reward systems in the brain

(Kometsky 1985). One common approach to the study of these

97
FIGURE 2. A comparison of the effects of saline vehicle, and acute

intravenous (1 mg/kg), and intraperitoneal(10 mg/kg)

administration of cocaine on local cerebral glucose

utilization (LCGU) in the nucleus accumbens and

substantia nigm pars reticulata in rats

*p<0.05, significantly different from saline vehide control, Bonferroni t-statistic.

NOTE: Values represent means and standard errors of rates of LCGU (μ mol/100

g/min) in groups of six, four, and four rats, respectively.

9 8
systems is through brain stimulation reward, a phenomenon first

demonstrated by Olds and Milner (1954), who showed that an animal

will work in order to obtain electrical stimulation to certain brain

sites.

Cocaine has been shown both to increase rates of responding for a

constant level of intracranial stimulation (Crow 1970) and to lower

the threshold for brain stimulation reward (Esposito et al. 1978). The

specific neural circuitry involved in mediating these effects, however,

has not yet been identified. To address this question, we have used

metabolic mapping with the 2-DG method to identify those areas

functionally involved in the interaction of cocaine and brain stimulation reward (Bain et al. 1987). Two groups of animals were tested:

(1) animals self-stimulating to the medial forebrain bundle; and (2)

animals self-stimulating to the medial forebrain bundle that received

10 mg/kg cocaine intraperitoneally. This dose and regimen were

chosen because previous work has clearly demonstrated its effectiveness in enhancing brain stimulation reward (Esposito et al. 1978).

Both groups of animals receiving electrical brain stimulation responded on the average at identical rates at similar current intensity

levels. In these preliminary experiments, these data were compared

to a group of animals with electrodes implanted in the medial

forebrain bundle but not stimulated during the experiment (Porrino,

unpublished observations).

Although the experiment is not complete, an interesting picture has

emerged. In most areas, rates of local cerebral glucose utilization

were the same in the two groups of animals responding to brain

stimulation reward. Increases in metabolic activity as compared to

rates in unstimulated rats were found throughout the rostral and

caudal extent of the medial forebrain bundle and in areas such as the

nucleus accumbens, prefrontal cortex, and amygdala. Marked bilateral

increases in rates of glucose utilization were found in the olfactory

tubercle and decreases were found in the ventral tegmental area of

cocaine-treated animals as compared with rates in animals receiving

electrical stimulation alone. Metabolic activity in these areas has

been reported not to be altered following intraperitoneal cocaine

administration without brain stimulation (London et al. 1986). lt is

worth noting that the effects of the interaction of cocaine and brain

stimulation reward on cerebral metabolism in the olfactory tubercle

are quite similar to those reported with amphetamine and brain

stimulation (Seeger et al. 1984). Finally, these data suggest that

cocaine enhances brain stimulation reward by acting on circuits

different from those that mediate its effects on locomotor activity.

99
The behavioral state of the animal during cocaine administration,

then, is a significant factor in determining the anatomical distribution

of local cerebral metabolic effects.

COMPARlSON WITH OTHER PSYCHOMOTOR STlMULANTS

Psychostimulants can be divided into two separate drug classes on the

basis of their actions at dopaminergic nerve terminals (McMillen 1983;

Scheel-Kruger 1971; Shore 1978). Drugs in the amphetamine class of

stimulants act via stimulation of release of newly synthesized

dopamine from an alpha-methylpara-tyrosine sensitive pool (Chiueh

and Moore 1975b; Scheel-Kruger 1971). In contrast, drugs in the

nonamphetamine class of stimulants (e.g., cocaine or methylphenidate)

release dopamine from reserpine-sensitive vesicular stores (Chiueh and

Moore 1975a; Scheel-Kruger 1971). Although there are differences in

the biochemical actions of these drugs, there are strong similarities

in their behavioral effects (e.g., reinforcing properties, locomotoractivating properties, and stereotypy-eliciting properties), all of which

are thought to be mediated through their effects on dopaminergic

systems (Creese and lverson 1975; DeWit and Wise 1977; Roberts

et al. 1975).

There have been a number of studies examining the effects of

psychostimulants other than cocaine on brain glucose utilization. The

dose-dependent nature of cocaine’s effects on local cerebral energy

metabolism is clearly analogous to the effects of other psychostimulants on functional activity. Low doses of either amphetamine

(Porrino et al. 1984) or methylphenidate (Porrino and Lucignani 1987),

for example, result in selective increases in glucose utilization in the

nucleus accumbens (figure 3). Again, as with cocaine, there appears

to be a greater sensitivity of anatomical portions of the mesocorticolimbic dopaminergic system, as compared with components of other

dopaminergic systems, to the actions of psychostimulants.

At higher doses, parallel effects on glucose utilization and behavior

also occur. Both amphetamine (Wechsler et al. 1979; Porrino et al.

1984) and methylphenidate (Bell et al. 1982; Porrino and Lucignani

1987) produce functional changes in many of the components of the

extrapyramidal system similar to those produced by cocaine (Porrino

et al. 1988; London et al. 1986). At these doses, all three psychostimulants elicit stereotypic behavior patterns that may include

repetitive sniffing, grooming, rearing, circling, and/or gnawing (Fog

1969).

100
FIGURE 3. Comparison of the alterations in local cerebral glucose

utilization accompanying the administration of selected

psychostimulants and dopamine receptor agonist drugs

NOTE: Values are percent change from control. AMPH, amphetamine (0.5 mg/kg)

(Ponino et al. 1984); COC, cocaine (1 mg/kg) (Ponino et al. in press): MPH,

methylphenidate (2.5 mg/kg) (Porrino and Lucignani 1987); APO, apomorphine

(0.5 mg/kg) (McCulloch et al. 1982; Porrino, unpublished observations): BROM,

bromocriptine (20 mg/kg) (Pizzolato et al. 1985).

Although this dose-dependent pattern of effects within the mesocorticolimbic and nigrostriatal dopaminergic systems seems to be

typical of psychostimulants, such a pattern is not characteristic of

other dopaminergic drugs. For example, apomorphine, a direct

dopamine receptor agonist, does not significantly alter glucose

utilization in the mesocorticolimbic system. It is without effect on

such areas as the nucleus accumbens (figure 3) and prefrontal cortex

(McCulloch et al. 1982). This pattern is similar to that obtained

following the administration of bromocriptine, another dopamine

receptor agonist (Piuolato et al. 1985) (figure 3). It appears that

101
psychostimulants have a unique metabolic action in the mesocorticolimbic dopaminergic system not observed with other dopaminergic

agonist drugs.

CONCLUSIONS

The quantitative 2-DG method maps changes in functional activity

based on experimentally measured alterations in local rates of

cerebral glucose utilization and has proved useful in pharmacological

applications. The series of studies reviewed here emphasize the

complexity of the neural circuitry that mediates the effects of

cocaine in the central nervous system. In particular, they show how

variables such as dose, route of administration, and behavioral state

can determine the neurochemical response to cocaine administration.

Finally, cerebral metabolic studies of psychostimulants and other

drugs acting at dopaminergic synapses demonstrate a unique sensitivity of the dopaminergic mesocorticolimbic system to the effects

of psychostimulants. In summary, the 2-DG method can provide a

comprehensive picture of the functional involvement of various brain

areas in response to pharmacological agents such as cocaine and may

be able to furnish information about the neural circuits involved in

cocaine use and abuse.

REFERENCES

Bain, G.T.; Porrino, L.J.; Caplan, C.; and Kometsky, C. Cocaine

effects on rewarding brain stimulation as assessed by the quantitative P-deoxyglucose method. Abstr Soc Neurosci 13:1322, 1987.

Bell, R.D.; Alexander, G.M.; Schwartzman, R.J.; and Yu, J. The

methylphenidate-induced stereotypy in the awake rat: Local

cerebral metabolism. Neurology 32:377-381, 1982.

Chiueh, C.C., and Moore, K.E. Blockade by reserpine of

methylphenidate-induced release of brain dopamine. J Pharmacol

Exp Ther 193:559-563, 1975a.

Chiueh, C.C., and Moore, K.E. d-Amphetamine-induced release of

newly synthesized and stored dopamine from the caudate nucleus in

vivo. J Pharmacol Exp Ther 192:642-653, 1975b.

Creese, I., and Iverson, S.D. The pharmacological and anatomical

substrates of the amphetamine response in the rat. Brain Res

83:419-436, 1975.

Crow, T.J. Enhancement by cocaine of intracranial self-stimulation in

the rat. Life Sci 9:375-381, 1970.

102
DeWit, H., and Wise, R.A. Blockade of cocaine reinforcement in rats

with the dopamine blocker, pimotide, but not with the noradrenergic blockers phentolamine or phenoxybentamine. Can J Psychol

31:195-204, 1977.

Esposito, R.U.; Motola, A.H.D.; and Kometsky, C. Cocaine: Acute

effects on reinforcement thresholds for self-stimulation behavior to

the medial forebrain bundle. Pharmacol Biochem Behav 8:437-439,

1978.

Fog, R. Stereotyped and non-stereotyped behavior in rats induced by

various stimulant drugs. Psychopharmacology (Berlin) 14:299-304,

1969.

Goeders, N.E., and Smith, J.E. Cortical dopaminergic involvement in

cocaine reinforcement. Science 221:773-775, 1983.

Jones, R. The pharmacology of cocaine. In: Grabowski, J., ed.

Cocaine: Pharmacology, Effects, and Treatment of Abuse. National

Institute on Drug Abuse Research Monograph 50. DHHS Pub. No.

(ADM)84-1326. Washington, DC: Supt. of Docs., U.S. Govt. Print.

off., 1984. pp. 34-53.

Kadekaro, M.; Crane, A.M.; and Sokoloff, L. Differential effects of

electrical stimulation of sciatic nerve on metabolic activity in spinal

cord and dorsal root ganglion in the rat. Proc Natl Acad Sci USA

82:6010-6013, 1985.

Kornetsky, C. Brain stimulation reward: A model for the neuronal

bases for drug-induced euphoria. In: Brown, R.M.; Friedman, D.P.;

and Nimit, Y., eds. Neuroscience Methods in Drug Abuse Research.

National Institute on Drug Abuse Research Monograph 62.

Washington, DC: Supt. of Docs., U.S. Govt. Print. Off., 1985. DHHS

Pub. No. (ADM)85-1415. pp. 30-50.

London, E.D.; Wilkerson, G.; Goldberg, S.R.; and Risner, M.E. Effects

of L-cocaine on local cerebral glucose utilization in the rat.

Neurosci Lett 68:73-78, 1986.

Mata, M.; Fink, D.J.; Gainer, H.; Smith, C.B.; Davidsen, L.; Savaki, H.;

Schwartz, W.J.; and Sokoloff, L. Activity-dependent energy

metabolism in rat posterior pitultary primarily reflects sodium pump

activity. J Neurochem 34:213-215, 1980.

McCulloch, J. Mapping functional alterations in the CNS with

[

14C]deoxyglucose. In: Iverson, L; Iverson, S.; and Snyder, S.,

eds. Handbook of Psychopharmacology. Vol. 15. New York:

Plenum Press, 1982. pp. 321-410.

McCulloch, J.; Savaki, H.E.; McCulloch, M.C.; Jehle, J.; and Sokoloff,

L. The distribution of alterations in energy metabolism in the rat

brain produced by apomorphine. Brain Res 243:67-80, 1982.

McMillen, B.A. CNS stimulants: Two distinct mechanisms of action

for amphetamine-like drugs. Trends Pharm Sci 4:429-432, 1983.

103
Nomikos, G.G., and Spyraki, C. Cocaine-induced place preference

conditioning: Importance of route of administration and other

procedural variables. Psychopharmacology (Berlin) 94:119-125, 1988.

Olds, J., and Miner, P. Positive reinforcement produced by electrical

stimulation of septal area and other regions of rat brain. J Comp

Physiol Psychol 47:419-427, 1954.

Paxinos, G., and Watson, C. The Rat Brain in Stereotaxic

Coordinates. New York: Academic Press, 1982. 83 pp.

Piuolato, G.; Soncrant, T.T.; and Rapoport, S.I. lime-course and

regional distribution of the metabolic effects of bromocriptine in

the rat brain. Brain Res 341:303-312, 1985.

Porrino, L.J.; Domer, F.R.; Crane, A.M.; and Sokoloff, L. Selective

alterations in cerebral metabolism within the mesocorticolimbic

dopaminergic system produced by acute cocaine administration in

rats. Neurcpsychopharmocology 1:109-118, 1988.

Porrino, LJ., and Lucignani, G. Different patterns of brain energy

metabolism associated with high and low doses of methylphenidate:

Implications for stimulant drug action in hyperactive children. Biol

Psychietry 22:125-138, 1987.

Porrino, L.J.; Lucignani, G.; Dow-Edwards, D.; and Sokoloff, L.

Dose-dependent effect of acute administration on functional brain

metabolism in rats. Brain Res 307:311-320, 1984.

Roberts, D.C.S.; Corcoran, M.E.; and Fibiger, H.C. On the role of

ascending catecholaminergic systems in intravenous self-administration of cocaine. Pharmacol Biochem Behav 6:615-620, 1977.

Roberts, D.C.S.; Koob, G.F.; Klonoff, P.; and Fibiger, H.C. Extinction

and recovery of cocaine self-administration following 6-hydroxydopamine lesions of the nucleus accumbens. Pharmacol Biochem

Behav 12:781-787, 1980.

Roberts, D.C.S.; Zis, A.P.; and Fibiger, H.C. Ascending catecholamine

pathways and amphetamine-induced locomotor activity: Importance

of dopamine and apparent non-involvement of norepinephrine.

Brain Res 93:441-454, 1975.

Scheel-Kruger, J. Comparative studies of various amphetamine

analogues demonstrating different interactions with the metabolism

of the catecholamines in the brain. Eur J Phefmacol 14:47-59,

1971.

Schwartz, W.J.; Smith, C.B.; Davidsen, L; Savaki, H.E.; Sokoloff, L;

Mata, M.; Fink, D.J.; and Gainer, H. Metabolic mapping of

functional activity in the hypothalamo-neurohypophysial system of

the rat. Science 205:723-725, 1979.

104
Seeger, T.F.; Porrino, L.J.; Esposito, R.U.; Crane, A.M.: Sullivan, T.L.;

and Pert, A. Amphetamine effects on intracranial self-stimulation

as assessed by the quantitative 2-deoxyglucose method. Abstr Soc

Neurosci 10:307, 1984.

Shore, P.A. Actions of amfonelic acid and other nonamphetamine

stimulants on the dopamine neuron. J Pharm Pharmacol 28:855-857,

1978.

Sokoloff, L. Mapping cerebral functional activity with radioactive

deoxyglucose. Trends Neurosci 1:75-79, 1978.

Sokoloff, L. The radioactive deoxyglucose method: Theory, procedure, and applications for the measurement of local glucose

utilization in the central nervous system. In: Agranoff, B.W., and

Aprison, M.H., eds. Advances in Neurochemistry, Vol. 4. New York:

Plenum Press, 1982. pp. 7-36.

Sokoloff, L., and Ponino, LJ. Some fundamental considerations in

the application of the deoxyglucose method to pharmacological

studies. In: Kriegelstein, I., ed. Pharmacology of Cerebral

Ischemia. Amsterdam: Elsevier, 1986. pp. 65-76.

Sokoloff, L.; Reivich, M.; Kennedy, C.; DesRosiers, M.H.; Patlak, C.S.;

Pettigrew, K.D.; Sakurada, O.; and Shinohara, M. The [14C]deoxyglucose method for the measurement of local cerebral glucose

utilization: Theory, procedure and normal values in the conscious

and anesthetized albino rat. J. Neurochem 28:897-916, 1977.

Spyraki, C.; Nomikos, G.G.; and Varonos, D.D. Intravenous cocaineinduced place preference: Attenuation by haloperidol. Behav Brain

Res 26:57-62, 1987.

Wechsler, LR.; Savaki, H.E.; and Sokoloff, L. Effects of Damphetamine and L-amphetamine on local cerebral glucose utilization in the

conscious rat. J. Neurochem 32:15-22, 1979.

ACKNOWLEDGMENTS

The preparation of this manuscript was supported in part by U.S.

Public Health Service grant DA 02326 from the National Institute on

Drug Abuse (NIDA) to C. Kornetsky and NIDA Research Scientist

Award DA 00099 to C. Kornetsky.

105
AUTHORS

Linda J. Porrino, Ph.D.

Neuropsychopharmacology Laboratory

Addiction Research Center

National Institute on Drug Abuse

P.O. Box 5180

Baltimore, MD 21224

and

Laboratory of Cerebral Metabolism

National Institute of Mental Health

Bethesda, MD 20892

Conan Kornetsky, Ph.D.

Laboratory of Behavioral Pharmacology

Boston University School of Medicine

80 East Concord Street, L-602

Boston, MA 02118

Address all correspondence to:

Dr. Linda J. Porrino

Unit on Brain Imaging

Clinical Neuroscience Branch

National Institute of Neurological and

Communicative Disorders and Stroke

Building 10, Room 5N-214

9000 Rockville Pike

Bethesda, MD 20892

106
Behavioral Studies of the

Reinforcing Properties of Cocaine

Chris E. Johanson

INTRODUCTION

This chapter will describe the behavioral characteristics of cocaine

self-administration and highlight the procedures that have been used

in experimental studies to measure its reinforcing efficacy. Since

this monograph is devoted in large measure to reviewing the status of

our understanding of the neuroanatomical and neurochemical bases of

cocaine’s properties that contribute to its abuse by humans, it is

necessary to clarify the intent of including a review of behavioral

studies on cocaine’s reinforcing efficacy.

Like all psychoactive drugs, cocaine has a complex, multiple set of

pharmacological properties mediated by a number of neurochemicals

with sites of action throughout the central nervous system. Not all

of these properties are related to the reinforcing efficacy of the

drug; in addition, it is likely that the mediation of cocaine’s

reinforcing properties is due to the interaction between multiple

systems. In studying the molecular basis of cocaine’s reinforcing

effects, the influence of this complexity on behavioral indices of

reinforcing efficacy is often ignored. Including a review of behavioral studies demonstrating that cocaine self-administration is not

solely determined by cocaine’s reinforcing effects and that reinforcing

properties can be influenced by a range of behavioral variables is a

reminder to neuroscientists that any molecular mechanism proposed to

underlie cocaine’s addictive properties must be able to account for

these complexities. This chapter is also intended to acquaint

neuroscientists who are studying the molecular basis of cocaine’s

reinforcing properties with the most sophisticated approaches for

measuring these effects, so that they can design the most definitive

studies.

107
ESTABLISHMENT OF COCAINE AS A REINFORCER

One of the definitions of a positive reinforcer is the ability to maintain responding leading to its presentation. The positive reinforcing

properties of cocaine have been investigated since the early 1960s,

using drug self-administration techniques. Pickens and Thompson

(1968) showed that rats equipped with an intravenous catheter would

press a lever if that response was followed by an intravenous injection of cocaine. In a series of manipulations, these investigators

carefully demonstrated that lever pressing was a function of contingent cocaine injection. For instance, when cocaine was injected

intermittently by the experimenter instead of the animal, the rats did

not continue to lever press. When saline was substituted for cocaine

or when only a light change followed each lever press, responding

declined. Finally, when two levers were available, the rat responded

on the lever that produced cocaine even when the experimenter

switched operative levers.

In another early self-administration study (Wilson et al. 1971), rhesus

monkeys were given 4 hours of daily access to cocaine during which

each lever press resulted in a drug injection. Interestingly, the monkeys regulated their drug intake to a remarkable degree. After

training, they showed stability in their daily intake of cocaine over

periods of months. There were no indications of changes in sensitivity to cocaine’s reinforcing effects as would be indicated by an

increase (tolerance) or a decrease (supersensitivity) in its rate of

self-administration. These investigators also demonstrated the constancy of cocaine intake by changing the dose injected after each

lever press. As dose per injection was increased, the number of

injections taken by the animals decreased, resulting in an almost constant intake in drug regardless of the dose per injection.

Another type of regulation is also evident in the pattern of cocaine

self-administration. As shown in figure 1, infusions of cocaine are

equally spaced across the experimental session almost as if the drug

were being injected under the control of a clock. Stability of intake

within sessions is not a characteristic shared by many other drug

reinforcers, as shown for pentobarbital in figure 1. Even a drug such

as amphetamine, which shares many pharmacological properties with

cocaine, shows variation in intake; within sessions, amphetamine is

taken in bursts of injections with long pauses between these bursts

(Balster and Schuster 1973a). On the other hand, a variety of other

drugs that function as positive reinforcers and also have some properties in common with cocaine, including procaine (Johanson 1980),

108
FIGURE 1. Representative cumulative response records of fixed-ratio

10 performence maintained by 0.2 mg/kg cocaine (top

panel) end 0.25 mg/kg pentobarbital (bottom panel) in 8

rhesus monkey

NOTE: Ordinate: cumulative responses. Abscissa: time. Each downward deflection of

the response pen indicates a 10-second injection. The response pen resets

every 30 minutes.

cathinone (Johanson and Schuster 1981 b), and propylbutyldopamine

(Woolverton et al. 1984), also show within-session regularity in

intake. The mechanisms resulting in this type of regularity are not

currently understood. It is unlikely, however, that it is completely

attributable to any single set of neurochemical events.

What must also be explained by a complete description of the molecular actions of cocaine is the disappearance of controlled intake across

sessions when access to cocaine is no longer limited to a few hours

each day. In a study by Johanson et al. (1976), untrained rhesus

monkeys were exposed to continuous around-the-clock access to one

of a variety of psychomotor stimulant drugs. Two monkeys given

access to 0.2 mg/kg cocaine began taking drug the very first day of

its availability, and immediately intake became erratic and excessive,

resulting in severe toxicity that led to death. Similar results were

noted with other psychomotor stimulants tested, including amphetamine, methamphetamine, and diethylpropion. Therefore, it appears

that, where there are no outside restraints on the availability of

psychomotor stimulant drugs, rhesus monkeys will suddenly increase

their drug-taking behavior to the point of severe toxicity. In contrast, the intake of cocaine under conditions of limited access is

surprisingly regulated. The mechanism underlying this loss of control

is not understood, but it is possible that it is due to psychomotor

109
stimulant drugs’ ability to produce stereotypic behavior, which may

include the drug-taking response.

REINFORCING PROPERTIES OF COCAINE ACROSS

CONDlTlONS

The results of the early studies of Pickens and Thompson (1968) and

Wilson et al. (1971) have been replicated in a number of ways. What

is impressive is the generality of the phenomenon. For instance,

cocaine is self-administered by every species of animal tested,

including rats (Pickens and Thompson 1968), squirrel monkeys

(Goldberg 1973; Katz 1979), rhesus monkeys (Woods and Schuster

1968), pigtail macaques (Young and Woods 1980), baboons (Grfffiths et

al. 1975), dogs (Risner and Jones 1975), and humans (Fischman 1984).

This concordance across species, both in terms of the ability of

cocaine to function as a reinforcer as well as the characteristics of

the maintained responding (Griffrths et al. 1980; Johanson and

Schuster 1981 a), is one type of evidence of the robustness of

cocaine’s ability to function as a reinforcer. This also implies that

similar molecular substrates are shared across species and that it is

unlikely that cocaine abuse in humans is due to some specific

pathological condition.

A second type of evidence that cocaine is a robust reinforcer is that

it maintains responding regardless of its route of delivery. Although

the IV route has been used most commonly in experimental studies,

cocaine also maintains responding when delivered intragastrically

(Woolverton and Schuster 1983), by chewing or smoking (Siegel et al.

1976), and intramuscularly (Goldberg et al. 1976).

Cocaine self-administration occurs not only with a variety of species

using several routes of administration but also under a variety of

environmental circumstances. In the terminology of behavior analysis,

this can be translated into schedule contingencies, i.e., the rule that

governs the relationship between behavior and the injection of the

reinforcer. The nature of that rule has overwhelming influence on

the reinforcing properties of cocaine, as evidenced by changes in the

shape and position of its dose-response function. Such an influence

is yet another of the behavioral characteristics of cocaine that

require elucidation by any proposed molecular mechanism.

In the simplest schedule, continuous reinforcement, every response is

followed by a reinforcer. This schedule was used by both Pickens

and Thompson (1968) and Wilson et al. (1971). However, the

110
relationship between responding and reinforcer presentation may be

made more complex. A response may be reinforced on the basis of

the number of responses emitted since the termination of the previous reinforcer presentation (a ratio schedule), or on the basis of the

time elapsed since the last reinforcer presentation (an interval

schedule). Many studies have shown that cocaine maintains responding under ratio schedules (Balster and Schuster 1973a). The pattern

of responding is characterized by an initial pause followed by a high

terminal rate of responding (figure 1). Although this pattern of ratio

responding is similar to that maintained by other events, such as food

and water, the rates of responding typically found in drug selfadministration studies have been low compared to rates maintained by

food, and increases in dose per injection further decrease rates, i.e.,

the relationship between dose and rate of responding is inverse.

These low rates are most likely due to the dual actions of the drug,

as shown in figure 2. On the one hand, cocaine serves as a reinforcer that increases rate of responding, but on the other hand, the

drug has the ability to disrupt ongoing behavior temporarily, and thus

has a rate-decreasing effect. As dose is increased, the latter effects

predominate, and responding becomes suppressed. Since increased

responding under ratio schedules results in increased rates of drug

intake, the problem is particularly striking under this schedule.

Therefore, using ratio schedules to determine the potency of cocaine

as a reinforcer may be inadequate because the dependent variable

REINFORCING EFFECTS

Increases in response leading to drug

OPERANT

RESPONSE

DIRECT EFFECTS

Temporary increase or decrease in all

ongoing responses regardless of

consequence

FIGURE 2. Two types of effects that a drug injection contingent on

an operant response can exert

111
itself, namely rate of responding, is not determined solely by reinforcing properties. In fact, at the highest doses, which may have the

maximum reinforcing effect, responding is predominantly influenced by

cocaine’s nonspecific rate-decreasing effects and is often suppressed.

This problem must be taken into consideration in interpreting studies

designed to determine the underlying molecular mechanism of any

specific behavioral effect such as reinforcing properties.

In an attempt to avoid a confounded measure of reinforcing properties, other schedules have been used in cocaine self-administration

studies. These studies are also important in terms of further demonstrating the generality of cocaine’s reinforcing properties. For

instance, cocaine has been demonstrated to maintain responding under

interval schedules. An important feature of interval schedules is that

rates of responding can change considerably without affecting rate of

reinforcement. One of the first studies using an interval schedule of

cocaine injections in monkeys was conducted by Balster and Schuster

(1973b). Responding was maintained under a fixed-interval, 9-minute

schedule of cocaine injection in one component and food delivery in

the other. In addition, there was a 15-minute timeout following the

presentation of each reinforcer. Responding was well maintained, and

the pattern of gradually accelerated responding over the interval with

cocaine was similar to that maintained by food. As dose per injection increased, rate of responding increased, i.e., the dose-response

function was direct. However, in a study by Johanson (1982) using a

fixed-interval, 5-minute schedule of cocaine injection without an

intervening timeout period, the shape of the dose-response function

was no longer a direct one but was inverted U-shaped. That is, rate

of responding initially increased as dose per injection was increased,

but rate then decreased with further increases in dose. This difference was most likely due to the more frequent injections in the

Johanson study. Despite the powerful nature of the contingencies

governing reinforcer presentation controlling responding, the nonspecific rate-modifying actions of cocaine also exerted an influence.

Once again, this complicates assessments of the molecular mechanisms

underlying the reinforcing properties of cocaine that rely on the

ability to compare potencies across measures.

Several studies with cocaine have used second-order schedules as a

way of minimizing the direct effects of cocaine, to get a less confounded estimate of the drug’s reinforcing actions. In this type of

schedule, responding specified by one particular schedule is treated as

a unitary response that is itself reinforced by another schedule

(Kelleher 1966). The responding treated as a unitary response can

112
also be followed by the presentation of a brief stimulus paired with

the presentation of the reinforcer. Goldberg (1973), using squirrel

monkeys, studied responding maintained by cocaine under a fixedratio (FR) 30 schedule of stimulus presentation (2-second yellow

light), which itself was maintained under a fixed-interval (FI)

6-minute schedule of cocaine injection. This schedule is designated a

second-order FI 5-minute (FR 30:s). Under this schedule, rates of

responding for cocaine were extremely high and similar to responding

maintained by other events, such as food, under an identical schedule.

Several additional studies have been conducted with this schedule,

using both squirrel and rhesus monkeys. Typical responding has been

maintained under FI (FR) schedules by IV and IM cocaine as well as

with FR schedules of FI components (Goldberg et al. 1981; Kelleher

and Goldberg 1977; Goldberg and Kelleher 1977; Goldberg et al. 1975;

Goldberg et al. 1976).

Kelleher and Goldberg (1977) demonstrated the importance of the

brief stimuli in maintaining responding under second-order schedules.

When these stimuli were removed following the FR components but

the drug was still injected, rate declined and patterning was disrupted. lf both drug injections and the brief stimuli were removed,

responding declined even further. However, when the brief stimuli

were then reinstated without the drug, responding increased. Similar

results have been found in other studies including one with IM

cocaine (Katz 1979). The fact that both the drug and the stimuli are

determinants of the rate of responding may explain the results when

dose is manipulated with this schedule. Although there is some tendency for rate to increase with increases in dose, in general, doss

response functions are flat relative to those generated by other

schedules. Therefore, if rate of responding reflects reinforcing

properties, the strength of cocaine’s ability to control responding

does not seem to change with its magnitude under these schedule

conditions. This is not the case under other schedules (Balster and

Schuster 1973b; lglauer and Woods 1974; Johanson and Schuster 1975),

and, under second-order schedules with different parameters, the

shape of the dose-response function is an inverted U-shape (Johanson

1982).

In summary, the studies reviewed in this section demonstrate that

cocaine self-administration occurs under a variety of experimental

circumstances and is not restricted to a narrow range of conditions.

lt is clear that persistent and excessive drug-seeking behavior is

determined by an interaction between the drug’s schedule of presentation and its specific pharmacological properties. The shape and

113
position of cocaine’s dose-response function can change dramatically

as a function of the parameters of the experimental conditions. To

some extent, this dynamic quality is due to the multiple actions of

cocaine (e.g., rate-suppressing effects, stereotypic effects), so it is

important in studies that are focused on elucidating the underlying

mechanism of cocaine’s reinforcing properties to bear in mind possible

confounds of the behavioral measure. In addition, however, it is possible that the reinforcing properties of cocaine as well as other drugs

are altered by a variety of factors including behavioral ones. One

example of a behavioral effect that has a profound influence on

cocaine self-administration is level of food deprivation. lt has been

shown (de la Garza et al. 1981) that the rate of responding increases

under both a ratio and an interval schedule of cocaine injection when

rhesus monkeys are food restricted. When molecular mechanisms of

action of cocaine’s reinforcing effects are proposed, it will be essential to determine how variables that alter the behavioral measure of

cocaine’s reinforcement alter the molecular substrate.

REINFORCING EFFICACY ASSESSED USING NONRATE

MEASURES

Because of the difficulty of using indices that can be influenced by

effects of cocaine other than reinforcing properties, the development

of additional approaches has been vigorously pursued. Two other

procedures that have been used to compare different doses of cocaine

(in the absence, it is hoped, of any confounding influence) are choice

paradigms and concurrent schedules. In these procedures, responding

on different levers is maintained by different doses, and the primary

dependent variable is the relative frequency of occurrence of the

alternative responses. The actual rate at which either of these

responses is made, which can be dramatically influenced by level of

drug intake, is irrelevant and does not contribute to an assessment of

reinforcing properties. These procedures have also been used with

other reinforcers, such as food and intracranial stimulation, and have

been found to be sensitive to differences in reinforcer magnitude.

With concurrent schedules, responding is maintained by two or more

simultaneously operating schedules. In a study by lglauer and Woods

(1974), responding was maintained in rhesus monkeys under a concurrent two-lever variable interval schedule of cocaine injections with a

5-minute timeout following each injection. In this study, relative

reinforcing efficacy was evaluated by comparing relative response

frequencies on the two levers. A standard dose of cocaine (0.05 or

0.1 mg/kg) was available under a variable interval 1-minute schedule

114
on one of two levers; the dose available under an identical schedule

on the second lever (variable-dose lever) was varied to include both

higher and lower doses of cocaine. The proportion of responses

occurring on the variable-dose lever increased as the dose available

on that lever increased; in all cases, the larger of the two doses

presented for comparison was preferred.

The second procedure designed to compare reinforcing properties

involves the use of discrete choice trials. In a study by Johanson

and Schuster (1975), rhesus monkeys were given an opportunity to

choose between two doses of cocaine, and injections were followed by

a 15-minute timeout period. The number of trials during which one

option rather than the other was selected was counted and used as

the measure of reinforcing properties. As in the lglauer and Woods

(1974) study, actual rate of responding did not influence the measure

of reinforcing properties, and again it was found that higher doses of

cocaine were preferred to lower doses. Similar results have been

found by Brady and Griffiths in baboons (1977).

The results that have been obtained with both the concurrent schedule and the choice paradigm have been encouraging. The assumption

in studies of drug self-administration is that reinforcing strength or

efficacy ought to increase with dose. As indicated in the prior section, dose-response relationships with cocaine are rarely direct, and it

has been assumed that this was due to rate-modifying effects of

cocaine’s action unrelated to reinforcing properties. To the extent

that the influence of these other effects has been eliminated in

studies using procedures that do not utilize rate as a measure of

reinforcing efficacy, these procedures are likely to be useful in the

elucidation of the molecular mechanisms underlying the reinforcing

properties of cocaine.

REINFORCING PROPERTlES ASSESSED BY RESISTANCE TO

PERTURBATION

In addition to the use of nonrate procedures, other approaches for

evaluating the reinforcing efficacy of cocaine and other drug reinforcers have been developed to a limited extent. These approaches have

in common the notion that the strength of a reinforcer can be measured in direct proportion to its ability to maintain responding, even

when that responding is perturbed by an outside influence. The perturbations that have been used in experimental studies include

increasing the work required to obtain the reinforcer (increased

115
response cost), providing other mutually exclusive reinforcer alternatives, and concurrent punishment.

The influence of response cost on drug self-administration has been

evaluated using progressive-ratio schedules. In such a schedule,

responding is maintained by a drug under a ratio schedule. After

responding is well established, the number of responses required for

each drug injection is systematically increased until responding

declines to below some criterion, i.e., animals at these high ratios no

longer continue to respond in order to get drug. The ratio value

that leads to this cessation in responding is called the breaking point.

Although responding is maintained under a ratio schedule, the breaking point, not rate of responding, is used as the index of reinforcing

efficacy. It does not matter how long an animal takes to complete

the ratio (within limits) but simply whether or not it is finished.

Using this procedure, Yanagita (1973) demonstrated that breaking

point was a direct function of the dose of cocaine, as would be

expected. At the highest dose (0.48 mg/kg), animals continued

responding even when 6,400 to 12,800 responses had to be made for

each drug injection. Johanson (1975) used a combination of the

previously described choice procedure and a progressive-ratio

schedule. Under the initial condition, animals given a choice between

a low and a high dose selected the high dose. Next, the fixed-ratio

requirement necessary to produce the preferred dose was systematically increased, while the behavioral requirements for the alternative

but less preferred dose of cocaine remained the same. lt was

reasoned that, although animals prefer higher doses of cocaine over

lower doses, if the behavioral requirements for the preferred dose

were great enough, the animals would choose the alternative. In

addition, the greater the difference between the sizes of the doses of

the two alternatives, the greater the increase in ratio necessary to

alter preference. In two of the four monkeys tested, the results

were as predicted (figure 3). However, for the remaining two

monkeys, the high dose continued to be selected even when ratios

were above 300 responses per injection. Given the results found by

Yanagita (1973), it is likely that higher ratios were required to alter

preference in these two monkeys.

Progressive-ratio schedules have also been used to compare the

reinforcing properties of cocaine to other drug reinforcers. For

instance Yanagita (1973) found that cocaine’s breaking point was 2 to

16 times higher than that for methamphetamine and amphetamine.

Similar results were found by Bedford et at. (1978). Griffiths and

colleagues determined that the breaking point for cocaine was higher

116
FR VALUE

FIGURE 3. Number of choice trials in which different doses of

cocaine (drug B) were chosen over 0.05 mg/kg of

cocaine as a function of fixed-ratio requirements for

the injection of drug B

NOTE: The fixed-ratio requirement for the injection of 0.05 mg/kg cocaine was 5.

The maximum number of choice trials programmed each session was 25.

than that for other stimulant or anorectic drugs including methylphenidate, diethylpropion, chlorphentermine, and fenfluramine

(Griffiths et al. 1975; Griffiths et al. 1978). Studies using dogs have

also demonstrated that cocaine sustains responding at higher FR

values than d-amphetamine, mazindol, fenfluramine (Risner and Silcox

1981), or nicotine (Risner and Goldberg 1983). The important point

of these studies is that cocaine is more efficacious than these other

drugs as a reinforcer, and the extent of the difference can be

expressed numerically by comparing maximum breaking points.

Another type of perturbation is the availability of alternative

reinforcers, the selection of which eliminates the opportunity for

cocaine self-administration. The choice procedure developed by

Johanson and Schuster (1975) has been used in this regard to compare

different drugs to cocaine. When differences in potency are taken

117
into account, cocaine was found to be more efficacious as a reinforcer than methylphenidate (Johanson and Schuster 1975), diethylpropion

(Johanson and Schuster 1977a), and procaine (Johanson and Aigner

1981). Interestingly, a choice procedure that compared cocaine to

dl-cathinone, the active alkaloid of a plant (khat) that is chewed recreationally by inhabitants of Africa and the Middle East, found that

the two drugs had similar efficacy (Woolverton and Johanson 1984).

In addition to comparisons between cocaine and other drugs, there

have been choice studies utilizing alternative nondrug reinforcers.

For instance, monkeys preferred even low doses of cocaine to the

opportunity to have visual contact with other monkeys (Woolverton,

personal communication). Even more compelling, monkeys given a

choice between food and cocaine preferred the latter and, without

experimenter intervention, might have starved (Aigner and Balster

1978). As with the progressive-ratio and drug/drug choice studies,

these results clearly indicate that cocaine is a powerful reinforcer,

i.e., its reinforcing efficacy exceeds that of most other reinforcers.

The third approach to assessing the strength of a reinforcer is to

determine its resistance to the effects of punishment. The effects of

punishment such as electric shock and time out from positive

reinforcement on behavior controlled by a variety of events other

than drugs have been studied extensively. The degree of response

suppression is dependent upon the intensity of the punishing event

and its schedule of presentation, as well as on the time between

response and consequence. All else being equal, it would be assumed

that the greater the difficulty in decreasing, using punishment, the

self-administration of a particular drug, the greater is that drug’s

reinforcing strength.

The effects of punishment on cocaine self-administration have been

demonstrated in several studies. Grove and Schuster (1974) examined

the ability of punishment to suppress responding maintained by

cocaine injections in monkeys under a FR 1 schedule during daily

3-hour sessions. Punishment was accomplished by delivering a brief

electric shock at the onset of each injection. Responding maintained

by both 0.1 and 0.2 mg/kg decreased as a function of the intensity of

the shock. However, the degree of suppression expressed as a percent of control rates was the same for the two doses. That is,

increasing the magnitude of reinforcement did not seem to attenuate

the effects of punishment, as might be expected if one assumes that

higher doses of cocaine have greater reinforcing efficacy. This

finding, however, is difficult to interpret because the baseline rates

118
of responding maintained by the different doses of cocaine were not

the same. Because responding was maintained under a ratio schedule,

the rates maintained by the higher dose were lower.

In an attempt to eliminate the problem of rate differences, Johanson

(1977) used the discrete trial choice procedure previously described,

with added punishment contingencies. Rhesus monkeys were given a

choice between two alternatives of IV cocaine. These alternatives

were initially equal in dose, but in subsequent comparisons they

differed in magnitude. Electric shock was delivered at the onset of

the injection of one of the alternatives. When the two doses were

equal, the nonshocked alternative was chosen. For some animals, the

shocked alternative was preferred even when the dose of this alternative was only twice as high. Other animals continued to select the

nonshocked alternative. However, as the dose of the shocked alternative further increased, all animals preferred the higher dose.

Figure 4 shows the results from one of these monkeys.

SESSION

FIGURE 4. The number of injections of the higher dose (indicated

above each panel) selected by an individual monkey

during consecutive sessions

NOTE: Electric shock was delivered at the onset of each injection of this higher dose.

The lower dose was 0.1 mg/kg cocaine during the entire session. There was a

maximum of 16 choices per session.

119
All of the studies reviewed in this section clearly lead to the conclusion that cocaine has strong and robust reinforcing properties, as

indicated by its ability to continue to maintain responding even when

response cost is high, even at the expense of refusing alternative

reinforcers as important as food, and even when self-administering

cocaine is punished. All of these approaches can be utilized to compare cocaine’s reinforcing efficacy to that of other drugs, but to date

not enough data have been generated to indicate unequivocally that

any of these approaches will be successful in quantitatively differentiating reinforcing efficacy across drugs. If these approaches are

sensitive to differences in reinforcing efficacy across drugs and yield

accurate quantitative indices of these differences, these values could

be used for correlating with the effects of cocaine on various molecular systems. However, it is important to note that the results of

these studies appear to indicate that even drugs with similar pharmacological actions (e.g., cocaine, mazindol, amphetamine, procaine)

differ not only in potency but also in efficacy. How drugs with

differing efficacy on a behavioral property (reinforcing properties)

can be compared in molecular studies that assess differences in

potency is not clear.

SUMMARY

In this chapter, the behavioral characteristics of cocaine selfadministration were described. Cocaine is a powerful positive

reinforcer, and its reinforcing properties have been demonstrated in

every species tested, across an enormous range of conditions in

experimental studies with animals. To compare the reinforcing efficacy of cocaine with that of other drugs, to determine how these

effects might change under different environmental conditions, and, in

the present context, to determine their underlying molecular mechanisms require that its reinforcing properties be assessed using indices

that are not influenced by the multitude of other pharmacological

effects that are produced by cocaine. Many procedures rely on rate

of responding or levels of cocaine self-administration as a measure of

reinforcing properties, but these may not be adequate. The use of

more sophisticated approaches that utilize nonrate measures or an

assessment of the resistance of drug-maintained responding to perturbations may enable neuroscientists to design more definitive studies.

REFERENCES

Aigner, T.G., and Balster, R.L. Choice behavior in rhesus monkeys:

Cocaine versus food. Science 201:534-535, 1978.

120
Balster, R.L, and Schuster, C.R. A comparison of d-amphetamine,

I-amphetamine and methamphetamine self-administration in rhesus

monkeys. Pharmacol Biochem Behav 1:67-71, 1973a.

Balster, R.L, and Schuster E.R. Fixed-interval schedule of cocaine

reinforcement: Effect of dose and infusion duration. Exp Anal

Behav 20:119-129, 1973b.

Bedford, J.A.; Baily, LP.; and Wilson, M.C. Cocaine reinforced

progressive ratio performance in the rhesus monkey. Pharmacol

Biochem Behav 9:631-638, 1978.

Brady, J.V., and Griffiths, R.R. Drug-maintained performance and the

analysis of stimulant reinforcing effects. In: Ellinwood, E.H., and

Kilbey, M.M., eds. Cocaine and Other Stimulants. New York:

Plenum Press, 1977. pp. 599-613.

de la Garza, R.; Bergman, J.; and Hartel, C.R. Food deprivation and

cocaine self-administration. Pharmacol Biochem Behav 15:141-144,

1981.

Fischman, M.W. The behavioral pharmacology of cocaine in humans.

In: Grabowski, J., ed. Cocaine: Pharmacology, Effects, and

Treatment of Abuse. National Institute on Drug Abuse Research

Monograph 50. DHHS Pub. No. (ADM)84-1326. Washington, DC:

Supt. of Docs, U.S. Govt. Print. Off., 1984. pp. 72-91.

Goldberg, S.R. Comparable behavior maintained under fixed-ratio and

second-order schedules of food presentation, cocaine injection, or

d-amphetamine injection in the squirrel monkey. J Pharmacol Exp

Ther 186:18-30, 1973.

Goldberg, S.R., and Kelleher, R.T. Reinforcement of behavior by

cocaine injections. In: Ellinwood, E.H., and Kilbey, M.M., eds.

Cocaine and Other Stimulants. New York: Plenum Press, 1977.

pp. 523-544.

Goldberg, S.R.; Kelleher, R.T.; and Goldberg, D.M. Fixed-ratio

responding under second-order schedules of food presentation or

cocaine injection. J Pharmacol Exp Ther 218:271-281, 1981.

Goldberg, S.R.; Kelleher, R.T.; and Morse, W.H. Sewndorder

schedules of drug injection. Fed Proc 34:1771-1776, 1975.

Goldberg, S.R.; Morse, W.H.; and Goldberg, D.M. Behavior maintained

under a second-order schedule of intramuscular injection of

morphine or cocaine in rhesus monkeys. J Pharmacol Exp Ther

199:278-286, 1976.

Griffiths, R.R.; Bigelow, G.E.; and Henningfield, J.E. Similarities in

animal and human drug taking behavior. In: Mello, N.K, ed.

Advances in Substance Abuse: Behavioral and Biological Research.

Vol. 1. Greenwich, CT: JAI Press, 1980. pp. 1-90.

121
Griffiths, R.R.; Brady, J.V.; and Snell, J.D. Progressive ratio

performance maintained by drug infusions: Comparison of cocaine,

diethylpropion, chlorphentermine and fenfluramine.

Psychopharmocology 56:5-13, 1978.

Griffiths, R.R.; Findley, J.D.; Brady, J.V.; Dolan-Gutcher, K; and

Robinson, W.W. Comparison of progressive-ratio performance

maintained by cocaine, methylphenidate, and secobarbital.

Psychopharmacologia 43:61-83, 1975.

Grove, R.N., and Schuster, C.R. Suppression of cocaine selfadministration by extinction and punishment. Pharmacol Biochem

Behav 2:199-208, 1974.

Iglauer, C., and Woods, J.H. Concurrent performances: Reinforcement by different doses of intravenous cocaine in rhesus monkeys.

J Exp Anal Behav 22:179-196, 1974.

Johanson, C.E. Pharmacological and environmental variables affecting

drug preference in rhesus monkeys. Pharmacol Rev 27:343-355,

1975.

Johanson, C.E. The effects of electric shock on responding maintained by cocaine injections in a choice procedure in the rhesus

monkey. Psychopharmacology 53:277-282, 1977.

Johanson, C.E. The reinforcing properties of procaine, chloroprocaine

and proparacaine in rhesus monkeys. Psychopharmacology 67:189

194, 1980.

Johanson, C.E. Behavior maintained under fixed-interval and secondorder schedules of cocaine or pentobarbital in rhesus monkeys.

J Pharmacol Exp Ther 221:384-393, 1982.

Johanson, C.E., and Aigner, T. Comparison of the reinforcing

properties of cocaine and procaine in rhesus monkeys. Pharmacol

Biochem Behav 15:49-53, 1981.

Johanson, C.E.; Balster, R.L; and Bonese, K. Self-administration of

psychomotor stimulant drugs: The effects of unlimited access.

Pharmacol Biochem Behav 4:45-51, 1976.

Johanson, C.E., and Schuster, C.R. A choice procedure for drug

reinforcers: Cocaine and methylphenidate in the rhesus monkey.

J Pharmacol Exp Ther 193:676-688, 1975.

Johanson, C.E., and Schuster, C.R. A comparison of cocaine and

diethylpropion under two different schedules of drug presentation.

In: Ellinwood, E., and Kilbey, M.M., eds. Cocaine and Other

Stimulants. New York: Plenum Press, 1977. pp. 545-570.

Johanson, C.E., and Schuster, C.R. Animal models of drug selfadministration. In: Mello, N.K, ed. Advances in Substance Abuse:

Behavioral and Biological Research. Vol. II. Greenwich, CT: JAI

Press, 1981a. pp. 219-297.

122
Johanson, C.E., and Schuster, C.R. A comparison of the behavioral

effects of I- and dl-cathinone and d-amphetamine. J Pharmacol

Exp Ther 219:355-362, 1981b.

Katz, J.L. A comparison of responding maintained under second-order

schedules of intramuscular cocaine injection or food presentation in

squirrel monkeys. J Exp Anal Behav 32:419-431, 1979.

Kelleher, R.T. Conditioned reinforcement in second-order schedules.

J Exp Anal Behav 9:475-485, 1966.

Kelleher, R.T., and Goldberg, S.R. Fixed-interval responding under

second-order schedules of food presentation or cocaine injection.

J Exp Anal Behav 28:221-231, 1977.

Pickens, R., and Thompson, T. Cocaine-reinforced behavior in rats:

Effects of reinforcement magnitude and fixed-ratio size.

J Pharmacol Exp Ther 161: 122-129, 1968.

Risner, M.E., and Goldberg, S.R. A comparison of nicotine and

cocaine self-administration in the dog: Fixed-ratio and progressiveratio schedules of intravenous drug infusion. J Pharmacol Exp Ther

224:319-326, 1983.

Risner, M.E., and Jones, B.E. Self-administration of CNS stimulants

by dog. Psychopharmacologia 43:207-213, 1975.

Risner, M.E., and Silcox, D.L Psychostimulant self-administration by

beagle dogs in a progressive-ratio paradigm. Psychopharmacology

75:25-30, 1981.

Siegel, R.K; Johnson, C.A.; Brewster, J.M.; and Jarvik, M.E. Cocaine

self-administration in monkeys by chewing and smoking. Pharmacol

Biochem Behav 4:461-467, 1976.

Wilson, M.C.; Hitomi, M.; and Schuster, C.R. Self-administration of

psychomotor stimulants as a function of unit dosage.

Psychopharmacologia 22:271-281, 1971.

Woods, J.H., and Schuster, C.R. Reinforcement properties of

morphine, cocaine, and SPA as a function of unit dose. Int J

Addict 3:231-237, 1968.

Woolverton, W.L.; Goldberg, L.I.; and Ginos, J.Z. Intravenous selfadministration of dopamine receptor agonists by rhesus monkeys.

J Pharmacol Exp Ther 230:678-683, 1984.

Woolverton, W.L., and Johanson, C.E. Preference in rhesus monkeys

given a choice between cocaine and d,/-cathinone. J Exp Anal

Behav 41:35-43, 1984.

Woolverton, W.L., and Schuster, C.R. lntragastric self-administration

in rhesus monkeys under limited access conditions: Methodological

studies. J Pharmacol Methods 10:93-106, 1983.

Yanagita, T. An experimental framework for evaluation of dependence liability in various types of drugs in monkeys. Bull Narc

25:57-64, 1973.

123
Young, A.M., and Woods, J.H. Behavior maintained by intravenous

injection of codeine, cocaine, and etorphine in the rhesus macaque

and the pigtail macaque. Psychapharmacology 70:263-271, 1980.

AUTHOR

Chris E. Johanson, Ph.D.

Present address:

Department of Psychiatry

Uniformed Services University

of the Health Sciences

4301 Jones Bridge Road

Bethesda, MD 20814

Department of Psychiatry

The University of Chicago

5641 South Maryland Avenue

Chicago, IL 60687

124
Pharmacological Modifications

of Cocaine and Opioid SelfAdministration

Gail Winger

INTRODUCTION

Two types of pharmacological treatment of drug abuse have received

the most scientific consideration. One is the use of agonists to

suppress drug self-administration, the other is the use of antagonists

to block drug effects. Agonist treatment approaches for opioid abuse

are typified by the use of methadone. Antagonist treatment

approaches have more promise under some circumstances, but early

attempts to use cyclazocine or naltrexone for opioid abuse were

unsuccessful, due primarily to lack of client acceptance of these

drugs (Archer 1981; Renault 1981).

Treatment of cocaine abuse with agonists such as methylphenidate has

also been attempted, but with little success except in patients with

attention deficit disorders (Gawin 1985). To my knowledge, use of

dopamine antagonists to treat cocaine abusers has not been attempted, perhaps because these agents can produce serious and permanent motor disorders, unpleasant subjective effects, or increases

rather than decreases in cocaine self-administration in experimental

animals.

This review discusses some of the animal literature on modification of

opioid agonists or opioid antagonists and compares these data to the

effects of stimulants or dopamine antagonists on behavior maintained

by cocaine administration. Since our understanding of receptor interactions in the opioid system is much clearer than is our understanding of the interactions between cocaine and the dopamine receptor

system, the opioid system may serve as a model of agonist-antagonist

interactions in the context of drug self-administration.

125
In using either agonist or antagonist administration to decrease drug

self-administration in an animal model, it is important to produce a

specific decrease in behavior maintained by the drug. Behaviors

maintained by other reinforcers should be left relatively intact by

these manipulations, since disruption of behavior in general would be

unacceptable in a treatment program for human drug abusers. There

are several ways in which specificity of action can be assessed

experimentally. One approach is to compare data obtained in animals

whose behavior is maintained by food with data from a separate

group of animals whose responding is maintained by drug. If similar

rates and patterns of behavior develop in both groups of subjects,

comparison of the effect of drug administration on behavior maintained by different reinforcers is quite straightforward. Alternatively, food and drug can be used to maintain responding in a single

group of subjects. Food and drug may be concurrently available or

each reinforcer may be sequentially available, usually by using a

multiple schedule of food- and drug-reinforced responding. Thus, the

effects of pretreatment drug, either agonist or antagonist, can be

seen on behavior maintained by both reinforcers.

AGONIST ADMINlSTRATlON

The effects of opioid agonists on opioid-reinforced responding have

been studied using concurrent or multiple schedules of food- or drugreinforced responding. John Carney, in his 1976 doctoral thesis

(Carney, unpublished), used a multiple schedule of reinforcement, in

which signalled periods of food availability alternated with differentially signalled periods of codeine availability. There were five food

components and four codeine components in a session, presented in

alternation. Each component lasted until 15 reinforcers (either food

or codeine) had been earned on a fixed-ratio-30 (FR 30) schedule, or

until 15 minutes had passed. The dose of codeine used to maintain

behavior in the periods of drug availability was 0.1 mg/kg/injection, a

dose that was high enough to result in decrements in codeinereinforced responding that increased progressively throughout the

session. Interestingly, even when rates of codeine-maintained

responding were near zero, when the light signalling food availability

was illuminated, the monkeys immediately resumed food-reinforced

responding at a high rate. Thus, codeine appeared to be selectively

reducing rates of responding maintained by the drug itself.

Administration of a number of opioid agonists, including morphine,

etorphine, and levorphanol, produced decrements in codeine-maintained responding at doses that produced little change in

126
food-maintained responding. Although this suggested a selective

action of opioid agonists on opioid-maintained responding, the rates

of responding maintained by food were considerably higher than rates

of responding maintained by codeine. lf the opioid agonists were

simply suppressing low rates of responding selectively, the apparent

differential effect of drug due to reinforcer type could be spurious.

Camey did a rate-dependent analysis of the effects of the agonists

on different rates of responding maintained by codeine and food and

concluded that rate differences alone could not account for the

differing effects of opioids on codeine- and food-maintained

responding.

Even more convincing data on a selective effect of opioid agonists on

opioid-maintained responding were reported by Griffiths et al. (1976).

These investigators provided baboons with a choice between food and

heroin during eight daily trials. The dose of heroin was selected so

that the animals typically selected drug on three of the trials, and

food on the remaining five trials. Administration of 8.3 mg/kg/day

methadone produced a decrement in choice of heroin trials and a

corresponding increase in selection of food.

With the exception of the study reported by Mello et al. (1983) in

which methadone did not affect self-administration of dilaudid, the

data are consistent with respect to the effects of opioid agonists on

opioid self-administration. There appears to be a selective suppression of opioid-maintained responding by opioid agonists that is not

necessarily accompanied by a similar suppression in food-maintained

responding. Thus, the evidence we have concerning the effectiveness

of methadone in suppressing opioid administration in human opioid

abusers is, for the most part, supported by the data obtained in

animals.

There are fewer published studies of the effects of cocaine agonists

on cocaine se&administration. Herling et al. (1979) evaluated the

effects of cocaine, amphetamine, and pentobarbital on behavior

maintained by cocaine in one group of monkeys and by food in a

second group. The schedule of reinforcement was the same in both

groups. lt was a second-order schedule in which a brief light

stimulus was presented following every 10th response, and the 19th

response following a fixed interval of 5 minutes was reinforced with

delivery of food or cocaine, accompanied by the light stimulus. Rates

of responding maintained by food or cocaine were variable from

monkey to monkey, but overlapped to a considerable degree. The

effects of each of the three pretreatment drugs were independent of

127
the reinforcer delivered. Rates of responding maintained by food or

cocaine were suppressed to an equal degree by similar doses of either

cocaine, d-amphetamine, or pentobarbital.

Although these data suggest that the rate-decreasing effects of cocaine or amphetamine on cocaine-maintained responding may not be

selective for cocaine, there are other data that indicate that dopamine

agonists may have a selective effect on cocaine-maintained responding. Woolverton and Kleven (1987) presented such evidence

using behavior maintained under a multiple schedule of food- and

cocaine-reinforced responding. The schedule was similar to that used

by Camey (unpublished) in which periods of food availability alternated with periods of drug availability. In Woolverton’s study, however, a single period of cocaine-reinforced responding was placed

between two periods of food-reinforced responding. Rates of responding under all conditions were high, and there was little difference in rates maintained by food and those maintained by cocaine.

Under these conditions, constant infusions of the dopamine agonist

bromocriptine produced a selective decrement in rates of responding

maintained by cocaine, while leaving rates maintained by food relatively intact. This very provocative report was tempered by the

additional information that the chronic infusion of bromocriptine

produced toxic effects of increased activity, stereotyped behavior, and

preconvulsive signs in some of the monkeys, and the infusions could

not be continued.

The primary difference between the effects of opioid agonists on

opioid self-administration and those of dopamine agonists on cocaine

self-administration is that the opioid agonists produce selective

decreases in opioid-maintained responding in the absence of clear

toxic effects on the animal. The dopamine agonist bromocriptine was

also able to suppress selectively cocaine-maintained responding but

produced marked toxic effects. Clearly, the data with both types of

drug reinforcers are very limited, and further study remains to be

done before it is clear what, if any, differences exist in these two

systems. Among other things, the current data should be replicated,

the effects of other dopamine agonists must be evaluated, and

comparisons should be made between the effects of acute and chronic

administration of the agonists in question.

In designing an agonist that might be capable of suppressing behavior

maintained by cocaine, it is important to keep in mind the aspects of

methadone that may be related to its success in the treatment of

128
opioid abuse. It has a sustained effect following a single administration, it has a slow onset of action that may decrease the likelihood of its having strong reinforcing effects in its own right, it has

few toxic effects on chronic administration, and it can be given for

sustained periods of time without endangering the individual. None

of these aspects is true of dopamine agonists such as bromocriptine.

Although it may be possible to identify or synthesize dopamine

agonists with long durations of action and/or slow onsets of action,

it may not be possible to identify a dopamine agonist that either does

not have or does not develop toxic effects with chronic administration. Tolerance to the effects of dopamine agonists has not been

clearly defined. In the case of some stimulants such as d-amphetamine, which may have a dopamine agonist component of action,

sensitivity to some actions has been described (Robinson and Becker

1986). Although it is much too soon to tell whether dopamine

agonists will have some use in the treatment of cocaine abuse, it

appears to this author that the problem of toxic effects on chronic

administration will be the most difficult to overcome.

ANTAGONIST ADMINISTRATlON: SELF-ADMINISTRATlON OF

SINGLE DOSES

The effects of antagonist administration on opioid self-administration

have been described in a relatively small number of experiments.

Using the schedule of alternating food- and 0.1 mg/kg/injection

codeine-maintained responding described above, Carney (unpublished)

showed that naltrexone produced marked alterations in codeinemaintained responding at doses that did not markedly alter foodmaintained responding. The codeine-selective nature of the effects of

naltrexone was most obvious at a dose of 0.32 mg/kg given prior to

an experimental session. Rates of behavior maintained by codeine

were almost completely eliminated, whereas rates of behavior maintained by food were relatively unaltered. Lower doses of naltrexone,

0.003 and 0.03 mg/kg, produced marked increases in rates of

responding maintained by codeine and increases in the amount of drug

self-administered. These doses also had little effect on behavior

maintained by food.

In support of Carney’s findings of increases in opioid selfadministration following administration of an antagonist, Griffiths

et al. (1976) reported a dose-related increase in choice of heroin over

food in baboons chronically infused with naltrexone.

129
The increase in opioid-maintained responding following administration

of opiate antagonists is much like the effects of dopamine antagonists

on cocaine-maintained responding. A recent review of the literature

on the effects of antagonists on cocaine self-administration and discrimination (Woods et al. 1987) indicated that, when dopamine

antagonists were given prior to opportunities for cocaine selfadministration, rates or amounts of cocaine self-administration were

increased. The dopamine antagonists used included haloperidol in

both rats and monkeys: chlorpromazine in both rats and monkeys;

pimozide, sulpiride, thioridazine, and flupenthixol in rats; and

pimozide in both dogs and squirrel monkeys.

Although these studies of cocaine and codeine self-administration

indicated that antagonists produced increases in behavior maintained

by these drugs, in most cases only a single dose of self-administered

drug was used. It is difficult to make generalizations about the

effects of antagonists on the reinforcing effects of agonists under

these circumstances. The general pattern of drug self-administration

under typical fixed-ratio schedules of reinforcement is an inverted

U-shape (e.g., Young and Woods 1980; Winger and Woods 1985). lf

antagonists act to shift this curve to the right in a parallel fashion,

then doses on the ascending limb of the inverted U-shaped curve

would maintain lower rates of responding following antagonist

administration, and doses on the descending limb would maintain

higher rates of responding following antagonist administration. We

have argued previously (Woods et al. 1987) that it is the ascending

limb that reflects most accurately the reinforcing function of cocaine

and other drugs, with the descending limb being modulated predominantly by the rate-decreasing effects of the drug. Thus, it is when

the ascending limb is shifted by pretreatment with potential

antagonists that a specific effect on reinforcing function can be

indicated most clearly. In studies where rates of cocaine- or opioidmaintained responding were increased by pretreatment with an

antagonist, the dose of the reinforcing drug was almost always

sufficiently high to be on the descending limb of a dose-effect curve,

had one been obtained. It is thus not surprising to find consistent

increases in behavior following administration of antagonists.

Unfortunately, these experiments did not reveal much about the

effects of antagonist administration on the reinforcing effects of the

agonists, as would have been shown if lower doses of the agonists

had been evaluated.

130
ANTAGONIST ADMINISTRATlON: MULTIPLE SELF-ADMINlSTERED

DOSES

The effects of infusions of naltrexone on self-administration of a

range of doses of morphine was evaluated by Harrigan and Downs

(1978). In the absence of naltrexone, behavior maintained by

morphine occurred at a variety of rates, depending on the dose of

morphine available. An inverted U-shaped curve relating dose to

rates of responding was observed. This curve was shifted to the

right with a slightly decreased maximum by an infusion of 1 μg/kg/hr

of naltrexone. An infusion rate of 20 μg/kg/hr of naltrexone produced an overall decrement in responding maintained by all tested

doses of morphine.

A similar pattern of nonsurmountable antagonism of opioid selfadministration by naltrexone was observed by Herling (1981). He

evaluated the effects of naltrexone on rates of food- and codeinemaintained responding, using a schedule of alternating periods of

food and codeine availability. A dose of naltrexone of 0.003 mg/kg

produced a slight shift to the right in both the ascending and

descending limbs of the codeine dose-rate function. A dose of 0.03

mg/kg naltrexone produced a further shift to the right, as would be

expected if naltrexone were antagonizing both the rate-reinforcing

and rate-decreasing effects of codeine. However, when the dose of

0.03 mg/kg naltrexone was administered on two further occasions,

rates of codeine-maintained responding were decreased and flattened

rather than being shifted in a parallel fashion to the right. Importantly, these doses of naltrexone had little effect on food-maintained

responding. In fact, administration of naltrexone was able to

antagonize the suppression of food-maintained responding that

developed following self-administration of fairly high doses of

codeine.

As an aside, it may be that the nonsurmountable effect of naltrexone

on opioid-maintained responding is due primarily to the nature of the

naltrexonsagonist interaction. Bertalmio and Woods (1987) have

shown parallel shifts to the right in behavior maintained by alfentanil

by administration of 0.1 and 1.0 mg/kg of the opiate antagonist

quadazocine. No tendency for a downward shift in the alfentanil

dose-rate function was observed over frequent administration of

different doses of quadazocine.

Using different doses of self-administered agonists to evaluate the

effects of antagonists has also been shown to be important with the

131
dopamine antagonist-cocaine interaction. One of the very few

exceptions to the finding of increases in cocaine self-administration

following administration of a dopamine antagonist was that reported

by Herling and Woods (1980). In this study, behavior was maintained

by either food in one group of animals or cocaine in another group

of animals. Reinforcement was delivered on a second-order schedule

for both food and cocaine; rates, as described earlier for this type of

schedule, were variable among animals, regardless of the reinforcer

used. Administration of chlorpromazine produced only decreases in

rates of behavior maintained by food. In the case of cocaine, however, the effects of chlorpromazine were highly dependent on the

dose of cocaine use to maintain the behavior. The effects of chlorpromazine on behavior maintained by low doses of cocaine were

identical to the effects of chlorpromazine on behavior maintained by

food: rates were decreased across a range of chlorpromazine doses.

With higher doses of cocaine maintaining behavior, however, the

effects of chlorpromazine were to increase rates of responding

maintained by cocaine. Interestingly, under this schedule of reinforcement, increasing doses of cocaine did not produce consistent

changes in rates of responding, although two of the four monkeys

showed increases in rates followed by decreases in rates, as the dose

of cocaine was increased. The authors interpreted their findings as

indicating a mutually antagonistic interaction between cocaine and

chlorpromazine. Whereas chlorpromazine was able to antagonize the

rate-maintaining and rate-decreasing effects of cocaine, cocaine

(when delivered in sufficiently high doses), was also able to reverse

the rate-decreasing effects of chlorpromazine.

Although the typical inverted U-shaped function relating dose of

cocaine to rate of responding was not shown by all animals in the

preceding study, such a curve was obtained (Herling, unpublished) in

a study evaluating the reinforcing effects of food and several doses

of cocaine using a fixed-ratio schedule of reinforcement. In this

series of experiments, periods of food-maintained responding alternated with periods of cocaine-maintained responding. As shown in

figure 1, the effects of various doses of chlorpromazine on rates of

responding maintained by increasing doses of cocaine was to shift, in

a fairly selective way, the ascending limb of the cocaine dose-rate

curve. Interestingly, the rate-decreasing effects of cocaine, as

indicated by the descending limb of the curve, were not modified

nearly as much. This is in contrast to virtually all of the previous

reports of a rate-increasing effect of chlorpromazine on cocaine selfadministration. The reason for the lack of effect of chlorpromazine

on the descending limb of the curve in this particular study is not

132
COCAINE (mg/kg/injection)

FIGURE 1. Effects of chlorpromazine on behavior maintained by

various doses of cocaine

NOTE: Abscissae: Dose of intravenous cocaine (mg/kg/inj) available on an FR 30

schedule of reinforcement. Ordinates: Rates of responding in responses per

second. Closed circles are the rates of responding maintained by cocaine

following pretreatment with saline. The other symbols are rates maintained

following pretreatment with 0.03 mg/kg chlorpromazine (open circles), 0.1 mg/kg

chlorpromazine (hexagons), 0.18 mg/kg chlorpromazine (inverted triangles), and

0.3 mg/kg chlorpromazine (closed triangles). The numbers 737, 1094, and 778

refer to individual monkeys.

SOURCE: Herling 1980, Copyright 1980, Seymore Herling.

133
understood. These data indicate, however, that chlorpromazine,

administered under these experimental conditions, could modify the

reinforcing effects of cocaine, as indicated by shifts to the right in

the ascending limb of the cocaine dose-effect curve.

The doses of chlorpromazine that were effective in decreasing

cocaine-maintained responding also produced decreases in foodmaintained responding in the same monkey. Thus, under the

particular experimental circumstances described, chlorpromazine was

effective in modifying the reinforcing effects of cocaine, but only at

doses that also decreased rates of food-maintained responding. This

contrasts with data obtained under very similar circumstances in

which, as described above, naltrexone suppressed codeine-maintained

responding but left food-reinforced responding unaffected.

CONCLUSIONS

The contrast between the effects of dopamine agonists on cocaine

self-administration and the effects of opioid agonists on opioid selfadministration is in keeping with the differences seen in studies with

antagonists. In the case of dopamine agonist and antagonist administration, it appears difficult to find doses that alter cocaine selfadministration without producing unacceptable effects on other

behavior. The agonists appear to produce toxic effects in the form

of increases in stereotyped behavior, and the antagonists produce

decreases in food-maintained responding. Neither of these kinds of

effects has been reported in carefully controlled studies within the

opioid system. Thus, whereas the dopamine agonist bromocriptine

appears to be able to produce selective alterations in cocainemaintained responding, and the dopamine antagonist chlorpromazine

appears able to produce selective effects on the reinforcing effects of

cocaine, neither of these drugs represents a satisfactory approach to

treatment of cocaine abuse.

These experiments represent only the first steps in evaluation of

pharmacological modification of cocaine self-administration. There

are some reports that antidepressant drugs, e.g., desipramine (Gawin

and Kleber 1984) and trazodone (Small and Purcell 1985) appear to

have some use in the clinic in reducing cocaine abuse. These drugs

must be evaluated in experimental animals using self-administration

studies to help us determine what types of experimental models and

situations are sensitive to the same types of manipulations that

appear effective in man. With this information, it may be possible

134
to determine the mechanism of action of these antidepressant medications, and perhaps understand more about how cocaine produces its

reinforcing effects.

REFERENCES

Archer, S. Historical perspective on the chemistry and development of

naltrexone. In: Willette, R.E., and Bamett, G., eds. Narcotic

Antagonists: Naltrexone Pharmacochemistry and Sustained-Release

Preparations. National Institute on Drug Abuse Research Monograph 28. DHHS Pub. No. (ADM)81-902. Washington, DC: Supt. of

Docs., U.S. Govt. Print. Off., 1981. pp. 3-10.

Bertalmio, A.J., and Woods, J.H. Alfentanil reinforcement is mediated

by the mu opioid receptor: Apparent pA2 analysis. Fed Proc

46:546, 1987.

Camey, J.M. Selective modulation of codeine-reinforced responding

in rhesus monkeys. Unpublished doctoral thesis, University of

Michigan, 1976.

Gawin, F.H., and Kleber, H.D. Cocaine abuse treatment. Arch Gen

Psychiatry 41:903-909, 1984.

Gawin, F.H.; Riordan, C.; and Kleber, H.D. Methylphenidate treatment

of cocaine abusers without attention deficit disorder: A negative

report. Am J Drug Alcohol Abuse 11:193-197, 1985.

Griffiihs, R.R.; Wurster, R.M.; and Brady, J.V. Discrete-trial choice

procedure: Effects of naloxone and methadone on choice between

food and heroin. Pharmacol Rev 27:357-365, 1976.

Harrigan, S.E., and Downs, D.A. Continuous intravenous naltrexone

effects on morphine self-administration in rhesus monkeys.

J Pharmacol Exp Ther 204:481-486, 1978.

Herling, S. An analysis of specificity of drug-induced changes in

drug-reinforced responding. Unpublished doctoral thesis, University

of Michigan, 1980.

Herling, S. Naltrexone effects on food- and codeine-maintained

responding in rhesus monkeys. Eur J Pharmecol 73:41-49, 1981.

Herling, S.; Downs, D.A.; and Woods, J.H. Cocaine, d-amphetamine,

and pentobarbital effects on responding maintained by food or

cocaine in rhesus monkeys. Psychopharmacology (Berlin) 64:261-269,

1979.

Herling, S., and Woods, J.H. Chlorpromazine effects on cocainereinforced responding in rhesus monkeys: Reciprocal modification

of rate-altering effects of drugs. J Pharmacol Exp Ther 214:354-

361,1980.

135
Mello, N.K.; Bree, M.; and Mendelson, J.H. Comparison of buprenorphine and methadone effects on opiate self-administration in

primates. J Pharmacol Exp Ther 255:378-386, 1983.

Renault, P. Treatment of heroindependent persons with antagonists:

Current status. In: Willette, R.E., and Bamett, G., eds. Narcotic

Antagonists: Naltrexone Pharmacochemistry and Sustained-Release

Preparations. National Institute on Drug Abuse Research Monograph 28. DHHS Pub. No. (ADM) 81-902. Washington, DC: Supt.

of Docs., U.S. Govt. Print. Off., 1981. pp. 11-22.

Robinson, T.E., and Becker, J.B. Enduring changes in brain and

behavior produced by chronic amphetamine administration: A

review and evaluation of animal models of amphetamine psychosis.

Brain Res Rev 11:157-198, 1986.

Small, G-W., and Purcell, J.J. Trazodone and cocaine abuse. Arch

Gen Psychiatry 42:524, 1985 (Letter).

Winger, G., and Woods, J.H. Comparison of fixed-ratio and progressive-ratio schedules of maintenance of stimulant drug-reinforced

responding. Drug Alcohol Depend 15:123-130, 1985.

Woods, J.H.; Winger, G.; and France, C.F. Reinforcing and discriminative stimulus effects of cocaine: Analysis of pharmacological

mechanisms. In: Fisher, S.; Raskin, A.; and Uhlenhuth, E.H., eds.

Cocaine: Clinical and Biobehavioral Aspects. New York: Oxford

University Press, 1987. pp. 21-85.

Woolverton, W., and Kleven, M.S. Effects of bromocriptine and

desmythlimipramine on cocaine self-administration in rhesus

monkeys. Paper presented at the International Study Group

Investigating Drugs As Reinforcers Scientific Meeting, Philadelphia,

PA, June 1987.

Young, A.M., and Woods, J.H. Behavior maintained by intravenous

codeine, cocaine, and etorphine in the rhesus macaque and the

pigtail macaque. Psychopharmacology (Berlin) 70:263-271, 1980.

AUTHOR

Gail Winger, Ph.D.

Assistant Research Scientist

Department of Pharmacology

University of Michigan Medical School

M6322 Medical Services Building I

Ann Arbor, Ml 48109

136
Reinforcement Pathways for Cocaine

George F. Koob and Carol B. Hubner

INTRODUCTION

Cocaine as a Psychomotor Stimulant

Cocaine produces effects similar to many other drugs classified as

psychomotor stimulants. For example, cocaine possesses both

anorectic and psychostimulant properties and, as a result, has for

centuries been used to suppress hunger and fatigue (Angrist and

Sudilovsky 1976). Clinical studies assessing its subjective effects

have demonstrated that cocaine has stimulant actions and produces

euphoria (Fischman et al. 1983). In animals, cocaine acutely increases

motor activity (Groppetti et al. 1973), decreases food intake

(Groppetti et al. 1973), has stimulantlike actions on operant behavior

(Spealman et al. 1977), enhances conditioned responding (Spealman

et al. 1977), decreases threshold for reinforcing brain stimulation

(Kornetsky and Esposito 1981), and readily acts as a reinforcer for

drug self-administration (Pickens and Thompson 1968). Cocaine

shares most of the psychomotor stimulant effects of low doses of

amphetamine except that it is shorter acting and less potent (Simon

1973). At higher doses, cocaine acutely produces the intense

stereotypy associated with amphetamine or apomorphine but is much

less potent (Randrup and Munkvad 1970) and may be less effective

(Simon 1973).

Cocaine is also a local anesthetic and has convulsant properties

(Matsuzaki 1978). With chronic administration, these convulsant

properties appear to become sensitized as do the motor effects of

cocaine (Post et al. 1976). This sensitization may be analogous to

“kindling” associated with repeated electrical brain stimulation.

137
Neuropharmacology of Cocaine

The local anesthetic properties of cocaine are not thought to be

important for the production of its acute psychological effects, since

similar subjective effects cannot be observed with other local

anesthetics (Fischman et al. 1983). It is now well established that

cocaine has important effects on monoamine metabolism. Cocaine

blocks norepinephrine and dopamine reuptake and increases catecholamine turnover (Groppetti et al. 1973). Cocaine also blocks the uptake

of tryptophan, thereby presumably reducing serotonin synthesis

(Knapp and Mandell 1972) and turnover (Friedman et al. 1975).

Impetus for a critical role for dopamine in the psychomotor stimulant

effects of cocaine was the observation that the locomotor activation

produced by cocaine could be blocked by 6-hydroxydopamine

(6-OHDA) lesions of the nucleus accumbens (Kelly and lversen 1976).

In that study, the lesion effect was thought to be largely due to

dopamine depletion, since norepinephrine forebrain depletions were

minimized by pretreatment with desmethylimipramine. Furthermore,

similar lesions block the locomotor stimulant effects of d-amphetamine but not of caffeine or scopolamine (Joyce and Koob 1981),

heroin, or corticotropin-releasing factor (Swerdlow and Koob 1985;

Vaccarino et al. 1986).

COCAINE AND REWARD THRESHOLD

Substrates of Reward-Intracranial Self-Stimulation

The discovery that electrical stimulation of particular areas of the

brain could reinforce behavior provided a unique means to assess the

motivation to respond to hedonic stimuli. Indeed, the discovery of

intracranial self-stimulation (ICSS) by Olds and Milner (1954)

suggested a “short-circuiting” of the reinforcement process. ICSS is

typically obtained from most regions of the limbic system. Although

electrodes placed in a midbrain-forebrain system (the medial forebrain

bundle), which courses through the lateral hypothalamus, produce the

highest rates of responding, ICSS has also been produced in regions

as far removed from the classical limbic system as the cerebellum and

nucleus solitarius.

The potency of ICSS as a reinforcer led to its rapid use as a tool to

measure activity in the brain “reward” systems, but also led to many

problems of measurement and interpretation. One of the earliest

measures of brain stimulation reward was the absolute rate of

138
responding. Here an experiment consisted of placing an animal in the

test situation for a specified period of time using a specified suprathreshold current intensity. Testing was continued until response

rates had stabilized from session to session. Generally, this rate of

responding was taken to reflect the reinforcing value of the stimulus

delivered. Any treatment that increased the rate of responding was

thought to reflect a facilitation of reward or a lowering of threshold

for reward. This principle, where the actual value of the reinforcer

would be directly related to the output of the organism to obtain said

reinforcer, has a theoretical basis in a motivational theory (Hemstein

1971).

One means of evaluating threshold for brain stimulation, or by

extrapolation measuring “hedonia,” has been to establish within each

test session the point at which the rat fails to consistently respond.

Not unlike psychophysical tests, rats are systematically subjected to

an ascending or descending series of rate-intensity functions. What

typically results is a sigmoidal function whose slope increases the

closer one approaches the low threshold areas of midbrain/lateral

hypothalamus (figure 1). Shifts of this function to the right or left

without changes in the maximal rate of responding presumably

reflect changes in threshold for rewarding brain stimulation.

More recently, Kornetsky and associates have exploited the highincentive, low-drive properties of ICSS to develop a rate-independent

measure of reward (Kornetsky and Esposito 1981). Rate measures per

se can lead to misleading results when attempting to measure

reinforcement threshold. For example, in an early study, Valenstein

(1964) showed that rats allowed a choice between brain stimulation in

two different sites showed a preference for the site that maintained

the lower rate of responding. Thus, rate-independent threshold

measures for a given reinforcer may be a more valid measure of the

motivation to respond for a reinforcer.

For the Kornetsky threshold procedure, trials begin with the delivery

of a noncontingent 0.5-second stimulus. If the animal responds by

turning a wheel manipulandum one-quarter turn within 7.5 seconds of

the onset of the “priming stimulus,” a second stimulus is delivered

that is identical to the first noncontingent stimulus. The intensity of

the stimulus (value of the reinforcer) is varied according to a

psychophysical procedure where stimuli are presented in an alternating descending and ascending series with five trials presented at

each intensity level. The threshold value for each series is

139
CURRENT INTENSITY LEVEL

FIGURE 1. Rate vs. intensity function for intracranial se/fstimulation in rats

NOTE: Eight rats had electrodes placed in the lateral hypothalamus (LH), and seven

rats had electrodes aimed at the locus coeruleus (LC). Each curve represents

one session with a descending sequence. Currents ranged from 5 to 40μ A

(60 Herz AC RMS) for the LH and 15 to 50μA for the LC.

defined as the midpoint in current intensity between the level at

which the animal made three or more correct responses out of five

stimulus presentations, and the level where fewer than three correct

responses were made (Marcus and Kornetsky 1974). This “rate-free”

measure of reward has proved sensitive to pharmacological manipulation, which is consistent with the hypothesis that it reliably measures

reward threshold.

140
Cocaine and Intracranial Self-Stimulation

In general, drugs that increase the availability of catecholamines at

the synapse facilitate ICSS, and those agents that deplete the brain

of catecholamines or block cate!cholamine transmission inhibit ICSS.

For example, it is well documented that psychomotor stimulants such

as amphetamine will further decrease responding when responding has

been suppressed by a punisher (Geller and Seifter 1960). These

effects have been described as “a sensitization of the neural systems

that mediate the processing of rewards” (Stein 1964). This facilitation of reward by amphetamine is evident in studies using ICSS as

the reinforcer (Cassens and Mills 1973; Goodall and Carey 1975; Koob

et al. 1977; Phillips and Fibiger 1973; Stein 1964; Stein and Ray 1960;

Wauquier and Niemegeers 1974a). Although most studies have

employed a lever press response as the operant and have shown that

lever-pressing rates for ICSS increase with injection of amphetamine,

other measures of threshold have produced similar consistent changes

in ICSS threshold (Kornetsky and Esposito 1979). Cocaine also

facilitates responding for brain stimulation reward (Crow 1970;

Wauquier and Niemegeers 1974b).

While little work has explored site-specific effects for the facilitation

of brain stimulation by drugs, one study using a choice procedure and

continuous access to brain stimulation from three separate electrodes

in the same animals showed that rats preferred to self-stimulate in

the ventral tegmental area at moderate and high doses of d-amphetamine (Koob et al. 1977). In that study, rats given a choice of selfstimulating in the septal area, anterior lateral hypothalamus, or

posterior lateral hypothalamus chose the posterior lateral hypothalamus under the influence of d-amphetamine. When given a choice of

self-stimulating in the posterior lateral hypothalamus, the ventral

tegmental area, or the region of the locus coeruleus, the rats chose

the ventral tegmental area under the influence of d-amphetamine.

The authors suggested that this pattern of preference choices under

the influence of the drug parallelled the nondrug thresholds.

Anatomically, this preference pattern also parallels the course of the

mesolimbic dopamine system.

Amphetamine also decreases ICSS threshold in self-titration procedures where the current delivered decreases systematically with each

lever press, but the animal can reset the current back to its original

level by pressing another lever (Stein and Ray 1960; Zarevics and

Setler 1979). Functions relating rate of responding to current level,

141
i.e., rate/intensity (R/l) functions, shift to the left with administration

of psychostimulants including cocaine (Steiner and Stokeley 1973;

Phillips and LePiane 1986). as can be seen in figure 2. Other tests

have been developed to minimize the contribution of nonspecific

activating effects of psychomotor stimulants on the facilitation of

ICSS. Amphetamine and cocaine also enhance reward in these “ratefree” measures of ICSS threshold (Liebman and Butcher 1974; Esposito

et al. 1978; Kometsky and Esposito 1979). Using the threshold

procedure, Kometsky and Esposito (1981) demonstrated that cocaine

lowered the reward threshold at doses that actually increased the

detection threshold (figure 3). For detection threshold measures, the

initial noncontingent stimulus varied in intensity (at subreward

levels), while the second or response-contingent stimulus was held

constant at a rewarding intensity (above threshold) in order to

maintain responding. Thus, the initial stimulus acted as a discriminative stimulus indicating the availability of brain stimulation as a

reward (Kornetsky and Esposito 1981). This suggests that cocaine

was having an effect on reward threshold that was different from its

effect on the ability of the animals to make a psychophysical

discrimination.

The mechanism of action for this threshold-lowering action of cocaine

is not well understood, but it is sensitive to moderate to high doses

of opioid antagonists. Naloxone injected intraperitoneally at doses of

2 to 8 mg/kg attenuated the threshold-lowering actions of cocaine

(10 to 15 mg/kg, intraperitoneally), with the most effective dose

being 4 mg/kg of naloxone (Bain and Kometsky 1987). Similar

effects of naloxone have been observed with other psychomotor

stimulant drugs. Amphetamine lowers brain stimulation threshold, and

this effect is also reversed by naloxone (Esposito et al. 1980). These

results suggest some form of functional catecholamine/opioid interaction in cocaine reward. However, the specific nature of this

interaction needs to be elucidated, particularly given the lack of

effect of opioid antagonists on cocaine self-administration.

INTRAVENOUS COCAlNE SELF-ADMINISTRATlON: NEURAL

SUBSTRATES

Pharmacological Studies

Catecholamines have been strongly implicated in the reinforcing

properties of psychomotor stimulants (Pickens et al. 1978), and, more

specifically, the reinforcing properties of psychomotor stimulants have

been linked to the activation of central dopamine neurons and their

142
CURRENT LEVEL

FIGURE 2. Effects of intraperitoneally injected cocaine (10 mg/kg) in

rats lever-pressing for brain stimulation reward from

electrodes in the lateral hypothalamus

NOTE: Cocaine was injected intraperitoneally 15 minutes prior to the test session.

Rats were tested for 5 minutes at each of eight current levels in a descending

series. Data are expressed as mean ± SEM of the percent of maximal responding for each rat under the saline condition (n=8).

143
DOSE (mg/kg)

FIGURE 3. Mean effect of various doses of cocaine on the threshold

for brain stimulation reward and on the threshold for

brain stimulation detection

NOTE: Data are expressed as Z-scores based on the respective mean and standard

deviation of the effects of saline (n=4).

SOURCE: Kometsky and Bain 1982, Copyright 1982, the New York Academy of

Sciences.

postsynaptic receptors. When the synthesis of catecholamines is

blocked by administering alpha-methyl-para-tyrosine, an attenuation

of the subjective effects of euphoria associated with psychomotor

144
stimulants occurs in man (Jonsson et al. 1971), and a blockade of

the reinforcing effects of methamphetamine occurs in animals

(Pickens et al. 1968). Furthermore, low doses of dopamine antagonists will increase response rates for intravenous injections of damphetamine (Risner and Jones 1976; Yokel and Wise 1975; Yokel and

Wise 1976). Noradrenergic antagonists such as phenoxybenzamine,

phentolamine, and propranolol have no effect on stimulant (amphetamine) self-administration (Risner and Jones 1976; Yokel and Wise

1976; DeWit and Wise 1977).

The partial blockade of dopamine receptors produced by low doses of

dopamine antagonists has been hypothesized to reflect a partial

blockade of the reinforcing effects of d-amphetamine (Yokel and Wise

1976; DeWit and Wise 1977). Thus, animals are thought to compensate for decreases in the magnitude of the reinforcer by increasing

their self-administration behavior. Similar results have been observed

on cocaine self-administration with alpha-flupenthixol (Ettenberg et

al. 1982) (figure 4) and many other dopamine receptor antagonists

including haloperidol, chlorpromazine, metoclopramide, thioridazine,

and sulpiride (Roberts and Vickers 1984). Recently, the selective D-1

antagonist SCH 23390 has been shown to increase cocaine selfadministration in rats at doses that did not impair motor function,

whereas spiperone, a D-2-selective compound, produced only small

increases in responding at doses close to those that reduced motor

function (Koob et al. 1987a) (figure 5). These results suggest that

dopamine receptor blockade, and particularly D-1 receptor blockade,

may be involved in the reinforcing effects of psychomotor stimulants

in rats. lt should be noted, however, that the SCH 23390 compound

failed to consistently produce this action when administered intravenously to rhesus monkeys self-administering cocaine (Woolverton

1986).

Lession Studies

The role of dopamine in the reinforcing properties of psychomotor

stimulants was extended by the observations that 6-OHDA lesions of

the nucleus accumbens produced extinctionlike responding that

extended into a long-lasting reduction in self-administration of

cocaine and d-amphetamine over days (Roberts et al. 1977; Lyness

et al. 1979). This effect was thought to be largely due to the

depletion of dopamine, since rats pretreated with desmethylimipramine

before the nucleus accumbens lesion (to protect norepinephrine and

serotonin from destruction with the 6-OHDA) showed an identical

decrease in cocaine self-administration (Roberts et al. 1980). Similar

145
FIGURE 4. The effects of naltrexone and alpha-flupenthixol on the

loading dose (infusions in the first 20 minutes of the

3-hour test sessions) in both heroin- and cocaine-selfadministering animals

NOTE: Increases in responding were only observed in cocaine animals pretreated with

opiate receptor antagonist. An asterisk In the base of a histogram Indicates

that the treatment dose was reliably different from the no-drug saline

condition p<0.05 Newman-Keuls test).

SOURCE: Ettenberg et al. 1992, Copyright 1992. Springer-Verlag.

146
Hours Hours

FIGURE 5. Effects of the D-1 dopamine receptor antagonist

SCH 23390 (left side) and the D-2 dopamine receptor

antagonist spiperone (right side) on cocaine selfadministration

NOTE: Each point represents the average hourly intake of cocaine by injection (n=5).

Doses are in micrograms per kilogram. For SCH 23390, doses of 5,10, and

20 μg/kg significantly increased cocaine self-administration (p<0.05 NewmanKeuls test following ANOVA). For spiperone, the dose of 10 μg/kg significantly increased cocaine self administration (p<0.05 paired t-test overall

ANOVA p>0.05).

SOURCE: Koob et al. 1987a, Copyright 1987, Elsevier Scientific Publishers Ireland Ltd.

147
results were obtained following 6-OHDA lesions of the ventral

tegmental area (Roberts and Koob 1982). Subsequent studies have

shown that 6-OHDA lesions of the frontal cortex (Martin-lverson et

al. 1986) and corpus striatum (Koob et al. 1987b) do not produce this

decrease in responding for cocaine.

These results with intravenous psychomotor stimulant selfadministration are consistent with earlier studies showing that the

mesocorticolimbic dopamine system has an important role in mediating

the activation associated with psychomotor stimulants (Kelly et al.

1975; Kelly and lversen 1976; Roberts et al. 1975). For example,

6-OHDA lesions of the region of the nucleus accumbens block the

locomotor activation due to amphetamine (Kelly et al. 1975; Roberts

et al. 1975) and cocaine (Kelly and lversen 1976). These lesions,

however, do not block the locomotor activation associated with

scopolamine and caffeine (Joyce and Koob 1981), heroin (Vaccarino et

al. 1986), or corticotropin-releasing factor (Swerdlow and Koob 1985).

This supports the hypothesis that these lesion effects on activation

produced by psychomotor stimulants are not nonspecific changes in

motor capability.

Nevertheless, the simple interpretation of a reduction in responding

as a reward deficit is confounded in the self-administration studies by

the observation that dopamine receptor antagonists produce an

increase in responding, which has been interpreted as a decrease in

the reinforcing properties of the drug. In the 6-OHDA lesion studies

discussed above, partial lesions led instead to a partial reduction in

responding. Further confounding the reinforcement deficit interpretation is the series of studies reported by Le Moal and colleagues

showing that mesocorticolimbic dopamine lesions facilitate acquisition

of d-amphetamine self-administration (Le Moal et al. 1979; Deminiere

et al. 1984). Thus, other measures of reinforcing value are needed

that do not depend on a simple continuous reinforcement procedure

where each lever press produces a single intravenous injection.

In a recent study in our laboratory, rats that had been trained to

self-administer cocaine intravenously were subjected to a progressive

ratio procedure following 6-OHDA lesions to the nucleus accumbens

or corpus striatum. The rats with lesions of the nucleus accumbens

showed a significant decrease in the highest ratio for which they

would respond to obtain cocaine (Koob et al. 1987b) (figure 6).

Furthermore, rats with 6-OHDA nucleus accumbens lesions increased

significantly the highest ratios for which they would self-administer

148
FIGURE 6. Effects of 6-OHDA lesions to the nucleus accumbens and

corpus striatum on responding in rats self-administering

cocaine

*Bignificantfy different from sham group, p<0.05 Newman-Keuls test.

NOTE: Top panel shows continuous reinforcement data averaged for the first 3 days

postlesion (mean ± SEM). Sham: vehicle (0.1 mg/ml ascorbic acid In salineinjected controls. Caudate: rats receiving 8 ug in 2 ul of 6-OHDA injected

into the nucleus accumbens. Middle panel shows the dose-effect functions for

each group. H, two times the normal 0.75 mg/kg/injection dose; M refers to

middle dose range, 0.75 mg/kg/ injection; L refers to one-half of the 0.75

mg/kg/injection dose. Bottom panel shows the mean injections and mean

highest ratio obtained by each group on the progressive ratio schedule.

SOURCE: Koob et al. 1987b, Copyright 1987, Raven press.

149
apomorphine using a similar progressive ratio procedure (Roberts and

Vickers 1987), suggesting that the direct dopamine agonist had

become more reinforcing in supersensitive 6-OHDA-lesion rats. This

motivational probe thus avoids many of the problems associated with

measuring simple rates of responding. The results in the progressive

ratio test suggest that the decrease in rate of responding for cocaine

observed previously does in fact represent a motivational deficit.

A few studies have examined the role of serotoninergic systems in

mediating stimulant self-administration. In one study, animals were

injected intracerebroventricularly with the serotonin neurotoxin 5, 7-

dihydroxytryptamine (80 to 90 percent forebrain serotonin depletion)

prior to self-administration training. This lesion produced an

increase in the amount of d-amphetamine self-administered during

acquisition and after baseline stabilization. These rats had an

increased tendency to overdose. The authors speculated that a

generalized serotonin depletion produced “a decreased reinforcing

effect of d-amphetamine” (Lyness et al. 1980, p. 940). This effect

could not be replicated by 5,7-dihydroxytryptamine injections into the

nucleus accumbens (Lyness et al. 1980). It is clear that general

serotonin neurotoxin lesions change psychomotor stimulant selfadministration in a manner opposite to that of dopamine neurotoxin

lesions. However, without other behavioral tests, it is difficult to

assess whether this reflects increases or decreases in the reinforcing

effects of the psychomotor stimulant.

Recent work examining the functional output of the nucleus accumbens has established the region of the substantia innominata (ventral

pallidum) as a critical connection in the expression of nucleus

accumbens behavioral stimulation (Mogenson and Nielson 1988). For

example, electrolytic or ibotenic acid lesions of the ventral pallidum

block the supersensitive locomotor response to apomorphine produced

in rats with destruction of dopamine terminals in the nucleus

accumbens. In addition, this nucleus accumbens-ventral pallidum

connection is thought to be GABAergic, since GABA infused into the

ventral pallidum blocks the locomotor activation produced by direct

application of dopamine into the nucleus accumbens (Mogenson and

Nielson 1983), and low doses of the GABA agonist muscimol similarly

block the supersensitive apomorphine response in nucleus accumbens

6-OHDA-lesion rats (Swerdlow and Koob 1984).

Studies designed to explore the role of the nucleus accumbens-ventral

pallidum connection in the reinforcing effects of cocaine have recently been completed. Kainic acid lesions of the nucleus accumbens

150
produce decreases in rates of cocaine self-administration in rats

trained on 1.5 mg/kg/injection cocaine (Zito et al. 1985). There was

a high positive correlation between the degree to which responding

decreased and the extent of the lesion in the nucleus accumbens (Zito

et al. 1985). Similar effects have now been observed in rats with

ibotenic acid lesions of the ventral pallidum (Hubner and Koob 1987).

In this study, rats were trained to self-administer intravenous cocaine

(0.75 mg/kg/injection) in 3-hour daily sessions on a fixed-ratio 5

schedule. After reaching steady baseline responding (± 10 percent of

the average for 3 consecutive days), rats received either bilateral

injections of vehicle or ibotenic acid. Subjects receiving an ibotenic

acid lesion in the ventral pallidum showed a significant decrease in

cocaine self-administration when compared to vehicle-injected control

animals. A cocaine dose-effect determination revealed an overall

decrease in responding for cocaine for the lesion group, but the

integrity of the typical dose-effect relationship was preserved. The

effect of ibotenic acid lesions on cocaine self-administration was also

assessed using a progressive ratio procedure. In this test, the lesion

group displayed a significant decrease in the highest fixed-ratio value

obtained. These results suggest that the neurons in the nucleus

accumbens that mediate the reinforcing actions of psychomotor

stimulants may project to the ventral pallidum, as do the neurons

responsible for locomotor activation (Swerdlow and Koob 1985).

SUMMARY AND CONCLUSIONS

Neurochemical Substrates of Cocaine Reward

There is some significant overlap in the brain substrates that appear

to be involved in cocaine’s effect on brain stimulation reward and

those sites involved in cocaine self-administration. Brain sites shown

to be sensitive to the threshold-reducing effects of cocaine are in

the medial forebrain bundle. The medial forebrain bundle represents

one of the important substrates for brain stimulation reward, and

lesions of the dopamine system ascending in the medial forebrain

bundle disrupt intravenous cocaine self-administration. While work

with brain stimulation reward has generally focused on the midbrain

source of the medial forebrain bundle (the ventral tegmental area),

self-administration studies have focused on the forebrain connections

of the extrapyramidal motor system, the nucleus accumbens. Linking

the two is the mesocorticolimbic dopamine system. Indeed, it may be

this distinction that can explain the interesting effects of naloxone

on psychomotor stimulant facilitation of brain stimulation. lt will be

151
of some interest to determine if the action of cocaine on brain

stimulation reward and its interaction with opioids is site specific.

Implications for Cocaine Dependence

The brain dopamine system as a focus for cocaine reward leads to a

number of important implications for our understanding of cocaine

dependence and possible treatment. Hypothetically, this same system

may be involved in the withdrawal symptoms of psychomotor

retardation and depression following cocaine intake cessation. Also,

this same system may be pathologically altered in drug-sensitive

individuals (see studies of Le Moal et al. (1979) above). Drugs that

act on the dopamine system may be useful in reducing the craving

associated with the drug as well as withdrawal signs. Thus, the

identification of an important, perhaps even critical, substrate for the

rewarding effects of cocaine provides a focal point for more preclinical questions. This is not to argue that other neurotransmitters

are not involved, but that these transmitters may ultimately act

through this final common substrate.

Implications for Neural Substrates of Reward

What are the implications of this work with psychomotor stimulants

for brain substrates of reward in general? There is little doubt that

psychomotor stimulants are reinforcing in man, i.e., they are selfadministered and they produce affective reactions variously described

as euphoria or pleasure (Jonsson et al. 1971).

Since the neurochemical sites for psychomotor stimulant reward are

likely to be the presynaptic dopamine terminals located in the region

of the nucleus accumbens, frontal cortex, and other forebrain structures that originate from the ventral tegmental area, this system may

be part of the circuitry involved in the processing of natural reinforcers. Note, however, that intracranial self-administration of

cocaine is elicited from the frontal cortex, but not the nucleus accumbens (Goeders and Smith 1983). Thus, concomitant activation of

structures other than the nucleus accumbens may be an important

part of the circuitry involved in initiation of cocaine self-administration and reward.

In addition, these studies provide evidence to show that, in the rat,

the neural/neurochemical substrates for processing the reinforcing and

stimulant properties of psychomotor stimulants may be similar, if not

identical. Parallel manipulations using dopamine receptor antagonists

152
and 6-OHDA lesions produce parallel results. How far this parallelism

continues in further processing is under current investigation.

However, this overlap brings additional impetus to earlier hypotheses

relating reinforcement and motor function (Glickman and Schiff 1967).

lt may be that reinforcers or rewards that produce an activation of

an organism rely on the same system that is exaggerated by

psychomotor stimulants (Wise and Bozarth 1987). Whether all reward

is processed through this same circuitry in a serial or parallel

manner is an area of intense investigation and controversy (Wise

1982; Wise and Botarth 1984; Koob 1987; Koob and Goeders 1988).

REFERENCES

Angrist, B., and Sudilovsky, A. Central nervous system stimulants:

Historical aspects and clinical effects. In: lversen, L.L; Iversen,

S.D.; and Snyder, S.H., eds. Handbook of Psychopharmacology.

Vol. 11. New York: Plenum Press, 1976. pp. 99-163.

Bain, G.T., and Kornetsky, C. Naloxone attenuation of the effect of

cocaine on rewarding brain stimulation. Life Sci 40:1119-1125, 1987.

Cassens, G.P., and Mills, A.W. Lithium and amphetamine: Opposite

effects on threshold of intracranial reinforcement.

Psychopharmacologia 30:283-290, 1973.

Crow, T.J. Enhancement by cocaine of intracranial self-stimulation in

the rat. Life Sci 9:375-381, 1970.

Deminiere, J.M.; Simon, H.; Herman, J.P.; and Le Moal, M. 6-Hydroxydopamine lesion of the dopamine mesocorticolimbic cell bodies

increases (+)-amphetamine self-administration. Psychopharmacology

(Berlin) 83:281-284, 1984.

DeWit, H., and Wise, R.A. Blockade of cocaine reinforcement in rats

with the dopamine receptor blocker pimozide, but not with the

noradrenergic blockers phentolamine and phenoxybentamine. Can J

Psychol 31:195-203, 1977.

Esposito, R.U.; Motola, A.H.D.; and Kornetsky, C. Cocaine: Acute

effects on reinforcement thresholds for self-stimulation behavior to

the medial forebrain bundle. Pharmacol Biochem Behav 8:437-439,

1978.

Esposito, R.U.; Perry, W.; and Kornetsky, C. Effects of d-amphetamine and naloxone on brain stimulation reward.

Psychopharmacology (Berlin) 69:187-191, 1980.

Ettenberg, A.; Pettit, H.O.; Bloom, F.E.; and Koob, G.F. Heroin and

cocaine intravenous self-administration in rats: Mediation by

separate neural systems. Psychopharmacology (Berlin) 78:204-209,

1982.

153
Fischman, M.W.; Schuster, C.R; and Hatano, Y. A comparison of the

subjective and cardiovascular effects of cocaine and lidocaine in

humans. Pharmacol Biochem Behav 18:123-127, 1983.

Friedman, E.; Gershon, S.; and Rotrosen, J. Effects of active cocaine

treatment on the turnover of 5-hydroxytryptamine in the rat brain.

Br J Pharmacol 54:81-64, 1975.

Geller, I., and Seifter, J. The effects of meprobamate, barbiturates,

d-amphetamine and promazine on experimentally induced conflict in

the rat. Psychopharmacologia 1:482-492, 1960.

Glickman, S.E., and Schiff, B.B. A biological theory of reinforcement.

Psychol Rev 74:81-108, 1967.

Goeders, N.E., and Smith, J.E. Cortical dopaminergic involvement in

cocaine reinforcement. Science 221:773-775, 1983.

Goodall, E.B., and Carey, R.J. Effects of d- versus l-amphetamine,

food deprivation, and current intensity on self-stimulation of the

lateral hypothalamus, substantia nigra, and medial frontal cortex of

the rat. J Comp Physiol Psychol 89:1029-1045, 1975.

Groppetti, A.; Zambotti, F.; Biazzi, A.; and Mantegazza, P.

Amphetamine and cocaine on amine turnover. In: Usdin, E., and

Snyder, S.H., eds. Frontiers in Catecholamine Research. Oxford:

Pergamon Press, 1973. pp. 917-925.

Hernstein, R.J. Quantitative hedonism. J Psychiatr Res 8:399-412,

1971.

Hubner, C.B., and Koob, G.F. Ventral pallidal lesions produce

decreases in cocaine and heroin self-administration in the rat.

Neurosci Abstr 13: 1717, 1987.

Jonsson, LE.; Anggard, E.; and Gunne, L.M. Blockade of intravenous

amphetamine euphoria in man. Clin Pharmacol Ther 12:889-896,

1971.

Joyce, E.M., and Koob, G.F. Amphetamine-, scopolamine-, and

caffeine-induced locomotor activity following 6-hydroxydopamine

lesions of the mesolimbic dopamine system. Psychopharmacology

(Berlin) 73:311-313, 1981.

Kelly, P.H., and Iversen, S.D. Selective 6-OHDA induced destruction

of mesolimbic dopamine neurons: Abolition of psychostimulant

induced locomotor activity in rats. Eur J Pharmacol 40:45-56, 1976.

Kelly, P.H.; Seviour, P.W.; and Iversen, S.D. Amphetamine and

apomorphine responses in the rat following 6-OHDA of the nucleus

accumbens septi and corpus striatum. Brain Res 94:507-522, 1975.

Knapp, S., and Mandell, A.J. Narcotic drugs: Effects on the

serotonin biosynthetic systems of the brain. Science 177:1209-1211,

1972.

154
Koob, G.F. Separate neurochemical substrates for cocaine and heroin

reinforcement. In: Church, R.M.; Commons, M.L; Stellar, J.; and

Wagner, A.R., eds. Quantitative Analyses of Behavior: Biological

Determinants of Behavior. Vol. 7. Hillsdale, NJ: Lawrence

Erlbaum Associates, Inc., 1987. pp. 134156.

Koob, G.F., and Goeders, N.E. Neuroanatomical substrates of drug

self-administration. In: Liebman, J.M., and Cooper, S.J. The

Neuropharmacological Basis of Reward. New York: Oxford

University Press, 1988.

Koob, G.F.; Le, H.T.; and Creese, I. D-1 receptor antagonist

SCH 23390 increases cocaine self-administration in the rat.

Neurosci Lett 79:315-321, 1987a.

Koob, G.F.; Vaccarino, F.J.; Amalric, M.; and Bloom, F.E. Positive

reinforcement properties of drugs: Search for neural substrates.

In: Engel, J., and Oreland, L., eds. Brain Reward Systems and

Abuse. New York: Raven Press, 1987b. pp. 35-50.

Koob, G.F.; Winger, G.D.; Meyerhoff, J.L; and Annau, Z. Effects of

d-amphetamine on concurrent self-stimulation of forebrain and

brainstem loci. Brain Res 137: 109-126, 1977.

Kornetsky, C., and Bain, G. Biobehavioral bases of the reinforcing

properties of opiate drugs. Ann NY Acad Sci 398:240-259, 1982.

Kornetsky, C., and Esposito, R.U. Euphorigenic drugs: Effects on

reward pathways of the brain. Fed Proc 38:2473-2476, 1979.

Kornetsky, C., and Esposito, R.U. Reward and detection thresholds

for brain stimulation: Dissociative effects of cocaine. Brain Res

209:496-500, 1981.

Le Moal, M.; Stinus, L; and Simon, H. Increased sensitivity to (+)-

amphetamine self-administration by rats following mesocorticolimbic

dopamine neurone destruction. Nature 280:156-158, 1979.

Liebman, J.M., and Butcher, L.L Comparative involvement of

dopamine and noradrenaline in rate-free self-stimulation in

substantia nigra, lateral hypothalamus and mesencephalic central

gray. Naunyn Schmiedebergs Arch Pharmacol 284:167-194, 1974.

Lyness, W.H.; Friedle, N.M.; and Moore, K.E. Destruction of

dopaminergic nerve terminals in nucleus accumbens: Effect on damphetamine self-administration. Pharmacol Biochem Behav 11:553-

556, 1979.

Lyness, W.H.; Friedle, N.M.; and Moore, K.E. Increased selfadministration of d-amphetamine after destruction of 5-

hydroxytryptaminergic neurons. Pharmacol Biochem Behav 12:937-

941, 1980.

Marcus, R., and Kornetsky, C. Negative and positive intracranial

reinforcement thresholds: Effects of morphine.

Psychopharmacologia 38:1-14, 1974.

155
Martin-lverson, M.T.; Szostak, C.; and Fibiger, H.C. 6-Hydroxy

dopamine lesions of the medial prefrontal cortex fail to influence

intravenous self-administration of cocaine. Psychopharmacology

(Berlin) 88:310-314, 1986.

Matsuzaki, M. Alteration in pattern of EEG activities and

convulsant effect of cocaine following chronic administration in the

rhesus monkey. Electroencephalogr Clin Neurophysiol 45:1-15, 1978.

Mogenson, G.J., and Nielson, M.A. Evidence that an accumbens to

subpallidal GABAergic projection contributes to locomotor activity.

Brain Res Bull 11:309-314, 1983.

Olds, J., and Milner, P. Positive reinforcement produced by electrical

stimulation of septal area and other regions of rat brain. J Comp

Physiol Psychol 47:419-427, 1954.

Phillips, A.G., and Fibiger, H.C. Dopaminergic and noradrenergic

substrates of positive reinforcement: Differential effects of d- and

/-amphetamine. Science 179:575-577, 1973.

Phillips, A.G., and LePiane, F.G. Effects of pimozide on positive and

negative incentive contrast with rewarding brain stimulation.

Pharmacol Biochem Behav 24:1577-1582, 1986.

Pickens, R.; Meisch, R.A.; and Dougherty, J.A. Chemical interactions

in amphetamine reinforcement. Psychol Rep 23:1267-1270, 1968.

Pickens, R.; Meisch, R.A.; and Thompson, T. Drug self-administration:

An analysis of the reinforcing effects of drugs. In: Iversen, L.L.;

Iversen, S.D.; and Snyder, S.H., eds. Handbook of

Psychopharmacology. Vol. 12. New York: Plenum Press, 1978. pp.

1-37.

Pickens, R., and Thompson, T. Cocaine reinforced behavior in rats:

Effects of reinforcement magnitude and fixed ratio size. J

Pharmacol Exp Ther 161:122-129, 1968.

Post, R.M.; Kopanda, R.T.; and Black, K.E. Progressive effects of

cocaine on behavior and central amine metabolism in rhesus

monkeys: Relationship to kindling and psychoses. Biol Psychiatry

11:405-419, 1976.

Randrup, A., and Munkvad, I. Biochemical, anatomical and

psychological investigations of stereotyped behavior induced by

amphetamines. In: Costa, E., and Garahini, S., eds. Amphetamines

and Related Compounds. New York: Raven Press, 1970. pp. 695-

713.

Risner, M., and Jones, B.E. Role of noradrenergic and dopaminergic

processes in amphetamine self-administration. Pharmacol Biochem

Behav 5:477-482, 1976.

156
Roberts, D.C.S.; Corcoran, M.E.; and Fibiger, H.C. On the role of

ascending catecholaminergic systems in intravenous selfadministration of cocaine. Pharmacol Biochem Behav 6:615-620,

1977.

Roberts, D.C.S., and Koob, G.F. Disruption of cocaine selfadministration following 6-hydroxydopamine lesions of the ventral

tegmental area in rats. Pharmacol Biochem Behav 17:901-904, 1982.

Roberts, D.C.S.; Koob, G.F.; Fibiger, H.C.; Corcoran, M.E.; and Bloom,

F.E. The effect of 6-OHDA lesions of the nucleus accumbens on

cocaine facilitation of intracranial self-stimulation and self-administration. Neurosci Abstr 5:660, 1979.

Roberts, D.C.S.; Koob, G.F.; Klonoff, P.; and Fibiger, H.C. Extinction

and recovery of cocaine self-administration following 6-OHDA

lesions of the nucleus accumbens. Pharmacol Biochem Behav

12:781-787, 1980.

Roberts, D.C.S., and Vickers, G. Atypical neuroleptics increase selfadministration of cocaine: An evaluation of a behavioral screen for

antipsychotic activity. Psychopharmacology (Berlin) 82:135-139,

1984.

Roberts, D.C.S., and Vickers, G. Increased motivation to self-administer apomorphine following 6-hydroxydopamine lesion of the

nucleus accumbens. Ann NY Acad Sci, in press.

Roberts, D.C.S.; Zis, A.P.; and Fibiger, H.C. Ascending catecholamine

pathways and amphetamine induced locomotor activity: Importance

of Dopamine and apparent noninvolvement of norepinephrine. Brain

Res 93:441-454, 1975.

Simon, P. Psychopharmacological profile of cocaine. In: Usdin, E.,

and Snyder, S.H., eds. Frontiers of Catecholamine Research.

Oxford: Pergamon Press, 1973. pp. 1043-1044.

Spealman, R.D.; Goldberg, S.R.; Kelleher, R.T.; Goldberg, D.M.; and

Charlton, J.P. Some effects of cocaine and two cocaine analogs on

schedule controlled behavior of squirrel monkeys. J Pharmacol Exp

Ther 202:500-509, 1977.

Stein, L. Self-stimulation of the brain and the central stimulant

action of amphetamine. Fed Proc 23:836-850, 1964.

Stein, L., and Ray, O.S. Brain stimulation reward “thresholds” selfdetermined in rat. Psychopharmacologia 1:251-256, 1960.

Steiner, S.S., and Stokely, S.N. Methamphetamine lowers selfstimulation thresholds. Physiol Psychol 1:161-164, 1973.

Swerdlow, N.R., and Koob, G.F. The neural substrates of

apomorphine- stimulated locomotor activity following denervation of

the nucleus accumbens. Life Sci 35:2537-2544, 1984.

157
Swerdlow, N.R., and Koob, G.F. Separate neural substrates of the

locomotor-activity properties of amphetamine, heroin, caffeine, and

corticotropin releasing factor (CRF) in the rat. Pharmacol Biochem

Behav 23:303-307, 1985.

Vaccarino, F.J.; Amalric, M.; Swerdlow, N.R.; and Koob, G.F.

Blockade of amphetamine- but not opiate-induced locomotion

following antagonism of dopamine function in the rat. Pharmacol

Biochem Behav 24:61-65, 1986.

Valenstein, E.S. Problems of measurement and interpretation with

reinforcing brain stimulation. Psychol Rep 71:415-437, 1964.

Wauquier, A., and Niemegeers, C.J.E. Intracranial self-stimulation in

rats as a function of various stimulus parameters. IV. Influence

of amphetamine on medial forebrain bundle stimulation with

monopolar electrodes. Psychopharmacologia 34:265-274, 1974a.

Wauquier, A., and Niemegeers, C.J.E. Intracranial self-stimulation in

rats as a function of various stimulus parameters. V. Influence

of cocaine on medial forebrain bundle stimulation with monopolar

electrodes. Psychopharmacologia 38:201-210, 1974b.

Wise, R.A. Neuroleptics and operant behavior: The anhedonia

hypothesis. Behav Brain Sci 5:39-88, 1982.

Wise, R.A., and Bozarth, M.A. Brain reward circuitry: Four circuit

elements “wired” in apparent series. Brain Res Bull 12:203-208,

1984.

Wise, R.A., and Bozarth, M.A. A psychomotor stimulant theory of

addiction. Psychol Rev 94:469-492, 1987.

Woolverton, W.L Effects of a D1 and D2 dopamine antagonist on the

self-administration of cocaine and piribedil by rhesus monkeys.

Pharmacol Biochem Behav 24:531-535, 1986.

Yokel, R.A., and Wise, R.A. Increased lever pressing for amphetamine

after pimozide in rats: Implications for a dopamine theory of

reward. Science 187:547-549, 1975.

Yokel, R.A., and Wise, R.A. Attenuation of intravenous amphetamine

reinforcement by central dopamine blockade in rats.

Psychopharmacology (Berlin) 48:311-318, 1976.

Zarevics, P., and Setler, P.E. Simultaneous rate-independent and

rate-dependent assessment of intracranial self-stimulation: Evidence

for the direct involvement of dopamine in brain reinforcement

mechanisms. Brain Res 169:499-512, 1979.

Zito, K.A.; Vickers, G.; and Roberts, D.C.S. Disruption of cocaine and

heroin self-administration following kainic acid lesions of the

nucleus accumbens. Pharmacol Biochem Behav 23:1029-1036, 1985.

158
ACKNOWLEDGMENTS

This research was supported in part U.S. Public Health Service grants

DA04398, DA04043, and DA05292 from the National Institute on Drug

Abuse.

This is publication number 5076BCR from the Research Institute of

Scripps Clinic. Conan Kornetsky, Department of Psychiatry, Boston

University School of Medicine, is acknowledged for helpful discussions

and the use of his data.

AUTHORS

George F. Koob, Ph.D.

Carol B. Hubner, Ph.D.

Division of Preclinical Neuroscience

and Endocrinology

Department of Basic and Clinical Research

Research Institute of Scripps Clinic

10666 North Torrey Pines Road

La Jolla, CA 92037

159
Multiple Dopamine Receptors and

the Behavioral Effects of Cocaine

W.L. Woolverton and M.S. Kleven

INTRODUCTION

Cocaine is a psychomotor stimulant and has behavioral effects that

are typical of that class of drugs. In addition, it has been known

for some time that cocaine blocks the uptake of monoamines into

neurons. An assumption of behavioral pharmacology has been that

this biochemical action in the brain is involved in most if not all of

the behavioral effects of cocaine. In particular, a substantial amount

of research suggests that a number of those effects are mediated by

an increase in the concentration of the neurotransmitter dopamine

(DA) in synapses in the brain.

To exert its effects, DA acts on DA receptors. In recent years, it

has become clear that there is more than one type of receptor for

DA. After some discussion as to whether there are two, three, or

four DA receptors, the current consensus is that two are sufficient to

account for most of the available data. lf other receptor systems

can serve as examples, it is likely that the ranks of DA receptors

will swell again in the future. In the brain, these receptor subtypes

have been designated, straightforwardly if somewhat unimaginatively,

as D-1 and D-2. Originally they were characterized biochemically

according to their relationship to the production of cAMP in brain

homogenates (Kebabian and Calne 1979). D-1 receptors are positively

linked to adenylate cyclase. Agonists at the D-1 receptor stimulate

the production of CAMP. D-2 receptors were originally said not to

be linked to adenylate cyclase but have recently been found to be

negatively linked to adenylate cyclase, i.e., D-2 agonists can inhibit

the production of cAMP (Stoof and Kebabian 1984). Defining the

function of D-1 and D-2 receptors in the brain is currently a very

active area of research.

160
The purpose of this paper is to briefly review the evidence that

cocaine is an indirect DA agonist and to relate some of the behavioral effects of cocaine to its indirect action at D-1 or D-2 receptors. Our approach will be a pharmacological one: to compare the

behavioral effects of cocaine to those of DA agonists with selective

D-1 or D-2 or mixed actions and to examine the interaction of

cocaine with antagonists with selective D-1 and D-2 or mixed actions.

The basic premise is simple: if a D-1 agonist mimics a behavioral

effect of cocaine and a D-1 antagonist blocks the effect, it suggests

that D-1 receptors are involved in that effect.

lt must be noted at the outset that there is precious little information available that bears directly on the role of DA receptor subtypes

in the behavioral effects of cocaine. A major reason for this deficiency, surely, is that it has been less than a decade since we have

been aware that there is more than one DA receptor. Although a

selective D-1 agonist (SW 38393) was discovered in 1976 (Setler et

al. 1978), the fact that it was a selective D-1 agonist was not appreciated until later. Even now it is one of the very few D-1 agonists

that cross the blood-brain barrier, a fact that makes problematic the

determination of an in vivo potency series that is essential to the

demonstration of a receptor-mediated effect. Moreover, an antagonist

selective for the D-1 receptor (SCH 23390) has been available for

only 5 years (lorio et al. 1983). (Sadly, the days of drug names like

arecoline, rauwolfia, and bufotenin seem to be gone.) It should also

be said that the recent intense interest in cocaine has made the

investigation of receptor mechanisms involved in its effects a viable

research issue.

NEUROCHEMICAL EFFECTS OF COCAINE

Cocaine has been known for many years to block the uptake of

monoamines (MacMillan 1959). Although initial research to demonstrate this effect was done in the periphery, it was not long before

researchers postulated that this was the mechanism of action in the

central nervous system (CNS). Indeed, cocaine has been shown to

block the uptake of norepinephrine (NE), DA, and 5-hydroxytryptamine (5-HT) into presynaptic terminals in vitro (Koe 1976; Taylor and

Ho 1978; Randrup and Braestrup 1977). Although it is often difficult

to distinguish between release and blockade of uptake, it is generally

believed that cocaine has little effect on monoamine release except at

very high doses (Heikkila et al. 1975; Scheel-Kruger et al. 1977).

161
of these neurochemical effects, attention has centered upon the DA

reuptake-blocking properties of cocaine. DA levels in the caudate

nucleus have been reported to increase by about 50 percent within

20 minutes after injection of 20 mg/kg cocaine (Pradhan et al. 1978;

Bhattacharyya et al. 1979). Within 60 minutes after injection, these

effects had returned to control values. Scheel-Kruger (1972) could

not find an effect of moderate doses of cocaine (e.g., 25 mg/kg) on

DA levels in rat brain 2 hours after injection. The reason for this

relatively minor change in brain DA levels may be explained by a

recent study. Church et al. (1987) used microdialysis to measure

extracellular concentrations of DA in rat striatum. Cocaine produced

as much as a ninefold increase in striatal DA, with a timecourse of

approximately 1 hour. Other DA uptake inhibitors (nomifensine and

benztropine) also increased extracellular DA with slight differences in

maximal effect and duration of effect. Apparently, a rather large

increase in extracellular DA was not detected in earlier studies using

ex vivo analysis. This effect could be due to a decrease in DA

metabolism subsequent to a blockade of DA reuptake. Cocaine has

also been reported to increase DA turnover (Patrick and Barchas

1977), perhaps by stimulating tyrosine hydroxylase activity. More

recently, cocaine has been found to increase the concentration of the

DA metabolite HVA in striatum, an effect that also implies that

turnover of DA was increased and would not be based solely upon

uptake inhibition (Hanson et al. 1987).

Other neurochemical actions of cocaine may be involved in some of

its behavioral effects. Although cocaine has been found to block the

uptake of NE and 5-HT, concomitant with the increase in DA described above, there was a decrease in brain NE and 5-HT levels

(Pradhan et al. 1978; Bhattacharyya et al. 1979). Within 20 minutes

after injection, there was a 40 percent depletion of 5-HT in the diencephalon-midbrain and pons-medulla and a 20 percent decrease of

NE levels in both regions. Within 60 minutes after injection, these

effects had returned to control values. The decrease in NE and

5-HT, an effect opposite that seen with DA, may reflect differences

in neuronal compensation and/or effects of cocaine beyond simple

blockade of monoamine reuptake. Recent studies of cocaine binding

by Reith and colleagues (Reith et al. 1983; Reith et al. 1986) suggest

the existence of several separate cocaine binding sites, each

associated with uptake of either DA or 5-HT. The existence of

5-HT-related binding sites may explain the effects of cocaine on 5-HT

uptake in vitro (Koe 1976; Taylor and Ho 1978; Randrup and Braestrup

1977). lt should be noted, however, that antidepressant drugs that

block neuronal uptake of NE and/or 5-HT, but do not affect DA

162
uptake, are not psychomotor stimulants. Moreover, cocaine has a

unique pharmacology, since it is a local anesthetic and decreases

5-HT synthesis (Knapp and Mandell 1972). lt should be emphasized

that cocaine’s behavioral effects may involve any or all of these

neuronal actions.

BEHAVlORAL EFFECTS OF COCAINE

lt has long been known that cocaine is a psychomotor stimulant.

Research with humans in the late 19th century demonstrated this fact

unequivocally (Freud 1884). Some 30 years later, when animal

researchers began working with cocaine, it was noted that cocaine

stimulated locomotor activity and produced stereotyped behavior in

rats and monkeys (Downs and Eddy 1932; Tatum and Seevers 1929).

More recently, it has been demonstrated that cocaine is selfadministered by a wide variety of species, and methods have been

developed for studying its prominent subjective effects, including

euphoria. These effects are certainly critical determinants of the

apparently exceptional abuse potential of cocaine.

Many of the behavioral effects of cocaine are believed to be the

result of enhanced DA neurotransmission in the CNS (Wise 1984). A

variety of types of experimental evidence converge upon this point.

Two of the major factors are that DA agonists have many of the

same behavioral effects as cocaine and that DA antagonists can block

these effects of DA agonists and of cocaine. What follows is a

review of the data showing that DA receptors are involved in several

of the more prominent behavioral effects of cocaine. To the extent

possible, we will relate these to the DA receptor subtypes that may

be involved. In several instances in which no data are available for

cocaine we will consider what, if anything, is known about

d-amphetamine, a compound that is often behaviorally interchangeable

with cocaine. In addition, we will consider the data examining other

neurochemical mechanisms, particularly noradrenergic, where they

have been implicated.

DA RECEPTORS AND THE REINFORCING EFFECTS OF COCAINE

lf a response that resulted in drug delivery increases in frequency,

the drug is defined as a positive reinforcer. Reinforcing effects are

a primary determinant of dependence potential. It has become clear

that cocaine is a particularly strong positive reinforcer. As with the

other behavioral effects of cocaine, it is suspected that this effect is

due to its effects on DA neurotransmission in the brain.

163
lf DA receptors mediate the reinforcing effects of cocaine, then

agonists that directly stimulate these receptors would be expected to

function as positive reinforcers. Baxter et al. (1974) reported that

the DA agonist apomorphine functioned as a positive reinforcer in

rats. Rats prepared with intravenous catheters self-administered

apomorphine at rates substantially above those maintained by the drug

vehicle. Apomorphine injections within a session were regularly

spaced, a pattern that was reminiscent of the pattern of cocaine selfadministration. Moreover, DA antagonists increased the rate of

apomorphine self-administration just as has been found with cocaine

self-administration (see below), further supporting the notion that DA

receptors are involved in this reinforcing effect (Baxter et al. 1974;

Gill et al. 1978; Yokel and Wise 1978). This was the first evidence

that a drug that is a direct agonist at DA receptors could function as

a positive reinforcer. Since that time, the finding has been confirmed several times in rats (Wise et al. 1976; Yokel and Wise 1978).

The discovery of a selective D-1 agonist (SKF 38393) and antagonist

(SCH 23390) allowed us to begin to examine the role of DA receptor

subtypes in the reinforcing effects of cocaine. In one study

(Woolverton et al. 1984), rhesus monkeys were prepared with intravenous catheters and allowed to self-administer cocaine or

d-amphetamine in daily experimental sessions that were 2 or 3 hours

long. When self-administration was stable under these baseline

conditions, drug vehicle or a dose of a DA agonist was made available

for several consecutive sessions. When drug vehicle was available,

responding declined to low levels over a period of 4 to 10 sessions,

as expected. When a DA agonist was available, responding was maintained at levels above those maintained by the vehicle in most of the

monkeys. The single, clear exception was the D-1 agonist SW 38393.

In all cases, responding for this agonist occurred at or below vehicle

levels over a wide range of doses. The other drugs had in common

agonist actions at the D-2 receptor. In fact, the compound that was

most reliably self-administered, piribedil, is the most selective in

terms of D-2 actions. This study clearly implicated D-2 receptors in

the reinforcing effects of cocaine.

Experiments with antagonists also suggest that DA receptors are

involved in the reinforcing effects of cocaine. The first experiments

to test this hypothesis involved administering catecholamine receptor

blockers before a session of cocaine self-administration. Wilson and

Schuster (1972) showed that when chlorpromazine (CPZ) was given

before a session of cocaine self-administration, the rate of cocaine

self-administration increased as though the unit dose of cocaine had

164
been reduced. That is, it appeared that the effects of cocaine had

been at least partially antagonized by CPZ. Of course, since CPZ

blocks a multitude of receptors, it was difficult to relate this result

to its effect at any single type of receptor. Nevertheless, the results

were consistent with the hypothesis that catecholamine receptors are

involved in the reinforcing effects of cocaine.

Research since this initial experiment has tended to implicate DA

receptors in this effect. Haloperidol, perphenazine, pimozide,

flupenthixol, (+)-butaclamol, sulpiride, thioridazine, and metoclopramide, all primarily DA receptor blockers, have been found to increase

the rate of cocaine self-administration in several different species (de

la Garza and Johanson 1982; Gill et al. 1978; Johanson et al. 1976; de

Wit and Wise 1977; Risner and Jones 1976; Roberts and Vickers 1984;

Woods et al. 1978). An interesting effect that has not been systematically pursued was the discovery by Wilson and Schuster (1973) that

atropine produces a similar increase in cocaine self-administration.

On the other hand, DA agonists have been shown to have the

opposite effect. The direct agonist bromocriptine (Hubner and Koob

1987; Woolverton and Kleven 1987), as well as the indirect DA

agonists amphetamine and phenmetrazine reduced the rate of cocaine

self-administration when they were administered in the same manner

as the DA antagonists described previously (Wilson and Schuster 1973;

Herling et al. 1979). In addition, a variety of other compounds

including clozapine (Roberts and Vickers 1982), pargyline (Wilson and

Schuster 1974), physostigmine (de la Garza and Johanson 1982; Wilson

and Schuster 1973), morphine, pentobarbital, and imipramine (Wilson

and Schuster 1973; Herling et al. 1979) have been found to have no

effect on or to reduce cocaine self-administration.

Virtually all of the compounds that have been shown to increase

cocaine self-administration are primarily D-2 antagonists. The

availability of a selective D-1 antagonist (lorio et al. 1983) made it

possible to compare the effects of this compound on cocaine selfadministration with those of D-2 antagonists. In another study

(Woolverton 1986), rhesus monkeys were allowed to self-administer

cocaine during daily 2-hour experimental sessions. When behavior

was stable, they were injected with the D-1 antagonist SCH 23390 or

the D-2 antagonist pimozide at a timepoint before the session that

assured maximal effect during the session. Intermediate doses of

pimozide generally increased the rate of responding for cocaine in the

same manner as has been found with other DA antagonists. On the

other hand, SCH 23390 usually failed to increase the rate of responding for cocaine. A high dose of pimozide resulted in a pattern of

165
responding for cocaine that was similar to what was seen when saline

was available for self-administration (figure 1). In contrast, a dose

of SCH 23390 that had comparable rate-reducing effects completely

suppressed responding early in the session and did not affect cocaine

self-administration later in the session. There was no suggestion of

pattern of responding similar to that seen when saline was available.

FIGURE 1. Effects of a high dose of pimozide or SCH 23390 on

responding maintained by cocaine or piribedil in rhesus

monkeys

NOTE: Diagonal marks represent drug injections. Responding on the inactive lever

was marked by the lower pen. DA antagonists were given IV either 2 hours

(pimozide) or 30 minutes (SCH 23390) before the 2-hour session.

166
The antagonists had effects on responding for the D-2 agonist

piribedil that were similar to their effects on cocaine-maintained

responding. These results are consistent with our findings with

agonist self-administration in suggesting a D-2 receptor role in the

reinforcing effects of cocaine. Goeders et al. (1986) reached similar

conclusions using rats as experimental subjects. The D-2 antagonist

sulpiride eliminated cocaine-maintained responding whereas

SCH 23390 did not.

Although these experiments with antagonists are consistent with the

hypothesis that D-2 receptors are involved in the reinforcing effects

of cocaine, they must be considered equivocal evidence of antagonism of the reinforcing effects of cocaine. Rate of drug selfadministration under the conditions utilized in these experiments is

determined by a number of drug effects in addition to the reinforcing

effects (Johanson 1976). For instance, if the effects of cocaine that

tend to reduce rate of responding regardless of the reinforcing

stimulus were antagonized, rate of responding for cocaine would

increase as well. In fact, Herling and Woods (1980) concluded that

the evidence from behavioral studies was most consistent with a

mutual antagonism of rate-reducing effects of cocaine and neuroleptics. In addition, we have recently examined the effects of SCH

23390 or pimozide on behavior maintained under a multiple schedule

of food or cocaine delivery (Woolverton and Virus, submitted).

Although it was possible to reduce cocaine self-administration with

either of these compounds as though the reinforcing effects of

cocaine were blocked, reductions occurred only at doses that comparably reduced responding maintained by food. Thus, the effects of DA

antagonist were not specific for cocaine but were generalized ratereducing effects.

As to NE mechanisms in the reinforcing effects of cocaine, both

direct and indirect NE agonists have been found not to be selfadministered by animals (Risner and Jones 1976; Woolverton 1987). In

addition, antagonists at NE receptors, including phenoxybenzamine,

phentolamine, propranolol, and prazosin, have been shown to have no

effect on or to reduce cocaine self-administration (de Wit and Wise

1977; Goldberg and Gonzalez 1976; Wilson and Schuster 1974;

Woolverton 1987). These findings suggest that the NE actions of

cocaine do not play a prominent role in its reinforcing effects.

In summary, much of the available evidence suggests that a D-2

receptor action is involved in the reinforcing effects of cocaine. It

should be pointed out that Koob et al. (1987) recently reported that

167
SCH 23390 consistently increased the rate of cocaine selfadministration in rats, while the D-2 antagonist spiperone only

occasionally had this effect. These results are in direct contrast to

our findings with rhesus monkeys. Although the reasons for this

discrepancy are unclear, it is possible that species differences played

a role. It is possible as well that the short duration of action of

SCH 23390 played a role in the negative effects we observed in

rhesus monkeys. In view of these conflicting results and the data

concerning the effects of SCH 23390 on the discriminative stimulus

properties of cocaine to be described in the next section, the effects

of D-1 antagonists on the reinforcing effects of cocaine bear further

investigation. We are currently investigating the effects of continuous infusion of SCH 23390 on cocaine self-administration.

DA RECEPTORS AND THE DISCRlMlNATlVE STIMULUS EFFECTS

OF COCAINE

Discriminative stimuli are stimuli that set the occasion for a particular behavior to be reinforced. For instance, if a behavior is reinforced in the presence of a red light, the red light is called the

discriminative stimulus for that response. lt has been shown

repeatedly in a number of species that a variety of psychoactive

drugs, including cocaine, can function as discriminative stimuli. lf

one behavior is reinforced following an injection of cocaine and a

second behavior reinforced following an injection of saline, the drug

comes to function as the discriminative stimulus that controls

differential responding. This paradigm has proven quite useful for

classifying drugs according to similarities in their discriminative

stimulus properties. Drugs that substitute for or block the effects of

the training drug may be considered to provide information relevant

to the mechanism of action of the training drug. In addition, since

the classification of drugs by animals trained in this paradigm is

similar to the classification of drugs according to their subjective

effects in humans, the paradigm is considered by many to be useful

as an animal model of subjective effects (Schuster et al. 1981). When

considered together, these two types of information may provide an

indication of the central actions involved in the subjective effects of

a drug. To the extent that subjective effects play a role in dependence potential, results of these experiments may help us understand

the central mechanisms involved in cocaine dependence.

Initial investigations of the central mechanisms of the discriminative

stimulus properties of cocaine examined the role of DA and DA

receptors by testing apomorphine in cocaine-trained animals.

168
Colpaert et al. (1976) and McKenna and Ho (1980) have reported that

the D-2 agonist apomotphine substituted for 10 mg/kg cocaine in

rats, findings that were consistent with the hypothesis that DA and

DA receptors play a primary role in the discriminative stimulus

properties of cocaine. However, further research has not supported

these findings. In a later study, Colpaert et al. (1979) tested the D-2

agonists apomorphine, piribedil, and bromocriptine in rats trained to

discriminate 10 mg/kg cocaine from saline. All three drugs only partially substituted for cocaiane. The most complete substitution for

cocaine was seen with apomorphine, which resulted in a maximum of

75 percent cocaine-appropriate responding. Piribedil engendered a

maximum of 57 percent, and only 29 percent drug-lever responding

was seen with bromocriptine. Similarly, Jarbe (1981) reported a maximum of 21 percent cocaine-appropriate responding with apomorphine

in pigeons trained to discriminate 5.6 mg/kg cocaine from saline. In

a later paper, Jarbe (1984) reported that apomorphine induced 56 percent drug-key responses in pigeons trained to discriminate 3.0 mg/kg

cocaine from saline. Similar results in pigeons trained to discriminate

2.0 mg/kg cocaine from saline were found by de la Garza and

Johanson (1985). Thus, virtually all of the available data demonstrate, at best, a partial substitution of D-2 agonists for cocaine.

Although there is substantially less data available with D-1 agonists,

the story to this point is similar to that with D-2 agonists. Wood

and Emmett-Oglesby (1987) have found that the D-1 agonist SKF

38393 partially substitutes for cocaine in rats trained to discriminate

10 mg/kg cocaine from saline. The results are consistent with what

has been found with this agonist in animals trained to discriminate

d-amphetamine from saline. We have found that SKF 38393 partially

substitutes for 0.5 mg/kg d-amphetamine in rats (figure 2) and

monkeys (Woolverton 1984).

Unfortunately, findings of partial substitution make it necessary to

equivocate on conclusions. The data would support the conclusion

that a D-1 or a D-2 agonist partially reproduces the effects of

cocaine. It should be noted that this partial substitution was often

the result of averaging different levels of responding in different

animals rather than the result of a consistent intermediate level of

responding in ail animals. It is possible, therefore, that different

animals learned to discriminate different components of what is a

complex stimulus, which includes both D1 and D-2 actions. Both

possibilities are consistent with the conclusion that D-1 and D-2

receptors are involved in the discriminative stimulus properties of

169
DRUG DOSE (MG/KG, IP)

FIGURE 2. Effects of SKF 38393 and piribedil alone or in combination

in rats trained to discriminate 0.5 mg/kg d-amphetamine

from saline

NOTE: Drugs were injected IP 10 minutes before the 15-minute session. When

combinations were given they were administered in separate syringes, one

immediately after the other.

cocaine and raise the possibility that a combination of D-1 and D-2

agonists might substitute for cocaine. Although this has not been

170
tested with cocaine, we have done this experiment with d-amphetamine, a drug that consistently substitutes for cocaine as a discriminative stimulus. When the D-2 agonist piribedil was combined

with the D-1 agonist SKF 38393 in rats trained to discriminate

d-amphetamine from saline, partial substitution was again the result

even up to doses that reduced rate of lever pressing (figure 2).

Thus, a combination of D-1 and D-2 actions was not similar to

d-amphetamine. Alternatively, it must be considered that partial

substitution may often represent a nonspecific effect of a drug that

is not like the training drug and at the same time is not like saline.

If this interpretation is accurate, it suggests that DA receptors are

not involved in this effect of cocaine. lt should also be mentioned

at this point that the neuropharmacology of a drug surely changes

with dose. A most graphic behavioral demonstration of this effect

was provided by Stolerman and D’Mello (1981), who demonstrated that

apomorphine substituted for a high dose of d-amphetamine but not

for a low dose and only partially for an intermediate dose. Such an

effect is possible with cocaine. Thus, although DA receptor mechanisms are not clear-cut when 10 mg/kg cocaine is used as the

training dose, they may be more apparent at higher doses. Clearly,

further research is needed to thoroughly address these issues.

Experiments with antagonists have also been used to examine receptor

mechanisms in the discriminative stimulus properties of cocaine. As

with other pharmacological effects, a competitive antagonist of cocaine would be expected to cause a parallel shift of the cocaine

dose-response function to the right. Unfortunately, in most of these

experiments, only the effects of combining an antagonist with the

training dose of cocaine were considered. Colpaert et al. (1976)

reported that haloperidol failed to block the effects of 5.0 mg/kg

cocaine. In contrast, Jarbe (1978) reported that pimozide, haloperidol, and CPZ all reduced the percentage of drug responses in rats

trained to discriminate 4.0 mg/kg cocaine from saline in a T-maze.

Colpaert and colleagues showed that haloperidol, pimozide, and

spiperone could reduce the percentage of responses that occurred on

the drug lever following the training dose of cocaine (10 mg/kg) in

rats (Colpaert et al. 1978a; Colpaert et al. 1978b). The most commonly reported effect has been a reduction in drug-appropriate

responding with the training dose of cocaine from 100 percent to

approximately 50 percent; that reduction occurred only at doses that

substantially reduced ongoing behavior. Complete saline-appropriate

responding has not been reported with combinations of a D-2 antagonist and cocaine. In the single case where this was examined, the

dose-response function for cocaine was shifted to the right, and the

171
maximum effect was reduced from 100 percent to approximately 50

percent when cocaine was combined with pimozide or haloperidol

(McKenna and Ho 1980). These results are suggestive of a noncompetitive antagonism of the discriminative stimulus properties of cocaine

by D-2 antagonists. Unfortunately, it is necessary to equivocate over

these findings of partial antagonism in much the same way as over

partial substitution with DA agonists.

The effects of the D-1 antagonist SCH 23390 on the discriminative

stimulus properties of cocaine are being examined in experiments

currently in progress in our laboratory (Kleven et al., in press).

Rhesus monkeys were trained to discriminate cocaine from saline and

were injected with combinations of SCH 23390 and cocaine. The

dose-response function for cocaine was shifted fourfold to eightfold

to the right when combined with SCH 23390. Two important factors

should be noted with the D-1 antagonist that have not been apparent

with D-2 antagonists. The first is that drug-lever responding was

consistently reduced from 100 percent to 0, evidence for complete

blockade of the effects of cocaine. Further, the D-1 antagonist

appeared to function as a competitive antagonist. That is, the doseresponse function for cocaine was shifted parallel to the right.

Similar results have been found with d-amphetamine as a discriminative stimulus in rhesus monkeys (Woolverton et al. 1987) and rats

(Nielsen and Jepsen 1985). These data strongly suggest that D-1

receptors are somehow involved in the discriminative stimulus

properties of cocaine and, perhaps, of psychomotor stimulants in

general. Further, the data suggest that this novel group of compounds (i.e., D-1 antagonists) may function as antagonists of the

subjective effects of cocaine. We feel that these are very exciting

findings.

In summary, research with DA agonists and antagonists has demonstrated partial substitution for cocaine by D-2 agonists and, at best,

partial blockade of the effect with D-2 antagonists. Although these

data are consistent with a role for D-2 receptors in the discriminative stimulus effects of cocaine, the data could not be described as

conclusive. Substitution data with a D-1 agonist are similar to those

with D-2 agonists, and the limited amount of data with D-1 antagonists suggests a role for D-1 receptors as well.

The possibility that other mechanisms are involved in the discriminative stimulus properties of cocaine should also be considered. Early

research with cocaine suggested that NE was probably not important.

Beta adrenergic agonists either failed to substitute or only partially

172
substituted for cocaine in rats (Colpaert et al. 1979) and the NE

antagonists phenoxybenzamine (alpha), propranolol (beta), and sotalol

(beta), have been reported not to block the discriminative stimulus

properties of cocaine (McKenna and Ho 1980; Jarbe 1978). However,

more recent research with d-amphetamine has made it clear that it is

important to consider the role of NE in the discriminative stimulus

properties of cocaine. The indirect NE agonist nisoxetine has been

shown to substitute for d-amphetamine as a discriminative stimulus in

several species (Evans and Johanson 1987; Snoddy and Tessel 1983;

Woolverton 1984) and the alpha NE antagonist prazosin has been

reported to antagonize the discriminative stimulus properties of

d-amphetamine in mice (Snoddy and Tessel 1985). Moreover, Colpaert

et al. (1980) reported that MAO-B inhibitors substitute for cocaine

and postulated that endogenous beta-phenylethylamine mediates the

effects of cocaine, a finding that should be pursued. Finally, it

should be mentioned that the antagonists cinanserin (5-HT) and

atropine (ACH) did not block the discriminative stimulus properties of

cocaine (McKenna and Ho 1980). Research with these neurotransmitter systems is extremely limited and should be expanded

(Lakoski and Cunningham, this volume).

DA RECEPTORS AND OTHER BEHAVIORAL EFFECTS OF COCAINE

Among the other behavioral effects of cocaine are stimulation of

locomotor activity, generation of stereotyped behavior, and reduction

in food intake (Scheel-Kruger 1972; van Rossum and Simons 1969;

Woolverton et al. 1978). In addition, cocaine disrupts schedulecontrolled behavior (Herling et al. 1979). There is considerable

evidence that CNS DA is involved in these effects (Creese and

lverson 1975; Kelly and lverson 1976; Menon et al. 1967; Ziegler et al.

1972). Like cocaine, the DA agonists apomorphine, piribedil, and

LY141865 have been shown to stimulate locomotor activity (Dourish

1983; Fuller et al. 1983; Offermeier and van Rooyen 1986). All are

primarily D-2 agonists. On the other hand, the D-1 DA agonist

SKF 38393 has little or no effect on locomotor activity in normal

rats (Breese et al. 1985). A number of different investigators have

reported that DA antagonists can block cocaine-induced locomotor

activity. The D-2 antagonists pimozide and haloperidol completely

antagonized the hyperactivity induced by cocaine and its analogs in

rats and mice (Bhattacharyya et al. 1979; Heikkila et al. 1979; ScheelKruger et al. 1977). Although these results suggest that D-2

receptors are involved in the locomotor effects of cocaine, interpretation of the interactions between cocaine and D-2 antagonists is

somewhat problematic. D-2 antagonists themselves reduce locomotor

173
activity, an effect that may give the appearance that locomotor

stimulation by cocaine is blocked.

At higher doses, psychomotor stimulants, including cocaine, induce a

syndrome of repetitive movements termed stereotyped behavior.

Although in some sense this could be considered locomotor activity, it

is generally treated as a different category of behavioral effect.

Available data suggest different areas of the brain are involved in

locomotor effects and stereotyped behavior (Kelly and lversen 1976).

High doses of D-2 agonists induce stereotyped behavior, and D-2

antagonists block the stereotyped behavior produced by cocaine and

by D-2 agonists (Porsolt et al. 1982; Scheel-Kruger et al. 1977).

Although D-1 agonists have been reported to induce a syndrome of

repetitive grooming and mouth movements, these effects are apparently not stereotypic in nature (Molloy and Waddington 1987;

Rosengarten et al. 1983; Starr and Starr 1986).

These studies are consistent in suggesting that a D-2 receptor action

is involved in the locomotor stimulant effects of cocaine. However,

it would be premature to suggest that the D-1 receptors do not play

a significant role. Although the effects of the D-1 antagonist

SCH 23390 on cocaine-induced locomotor stimulation or stereotypy

have not been determined, Mailman et al. (1984) reported that SCH

23390 blocked d-amphetamine-induced locomotor effects. In addition,

the behavioral effects of D-2 agonists can be blocked by the D-1

antagonist SCH 23390 (Mailman et al. 1984; Pugh et al. 1985; Walters

et al. 1987). For instance, it has been reported that D-1 and D-2

agonists must be combined to produce the full spectrum of

DA-mediated behaviors, including stereotyped behaviors (Arnt et al.

1987; Braun and Chase 1986). It is also of interest to note that the

D-1 agonist stimulates locomotor activity in rats with lesions of the

nigrostriatal pathway (Breese et al. 1985; Setler et al. 1978) suggesting that D-1 receptors do have some role in the regulation of

DA-mediated effects. A number of investigators have interpreted

these data to suggest that there are important interactions between

D-1 and D-2 receptors in the expression of DA-mediated behavioral

effects and that D-1 receptors are essential to the expression of

those effects. Thus, a growing body of evidence suggests that D-1

receptors may be involved in the expression of dopaminergic drug

effects. The role of D-1 receptors in the locomotor effects of

cocaine is an area where research is needed.

Regarding other behavioral effects of cocaine, both D-1 and D-2

agonists can reduce food intake in the same manner as cocaine

174
(Gilbert and Cooper 1985; Barzaghi et al. 1973). Since this is often a

nonspecific effect, it probably reveals little about receptor mechanisms. The interaction of cocaine with DA antagonists has not been

examined in a food intake paradigm. lt has been shown, however,

that a D-1 antagonist can block the effects of amphetamines on food

intake (Gilbert and Cooper 1985). The data with D-2 antagonists is

more controversial, perhaps because there is more of it (Foltin et al.

1983; Gilbert and Cooper 1985). Similarly, apomorphine has some

effects of schedule-controlled behavior that are similar to those of

cocaine (Abelson and Woods 1980), although the specificity of these

effects must be questioned as well. The effects of D-1 agonists on

schedule-controlled responding have not, to our knowledge, been

reported. The effects of cocaine on schedule-controlled behavior that

can be partially blocked by D-2 antagonists and a mutual antagonism

of rate-reducing effects of cocaine and D-2 antagonists has been

postulated to account for these effects (Herling and Woods 1980).

Similarly, in the Kleven et al. (in press) drug discrimination experiment, cocaine clearly reversed the effects of SCH 23390 on response

rate and vice versa. In addition, NE may be important in the effects

of cocaine on schedule-controlled behavior. In a recent paper, Tessel

and Barrett (1986) showed that prazosin could antagonize the effects

of cocaine on operant behavior. Further research is needed to follow

up these provocative findings.

SUMMARY AND CONCLUSIONS

Cocaine has a number of behavioral effects that are typical of

psychomotor stimulants and likely involve DA mechanisms in the

brain. These effects include, but are not limited to, reinforcing

effects, discriminative stimulus effects, locomotor effects, effects on

food intake, and effects on schedule-controlled behavior. The available data suggest that D-2 receptors are involved in the reinforcing

effects of cocaine, although the jury is still out concerning the role

of D-1 receptors. NE receptors apparently do not play a major role

in this effect. The role of D-2 receptors in the discriminative stimulus effects is unclear because of partial substitution and partial

blockade effects with D-2 agonists and antagonists, respectively.

Data with D-1 agonists and antagonists suggest that D-1 receptors

may play a necessary-but-not-sufficient role in the discriminative

stimulus effects of cocaine. Moreover, results in several species

suggest that NE may be important in this effect of cocaine. Very

little data currently exist concerning the role of DA receptor

subtypes in the other behavioral effects of cocaine. However, data

with d-amphetamine suggest that D-1 as well as D-2 receptors should

175
be investigated. In addition, there is evidence to suggest that NE is

involved in some of cocaine’s effects as well, an action that should

be considered further.

So, what mediates the behavioral effects of cocaine? Clearly, the

answer is not a simple one, basically because the CNS pharmacology

of cocaine is complex. In addition to the effects we have concentrated on, cocaine has effects on other neurotransmitters and local

anesthetic effects that must be considered. It is impossible to say

that any one pharmacological effect of cocaine mediates its behavioral

effects, indeed even that it mediates any one behavioral effect. In

fact, the word “mediates” is surely virtually meaningless in the

context of the CNS. Perhaps the best that can be said about the

role of DA receptors in the reinforcing effects of cocaine is they are

likely a link in a chain of events that ultimately results in the sensations produced by cocaine. Surely numerous other, perhaps more critical, links remain to be discovered. It is an exciting time to be

doing research with cocaine.

REFERENCES

Abelson, J.S., and Woods, J.H. Effects of apomorphine on elicited and

operant pecking in pigeons. Psychopharmacology (Berlin) 71:237-

241, 1980.

Arnt, J.; Hyttel, J.; and Perregaard, J. Dopamine D1 receptor

agonists combined with the selective D2 agonist quinpirole facilitate

the expression of oral stereotyped behavior in rats. Eur J

Pharmacol 133:137-145, 1987.

Barzaghi, F.; Gropetti, A.; Mantegazza, P.; and Muller, E.E.

Reduction of food intake by apomorphine: A pimozide-sensative

effect. J Pharm Pharmacol 25:909-911, 1973 (Letter).

Baxter, B.L.; Gluckman, M.I.; Stein, L.; and Scerni, R.A. Selfinjection of apomorphine in the rat: Positive reinforcement by a

dopamine receptor stimulant. Pharmacol Biochem Behav 2:387-391,

1974.

Bhattacharyya, A.K; Aulakh, C.S.; Pradhan, S.; Ghosh, P.; and

Pradhan, S.N. Modification of behavioral and neurochemical effects

of cocaine by haloperidol. Arch Int Pharmacodyn Ther 238:71-80,

1979.

Braun, A.R., and Chase, T.N. Obligatory D-1/D-2 receptor interaction

in the generation of dopamine agonist related behaviors. Eur J

Pharmacol 131:301-306, 1986.

176
Breese, G.R.; Baumeister, A.; Napier, T.C.; Frye, G.D.; and Mueller,

R.A. Evidence that D-1 dopamine receptors contribute to the

supersensitive behavioral responses induced by

I-dihydroxyphenylalanine in rats treated neonatally with

6-hydroxydopamine. J Pharmacol Exp Ther 235:287-295, 1985.

Church, W.H.; Justice, J.B., Jr.: and Byrd, L.D. Extracellular dopamine

in rat striatum following uptake inhibition by cocaine, nomifensine

and benztropine. Eur J Pharmacol 139:345-348, 1987.

Colpaert, F.C.; Niemegeers, C.J.E.; and Janssen, P.A.J. Cocaine cue in

rats as it relates to subjective effects: A preliminary report. Eur

J Pharmacol 40:195-199, 1976.

Colpaert, F.C.; Niemegeers, C.J.E.; and Janssen, P.A.J. Neuroleptic

interference with the cocaine cue: Internal stimulus control of

behavior and psychosis. Psychopharmacology (Berlin) 58:247-255,

1978a.

Colpaert, F.C.; Niemegeers, C.J.E.; and Janssen, P.A.J. Discriminative

stimulus properties of cocaine and amphetamine, and antagonism by

haloperidol: A comparative study. Neuropharmacology 17:937-942,

1978b.

Colpaert, F.C.; Niemegeers, C.J.E.; and Janssen, P.A.J. Discriminative

stimulus properties of cocaine: Neuropharmacological characteristics

as derived from stimulus generalization experiments. Pharmacol

Biochem Behav 10:535-546, 1979.

Colpaert, F.C.; Niemegeers, C.J.E.; and Janssen, P.A.J. Evidence that

a preferred substrate for type B monoamine oxidase mediates

stimulus properties of MAO inhibitors: A possible role for

-phenylethylamine in the cocaine cue. Pharmacol Biochem Behav

13:513-517, 1980.

Creese, I., and lversen, S.D. The pharmacological and anatomical

substrates of the amphetamine response in the rat. Brain Res

83:419-436, 1975.

de la Garza, R., and Johanson, C.E. Effects of haloperidol and

physostigmine on self-administration of local anesthetics.

Pharmacol Biochem Behav 17:1295-1299, 1982.

de la Garza, R., and Johanson, C.E. Discriminative stimulus

properties of cocaine in pigeons. Psychopharmacology (Berlin)

85:23-30, 1985.

de Wit, H., and Wise, R.A. Blockade of cocaine reinforcement in rats

with the dopamine receptor blocker pimozide, but not with the

noradrenergic blockers phentolamine and phenoxybenzamine.

Can J Psychol 31:195-203, 1977.

Dourish, C.T. Piribedil: Behavioral, neurochemical and clinical

profile of a dopamine agonist. Prog Neuropsychopharmacol Biol

Psychiatry 7:3-27, 1977.

177
Downs, A.W., and Eddy, N.B. The effect of repeated doses of cocaine

in the rat. J Pharmacol Exp Ther 48:199-200, 1932.

Evans, S.M., and Johanson, C.E. Amphetamine-like effects of

anorectics and related compounds in pigeons. J Pharmacol Exp

Ther 241:817-825, 1987.

Foltin, R.W.; Woolverton, W.L.; and Schuster, C.R. The effect of

d-amphetamine and haloperidol alone and in combination on milk

drinking in rats. Psychopharmacology (Berlin) 80:342-344, 1983.

Freud, S. Uber Coca., 1884. In: Byck, R., ed. Cocaine Papers:

Sigmund Freud. New York: Stonehill Publishing Co., 1974. pp. 47-

73.

Fuller, R.W.; Hemrick-Luecke, SK; Wong, D.T.; Pearson, D.;

Threlkeld, P.G.; and Hynes, M.D. Altered behavioral response to a

D2 agonist, LY141865, in spontaneously hypertensive rats exhibiting

biochemical and endocrine responses similar to those in normotensive rats. J Pharmacol Exp Ther 227:354-359, 1983.

Gilbert, D.B., and Cooper, S.J. Analysis of dopamine D1 and D2

receptor involvement in d- and l-amphetamine-induced anorexia in

rats. Brain Res Bull 15:385-389, 1985.

Gill, C.A.; Holz, W.C.; Zirkle, C.L.; and Hill, H. Pharmacological

modification of cocaine and apomorphine self-administration in the

squirrel monkey. In: Deniker, P.; Radouco-Thomas, C.; and

Villeneuve, A., eds. Proceedings of the Tenth Congress of the

Collegium international Neuropsychopharmacologicum. New York:

Pergamon Press, 1978. pp. 1477-1484.

Goeders, N.E.; Dworkin, S.I.; and Smith, J.E. Neuropharmacological

assessment of cocaine self-administration into the medial prefrontal

cortex. Pharmacol Biochem Behav 24:1429-1440, 1986.

Goldberg, S.R., and Gonzalez, F.A. Effects of propranolol on

behavior maintained under fixed-ratio schedules of cocaine injection

or food presentation in squirrel monkeys. J Pharmacol Exp Ther

198:626-634, 1976.

Hanson, G.R.; Matsuda, L.A.; and Gibb, J.W. Effects of cocaine on

methamphetamine-induced neurochemical changes: Characterization

of cocaine as a monoamine uptake blocker. J Pharmacol Exp Ther

242:507-513, 1987.

Heikkila, R.E.; Orlansky, H.; and Cohen, G. Studies on the distinction

between uptake inhibition and release of dopamine in rat brain

tissue slices. Biochem Pharmacol 24:847-852, 1975.

Heikkila, R.E.; Cabbat, F.S.; Manzino, L.; and Duvoisin, R.C.

Rotational behavior induced by cocaine analogs in rats with

unilateral 6-hydroxydopamine lesions of the substantia nigra:

Dependence upon dopamine uptake inhibition. J Pharmacol Exp

Ther 211:189-194, 1979.

178
Herling, S.; Downs, D.A.; and Woods, J.H. Cocaine, d-amphetamine

and pentobarbital effects on responding maintained by food or

cocaine in rhesus monkeys. Psychopharmacology (Berlin) 64:261-269,

1979.

Herling, S., and Woods, J.H. Chlorpromazine effects on cocainereinforced responding in rhesus monkeys: Reciprocal modification

of rate-altering effects of the drugs. J Pharmacol Exp Ther

214:354-361, 1980.

Hubner, C.B., and Koob, G.F. Effect of bromocriptine pretreatment

on intravenous cocaine self-administration in the rat.

Pharmacologist 29:158, 1987.

lorio, L.C.; Barnett, A.; Leitz, F.H.; Houser, V.P.; and Korduba, C.A.

SCH 23390, a potential benzazepine antipsychotic with unique

interactions on dopaminergic systems. J Pharmacol Exp Ther

226:462-468, 1983.

Jarbe, T.U.C. Cocaine as a discriminative cue in rats: Interactions

with neuroleptics and other drugs. Psychopharmacology (Berlin)

59:183-187, 1978.

Jarbe, T.U.C. Cocaine cue in pigeons: Time course studies and

generalization to structurally related compounds (norcocaine,

WIN 35,428 and 35,065-2) and (+)-amphetamine. Br J Pharmacol

73:843-852, 1981.

Jarbe, T.U.C. Discriminative stimulus properties of cocaine. Effects

of apomorphine, haloperidol, procaine and other drugs.

Neuropharmacology 23:899-907, 1984.

Johanson, C.E. Drugs as reinforcers. In: Blackman, D.E., and

Sanger, D.J., eds. Contemporary Research in Behavioral

Pharmacology. New York: Plenum Press, 1978. pp. 325-390.

Johanson, C.E.; Kandel, D.A.: and Bonese, K. The effects of perphenazine on self-administration behavior. Pharmacol Biochem

Behav 4:427-433, 1976.

Kebabian, J.W., and Calne, D.B. Multiple receptors for dopamine.

Nature 277:93-96, 1979.

Kelly, P.H., and Iversen, S.D. Selective 6-OHDA-induced destruction

of mesolimbic dopamine neurons: Abolition of psychostimulantinduced locomotor activity in rats. Eur J Pharmacol 40:45-56, 1976.

Kleven, M.S.; Anthony, E.W.; Goldberg, L.I.; and Woolverton, W.L

Blockade of the discriminative stimulus effects of cocaine in rhesus

monkeys with the D1 dopamine antagonist SCH 23390.

Psychopharmacology (Berlin), in press.

Knapp, S., and Mandell, A.J. Narcotic drugs: Effects on the

serotonin biosynthetic systems of the brain. Science 177: 1209-1211,

1972.

179
Koe, B.K. Molecular geometry of inhibitors of the uptake of catecholamines and serotonin in synaptosomal preparations of rat brain.

J Pharmacol Exp Ther 199:649-661 ,1976.

Koob, G.F.; Le, H.T.; and Creese, I. The D-1 dopamine receptor

antagonists SCH 23390. Neurosci Lett 79:315-320, 1987.

MacMillan, W.H. A hypothesis concerning the effect of cocaine on

the action of sympathomimetic amines. Br J Pharmacol 14:385,

1959.

Mailman, R.B.; Schulz, D.W.; Lewis, M.H.; Staples, L; Rollema, H.; and

Dehaven, D.L. SCH 23390: A selective D1 dopamine antagonist

with potent D2 behavioral actions. Eur J Pharmacol 101:159-160,

1984.

McKenna, M.L., and Ho, B.T. The role of dopamine in the discriminative stimulus properties of cocaine. Neuropharmacology 19:297-

303, 1980.

Menon, M.K.; Dandiya, P.C.; and Bapna, J.S. Modification of the

effect of some central stimulants in mice pretreated with alphamethyl-p-tyrosine. Psychopharmacologia 10:437-444, 1967.

Molloy, A.G., and Waddington, J.L. Assessment of grooming and other

behavioral responses to the D-1 dopamine receptor agonist SK&F

38393 and its R- and S-enantiomers in the intact adult rat.

Psychopharmacology (Berlin) 92:164-168, 1987.

Nielsen, E.B., and Jepsen, S.A. Antagonism of the amphetamine cue

by both classical and atypical, antipsychotic drugs. Eur J

Pharmacol 111:167-176, 1985.

Offermeier, J., and van Rooyen, J.M. A comparative study of the

locomotor effects of apomorphine and the “atypical dopamine

agonists” (piribedil and S3608). Life Sci 38:895-903, 1986.

Patrick, R.L., and Barchas, J.D. Potentiation by cocaine of the

stimulus-induced increase in dopamine synthesis in rat brain striatal

synaptosomes. Neuropharmacology 16:327-332, 1977.

Porsolt, R.D.; Roux, S.; and Jalfre, M. Antiapomorphine and locomotor effects of neuroleptics in rhesus monkeys. Pharmacol

Biochem Behav 17:1309-1312, 1982.

Pradhan, S.; Roy, S.N.; and Pradhan, S.N. Correlation of behavioral

and neurochemical effects of acute administration of cocaine in

rats. Life Sci 22: 1737-1744, 1978.

Pradhan, S.N.; Battacharyya, A.K: and Pradhan, S. Serotonergic

manipulation of the behavioral effects of cocaine in rats. Commun

Psychopharmacol 2:481-486, 1978.

Pugh, M.T.; O’Boyle, K.M.; Molloy, A.G.; and Waddington, J.L. Effects

of the putative D-1 antagonist SCH 23390 of stereotyped behavior

induced by the D-2 agonist RU24213. Psychopharmacology (Berlin)

87:308-312, 1985.

180
Randrup, A., and Braestrup, C. Uptake inhibition of biogenic amines

by newer antidepressant drugs: Relevance to the dopamine

hypothesis of depression. Psychopharmacology (Berlin) 53:309-314,

1977.

Reith, R.E.A.; Meisler, B.E.; Sershen, H.; and Lajtha, A. Structural

requirements for cocaine congeners to interact with dopamine and

serotonin uptake sites in mouse brain and to induce stereotyped

behavior. Biochem Pharmacol 35:1123-1129, 1986.

Reith, R.E.A.; Sershen, H.; Allen, D.L.; and Lajtha, A. A portion of

[

3H]cocaine binding in brain is associated with serotonergic

neurons. Mol Pharmacol 23:600-606, 1983.

Risner, M.E., and Jones, B.E. Role of noradrenergic and dopaminergic

processes in amphetamine self-administration. Pharmacol Biochem

Behav 5:477-482, 1976.

Roberts, D.C.S., and Vickers, G. Atypical neuroleptics increase selfadministration of cocaine: An evaluation of a behavioral screen for

antipsychotic activity. Psychopharmacology (Berlin) 82:135-139,

1984.

Rosengarten, H.; Schweitzer, J.W.; and Friedhoff, A.J. Induction of

oral dyskinesias in naive rats by D1 stimulation. Life Sci 33:2479-

2482, 1983.

Scheel-Kruger, J. Behavioral and biochemical comparison of amphetamine derivatives, cocaine, benztropine and tricyclic antidepressant

drugs. Eur J Pharmacol 18:63-73, 1972.

Scheel-Kruger, J.; Braestrup, C.; Nielson, M.; Golembiowska, K.; and

Mogilnicka, E. Cocaine: Discussion on the role of dopamine in the

biochemical mechanism of action. In: Ellinwood, E.H., Jr., and

Kilbey, M.M., eds. Cocaine and Other Stimulants. New York:

Plenum Press, 1977. pp. 373407.

Schuster, C.R.; Fischman, M.W.; and Johanson, C.E. Internal stimulus

control and subjective effects of drugs. In: Thompson, T., and

Johanson, C.E., eds. Behavioral Pharmacology of Human Drug

Dependence. National Institute on Drug Abuse Research Monograph

37. DHHS Pub. No. (ADM) 81-1137. Rockville, MD: the Institute,

1981. pp. 116-129.

Setler, P.E.; Sarau, H.M.; Zirkle, C.L.; and Saunders, H.L. The central

effects of a novel dopamine agonist. Eur J Pharmacol 50:419-430,

1978.

Snoddy, A.M., and Tessel, R.E. Nisoxetine and amphetamine share

discriminative stimulus properties in mice. Pharmacol Biochem

Behav 19:205-210, 1983.

Snoddy, A.M., and Tessel, R.E. Effect on psychomotor stimulant cues

and locomotor activity in mice. Eur J Pharmacol 116:221-228, 1985.

181
Starr, B.S., and Starr, M.S. Grooming in the mouse is stimulated by

the dopamine D1 agonist SKF 38393 and by low doses of the D1

antagonist SCH 23390, but is inhibited by dopamine D2 agonists, D2

antagonists and high doses of SCH 23390. Pharmacol Biochem

Behav 24:637-639, 1986.

Stolerman, I.P., and D’Mello, G.D. Role of training conditions in

discrimination of central nervous system stimulants by rats.

Psychopharmacology (Berlin) 73:295-303, 1981.

Stoof, J.C., and Kebabian, J.W. Two dopamine receptors:

Biochemistry, physiology and pharmacology. Life Sci 35:2281-2296,

1984.

Tatum, A.L., and Seevers, M.H. Experimental cocaine addiction. J

Pharmacol Exp Ther 36:401-410, 1929.

Taylor, D., and Ho, B.T. Comparison of inhibition of monoamine

uptake by cocaine, methylphenidate and amphetamine. Res Commun

Chem Pathol Pharmacol 21:67-75, 1978.

Tessel, R.E., and Barrett, J.E. Antagonism of the behavioral effects

of cocaine and d-amphetamine. Psychopharmacology (Berlin) 90:436-

440, 1986.

van Rossum, J.M., and Simons, F. Locomotor activity and anorexigenie action. Psychopharmacologia 14:248-254, 1969.

Walters, J.R.; Bergstrom, D.A.; Carlson, J.H.; Chase, T.N.; and Braun,

A.R. D1 dopamine receptor activation required for ostsynaptic

expression of D2 agonist effects. Science 236:719-722, 1987.

Wilson, M.C., and Schuster, C.R. The effects of chlorpromazine on

psychomotor stimulant self-administration in the rhesus monkey.

Psychopharmacologia 26:115-126, 1972.

Wilson, M.C., and Schuster, C.R. The effects of stimulants and

depressants on cocaine self-administration behavior in the rhesus

monkey. Psychopharmacologia 31:291-304, 1973.

Wilson, M.C., and Schuster, C.R. Aminergic influences on intravenous

cocaine self-administration by rhesus monkeys. Pharmacol Biochem

Behav 2:563-571, 1974.

Wise, R.A. Neural mechanisms of the reinforcing actions of cocaine.

In: Grabowski, J., ed. Cocaine: Pharmacology, Effects and

Treatment of Abuse. National Institute on Drug Abuse Research

Monograph 50. DHHS Pub. No. (ADM) 84-1326. Washington, DC:

Supt. of Docs., U.S. Govt. Print. Off., 1984. pp. 15-33.

Wise, R.A.; Yokel, R.A.; and de Wit, H. Both positive reinforcement

and conditioned aversion from amphetamine and apomotphine in

rats. Science 191:1273-1275, 1976.

Woods, D.M., and Emmett-Oglesby, M.W. Evidence for D2, receptor

involvement in tolerance to the discriminative stimulus properties of

cocaine. Abstr Soc Neurosci 13:1717, 1987.

182
Woods, J.H.; Herling, S.; and Winger, G. Chlorpromazine and

haloperidol-induced changes in some behavioral effects of cocaine

and amphetamine. In: Deniker, P.; Radouco-Thomas, C.; and

Villeneuve, A., eds. Proceedings of the Tenth Congress of the

Collegium international Neuropsychopharmacologicum. New York:

Pergamon Press, 1978. pp. 1485-1502.

Woolverton, W.L. Pharmacological analysis of the discriminative

stimulus properties of d-amphetamine in rhesus monkeys.

Pharmacologist 26:161, 1984.

Woolverton, W.L. Effects of a D1 and a D2 dopamine antagonist on

the self-administration of cocaine and piribedil by rhesus monkeys.

Pharmacol Biochem Behav 24:531-535, 1986.

Woolverton, W.L.; Goldberg, L.I.; and Ginos, J.Z. Intravenous selfadministration of dopamine receptor agonists by rhesus monkeys.

J Pharmacol Exp Ther 230:678-683, 1984.

Woolverton, W.L.; Kamien, J.B.; and Kleven, M.S. Blockade of the

discriminative stimulus (DS) effects of cocaine and d-amphetamine

in rhesus monkeys with the D1 dopamine antagonist SCH 23390.

Pharmacologist 29:158, 1987.

Woolverton, W.L.; Kandel, D.A.; and Schuster, C.R. Tolerance and

cross-tolerance to cocaine and d-amphetamine. J Pharmacol Exp

Ther 205:525-536, 1978.

Woolverton, W.L., and Kleven, M.S. Effects of bromocriptine and

desmethylimipramine on cocaine self-administration in rhesus

monkeys. Presented at the Annual Meeting of the International

Study Group Investigating Drugs as Reinforcers, Philadelphia, PA,

1987.

Woolverton, W.L. Evaluation of the role of norepinephrine in the

reinforcing effects of psychomotor stimulants in rhesus monkeys.

Pharmacol Biochem Behav 26:835-839, 1987.

Yokel, R.A., and Wise, R.A. Amphetamine-type reinforcement by

dopaminergic agonists in the rat. Psychopharmacology (Berlin)

58:289-296, 1978.

Ziegler, H.; Del Basso, P.; and Longo, V.G. Influence of

6-hydroxydopamine and of a-methyl-p-tyrosine on the effects of

some centrally acting agents. Physiol Behav 8:391-396, 1972.

ACKNOWLEDGMENTS

Research was supported by U.S. Public Health Service grants

DA 00250 and DA 00085 from the National Institute on Drug Abuse.

183
AUTHORS

William L. Woolverton, Ph.D.

Mark S. Kleven, Ph.D.

Department of Pharmacological and Physiological Sciences

Drug Abuse Research Center

The University of Chicago

Chicago, IL 60637

184
Neurobehavioral Pharmacology of

Cocaine

Steven I. Dworkin and James E. Smith

INTRODUCTION

Behavioral pharmacologists have developed five behavioral screens

that are widely used to investigate the abuse liability of pharmacological agents. These procedures include drug self-administration,

conditioned place preference, drug discrimination, electrical brain

stimulation, and the use of schedule-controlled behavior. Each of the

five behavioral procedures has its own strengths and weaknesses, a

fact that necessitates the use of several procedures to thoroughly

investigate the behavioral effects of any drug. All of these procedures have been, or are currently, used to investigate the abuse

liability of cocaine and the neurobiological mechanisms involved in

drug abuse. This chapter will review procedures used to investigate

the reinforcing, rate modulating, and stimulus effects of cocaine.

Furthermore, this chapter provides an example of an integrated

neurobiological approach to determining the neurobiology of cocaine

abuse.

DRUG SELF-ADMINISTRATlON

The drug self-administration procedure that was developed more than

30 years ago has been used extensively to determine the reinforcing

effects of drugs. Several different routes of drug administration,

including oral, intramuscular, intranasal, intravenous, inhalation, and

direct delivery into various brain sites have been studied using

several species of subjects including humans (Brady and Lukas 1984).

A typical experimental protocol involves surgically preparing subjects

with chronic indwelling catheters that allow for the intravenous

presentation of a drug. The subject is then placed in an operant

conditioning chamber containing a response lever. Responses on this

185
lever result in the activation of a syringe pump that injects a precise

amount of the drug into the subject.

Studies investigating drugs that are self-administered by nonhuman

subjects have provided a list of pharmacological agents that are

abused by humans. This procedure thus provides a general screen for

substances that have a potential for being abused. Cocaine in particular has been shown to quickly engender robust self-administration

patterns and has become the standard to compare other drugs

against (Woods et al. 1987). However, one of the limitations of the

self-administration procedure is that it provides a confounded measure

of reinforcing efficacy. For example, when a relatively large range

of cocaine doses is studied in the self-administration paradigm, an

inverted U-shaped dose-effect curve is obtained. That is, low and

high doses of the drug maintain lower rates of responding than

moderate doses. Since it has been determined that high doses of the

drug are preferred (Johanson and Schuster 1975; lglauer and Woods

1974), the descending limb of a self-administration dose-effect curve

is not indicative of a decrease in reinforcing efficacy but is more

likely the result of unconditioned (rate decreasing) drug effects

(Woods et al. 1987). The ascending limb of the curve may also be

partially influenced by unconditioned drug effects that result in

nonspecific increases in motor activity. Thus, the dose-effect curve

obtained from self-administration procedures is a function of both the

reinforcing and response-rate-modulating effects of the drug.

Another factor present in the self-administration procedure is the

discriminative stimulus functions of the drug. Drugs that are selfadministered also result in interoceptive stimulus changes that allow

an organism to accurately detect the delivery of a drug. However,

the role of stimulus effects in drug self-administration has not been

thoroughly investigated. Some questions that remain unanswered

include the precise involvement of stimulus functions in the regulation of drug intake and the influence of stimulus effects in modulating the reinforcing efficacy of a drug. The self-administration

procedure clearly involves several factors including reinforcing, ratemodulating, and stimulus effects of a drug, all of which may require

independent investigation.

DRUG DlSCRlMlNATlON

The drug discrimination paradigm has been used to evaluate the

stimulus functions of cocaine. A typical drug discrimination

procedure involves the use of an operant chamber containing two

186
response manipulanda. Prior to the observation period, subjects are

trained to make one response if a drug is administered or the other

if saline is delivered. Subjects quickly learn to detect and report the

presence or absence of interoceptive drug-related stimuli. Once the

animals are trained, the effects of other doses and other drugs, both

alone and in combination with the training drug, can be evaluated.

The degree to which they produce responding on the drug-associated

option is used as an index for assessing stimulus properties.

Cocaine has robust stimulus properties, and animals are easily trained

to discriminate cocaine from vehicle infusions (Woods et al. 1987).

Studies investigating drugs that substitute for cocaine (i.e., result in

responding on the cocaine-associated lever), or drugs that do not

substitute for or block the discriminative stimulus effects of cocaine,

provide data that implicate the activation of dopaminergic systems as

one of the major factors responsible for the stimulus functions of the

drug. However, the precise behavioral and neurobiological mechanisms involved in the discriminative stimulus functions of cocaine are

still being intensely investigated (Woolverton and Kleven, this

volume). Furthermore, an index of stimulus properties provides only

an indirect assessment of abuse liability. The relationship between

the stimulus effects and reinforcing effects of a substance is not

clear. There are several classes of drugs that have stimulus properties and are not self-administered. Furthermore, drugs that substitute

for cocaine in the drug discrimination task typically result in lower

rates of self-administration or are sometimes not self-administered.

Moreover, drugs that can attenuate the stimulus properties of cocaine

may increase, have no effect on, or decrease self-administration.

One final point about drug discrimination is that the procedure can

result in a rate-independent assessment of stimulus properties.

Doses of the drug that do not alter response rates result in drugappropriate responding.

RATE DEPENDENCY

The observation that the ongoing rate of responding is an important

determinant of the behavioral effect of a drug (Thompson et al. 1981)

is one of the cornerstones of behavioral pharmacology. Studies on

the effects of different classes of pharmacological agents on schedule-controlled behavior have continued to increase our understanding

of drug action as well as behavioral mechanisms. The effects of cocaine on schedule-controlled behavior can be summarized as follows:

(1) low doses capable of increasing motor activity have little or no

effect on operant behavior; (2) intermediate doses increase low rates

187
of responding but produce less of an increase or have no effect on

high rates; (3) larger doses increase low rates and decrease high

rates; and (4) very large doses decrease responding regardless of rate.

These effects have been observed using both simple and complex

schedule contingencies and a variety of experimental subjects

(Gonzalez and Goldberg 1977). A general conclusion is that, with few

exceptions (Johanson 1978), regardless of how responding is engendered or maintained, cocaine results in comparable rate-dependent

effects (Barrett 1976). These effects, however, are not specific to

cocaine and are observed after the administration of other central

nervous system stimulants.

A limited number of studies have investigated the effects of

combinations of cocaine with other drugs on schedule-controlled

behavior. Both the alpha1-adrenergic antagonist prazosin (Tessel and

Barrett 1986) and the neuroleptic chlorpromazine (Wilson and Schuster

1975) antagonize the rate-decreasing effects of cocaine, while atropine (Wilson and Schuster 1975) and ethanol (Aston-Jones et al. 1984)

enhance the rate-decreasing effects of cocaine. The combination of

diazepam and cocaine on schedule-controlled behavior can either

attenuate or enhance the rate-increasing effects of diazepam alone,

depending on the dose of cocaine administered (Ford et al. 1980).

Investigations of the effects of chlorpromazine in modulating the

behavioral effects of cocaine epitomize the strength of a multifaceted

behavioral approach. Chlorpromazine generally decreases responding

regardless of the rate of responding or the event that is used to

maintain responding. Furthermore, chlorpromazine can antagonize the

rate-decreasing effects of cocaine on food-maintained responding

(Wilson and Schuster 1975). Therefore, it is not surprising that the

drug can alter cocaine self-administration (Gill et al. 1977; Herling

and Woods 1980; Roberts and Vickers 1984). However, chlorpromazine

can decrease the self-administration of low to moderate doses of cocaine (ascending limb of the dose-effects function), while not increasing the intake of higher doses (descending limb). In addition,

chlorpromazine can also attenuate the discriminative stimulus effects

of cocaine (Jarbe 1978). Thus, chlorpromazine attenuates the ratedecreasing effects of cocaine, decreases the self-administration of

low to moderate doses, and attenuates the stimulus functions of the

drug. Consequently, the effect of chlorpromazine on cocaine selfadministration appears to be related to a decrease in both the

reinforcing efficacy and stimulus effects of cocaine.

188
In summary, drugs that are abused, including cocaine, have at least

three behavioral effects in common. They can modify the ongoing

rate of behavior, they can have discriminative stimulus functions, and

they are self-administered. The relative importance of these three

behavioral effects in determining the abuse liability of cocaine

requires further investigation. A neurobiological approach may

provide the answers to some unresolved questions concerning the

relative importance of these three behavioral properties as well as

the neurobiological constituents of these behavioral effects.

NEUROBlOLIGlCAL COMPONENTS OF COCAINE SELFADMINISTRATlON

The neurobiological components of a phenomenon include: (1) the

delineation of circuits and pathways involved in the event; (2) specification and localization of critical neurotransmitter receptors; and

(3) evaluation of the influence of additional afferents and efferents.

Examples of several procedures and the results that can be obtained

concerning cocaine abuse are presented in several chapters in this

monograph. This chapter will focus on the use of neurotoxin lesions

to delineate the neurobiological aspects of the behavioral effects of

cocaine.

Since a more complete presentation of this topic is included in the

monograph (Koob and Hubner, this volume), a brief description and

preliminary report using a procedure to simultaneously assess the

neurobiological components of responding maintained by several

reinforcers follows.

Rats were trained on a concurrent chained fixed-ratio (FR) 1, FR 9

(30-second timeout, 100-second limited hold) schedule using an

operant conditioning chamber containing three retractable levers.

Initially the three levers were available. A response on one of them

(food, water, or drug) made the other two retract and set the

contingency for the chosen lever as FR 9. Responding on the drug

lever resulted in a 0.2-ml infusion of cocaine, responding on the food

lever produced a 45-mg food pellet, and pressing the water lever

resulted in presentation of a 0.1-ml water dipper.

The rats were first trained on the schedule using only the food and

water levers with the cocaine lever retracted. They were implanted

with chronic intravenous jugular catheters as well as bilateral guide

cannulae, which were aimed into the central medial nucleus accumbens. Following recovery from surgery, the drug lever was extended

189
into the chamber, and a cocaine option was added to the food and

water schedule. Since a preliminary study showed that rats given

continuous access to cocaine would overdose themselves within 15

days (Dworkin et al. 1987), a restricted access condition to cocaine

was imposed. The cocaine lever was available from 9 a.m. until

3 p.m., and the food and water levers were continuously available.

The initial dose of cocaine used to engender and maintain responding

on the cocaine lever was 0.33 mg/infusion. The effects of substituting other doses of cocaine (0.08 to 0.83 mg/infusion) on responding maintained by the three reinforcers were determined (figure

1). Changing the cocaine dose that was available did not alter

responding on either the food or water lever. However, the rate of

self-administration showed an inverted V-shaped function. The highest

rate of self-administration occurred at the 0.17 mg/infusion dose.

Substitutions of saline for cocaine in the injection system resulted in

a decrease in responding on the cocaine lever and no significant

changes in responding on the food and water levers.

The effects of 6-hydroxydopamine (6-OHDA) lesions of the nucleus

accumbens on responding maintained by the three reinforcers were

determined. The rats received two bilateral injections of 4 g in

0.5 I over 7.5 minutes after desipramine pretreatment (to protect

noradrenergic neurons). The lesions were separated by 2 days.

Following recovery from the lesions, the animals were again placed

into the three-lever chamber and allowed to self-administer food,

water, and cocaine. The lesions did not alter responding maintained

by either food or water. However, the lesions flattened the doseeffect curve for cocaine self-administration by producing a specific

effect on the ascending limb of the curve.

These investigations of the effects of 6-OHDA lesions on responding

simultaneously maintained by food, water, and cocaine indicated that

the lesions may have specifically attenuated the reinforcing effects of

cocaine. The decrease in cocaine self-administration was not related

to a nonspecific motor effect, since no effect was observed on foodor water-maintained responding. Furthermore, the lesions did not

alter the unconditioned effects of cocaine on the descending limb of

the self-administration dose-effect curve. Thus, the dopaminergic

innervations of the nucleus accumbens destroyed by the injection of

6-OHDA appear to be involved in regulating cocaine intake as a

result of direct modulation of the reinforcing effects of the drug.

190
FIGURE 1. The mean number of ratios completed on the food, water,

and cocaine levers during a 24-hour period as a function

of the dose of cocaine that was available for selfadministration

NOTE: Points collected before the lesions are indicated by the open circles, and the

open squares Indicate the effects of the lesions. The vertical lines show 1

SEM. The data presented are the means from three rats exposed to each dose

for at least 3 days. Points above "S" show the effects of saline infusions.

191
NEUROLOGICAL DETERMINANTS OF THE RATE-DEPENDENT

EFFECTS OF COCAlNE

In self-administration studies, the effects of neurotoxin lesions are

not isolated from the influence of the reinforcing effects of the drug.

Therefore, a second behavioral procedure was used to investigate the

potential effects of the lesions on modifying the rate-dependent

effects of cocaine. Rats were trained on what has become a standard

operant baseline to investigate the rate-dependent effects of abused

drugs. A multiple fixed-interval (FI), 4-minute FR, 15-response

schedule of food presentation was used to maintain responding on a

lever. At the start of the session, the first response after 4 minutes

resulted in a food pellet delivery, and the next food pellet was

delivered after the rat completed 15 responses. The rats were given

60 seconds from the end of the FI schedule or start of the FR to

complete the schedule component (limited hold). lf the schedule

requirement was not completed by the end of the limited hold, the

other component was scheduled. The schedule components alternated

until 10 components of each were presented.

The schedule parameters used generated two different mean rates of

responding. Responding maintained by the interval schedule was

significantly lower than responding maintained by the ratio. However, cocaine resulted in similar response-rate effects on performance generated by both schedules. Doses of cocaine from 5.6 to 17

mg/kg had little or no effect on response rates, and the largest dose

investigated generally decreased responding (figure 2).

After the prelesion dose-effect curves were determined, the rats were

pretreated with desipramine and lesioned with 6-OHDA. Bilateral

injections of 4 μg in 0.5 μl of the neurotoxin or its vehicle were

delivered into the nucleus accumbens. The dose-effect curves were

again determined following recovery from the lesion. The lesion did

not result in any significant changes in the effects of cocaine on

food-maintained responding. However, there was a trend for cocaine

to result in a slightly greater decrease in responding after the lesion.

NEUROBIOLOGICAL COMPONENTS OF THE DISCRIMINATIVE

STlMULUS EFFECTS OF COCAlNE

Rats were trained to discriminate cocaine (10 mg/kg IP) from saline

using a standard two-lever procedure. Every 10th response (FR 10)

on one of the levers (drug lever) resulted in the delivery of a food

pellet if cocaine was injected before the session. Responding on the

192
FIGURE 2. Dose-response curves for the effects of cocaine on

responding maintained by the multiple Fl 4-minute,

FR 15 schedule of food presentation

NOTE: The circles Indicate the effects of the drug before the lesion, and the squares

show the postlesion effects. The points are the means from six rats, and the

vertical lines indicate ±1 SD. The top panels show the effect on re-sponding

maintained by the interval schedule, and the bottom panels indicate the effects

of cocaine on responding maintained by the ratio schedule. The left and right

panels show the effects of a vehicle and 6-OHDA lesion, respectively.

other lever (saline lever) resulted in reinforcement if the rat was

pretreated with saline. Training was continued until the rats emitted

the criteria of three or fewer responses on the incorrect lever before

the first food presentation and more than 95 percent of the responses

during a session occurred on the correct lever for five consecutive

sessions.

The subjects were then tested using a multiple dosing procedure.

The rats were injected with saline and immediately placed in the

conditioning chamber. The session was started 1 minute following the

injection and was terminated after 30 food pellets were obtained or 4

193
minutes of the session had elapsed. Five minutes after the first

injection, the rats were injected with 0.82 mg/kg of cocaine, and a

second session was started after 1 minute. This procedure was repeated with additional injections of 0.36, 0.50, 0.72, 1.0, 1.15, 2.1, and

3.0 mg/kg of cocaine 1 minute prior to the start of the third through

ninth sessions. During testing sessions, responding on both levers

was reinforced on an FR 10 schedule. Daily testing sessions were

terminated when the subjects completed the first ratio on the drug

lever after two consecutive injections. Training sessions were

occasionally run between testing sessions.

Each rat was tested at least eight times before being lesioned with

6-OHDA or its vehicle. Bilateral injections of 6 μg in 0.4 ml over

7.5 minutes were made into the nucleus accumbens, following pretreatment with desipramine. Three days were allowed for recovery

from the lesion. The animals were then tested again using the

multiple dosing procedure for four consecutive testing sessions.

The multiple dosing procedure resulted in generalization gradients

similar to those obtained when only single injections per day were

evaluated (figure 3). All but one of the rats tested chose the saline

lever after saline was injected and switched to the cocaine lever

after the fifth cocaine injection. The vehicle lesion did not result in

any significant changes in the discriminative stimulus properties of

cocaine.

In contrast to sham animals, the 6-OHDA lesion severely attenuated

the stimulus effects of cocaine. After the lesion, two of the subjects

occasionally selected the cocaine lever after the first (saline)

injection. In addition, these rats also completed the first ratio of

the session on the cocaine lever after receiving low doses of the

drug. Three rats, however, continued to choose the saline lever after

saline injections and persisted in choosing the saline lever or showed

no preference for either lever after additional injections of cocaine.

Similar changes were seen in measures of the percent of responding

on the drug lever during the testing sessions. Furthermore, neither

the multiple injections nor the lesions produced any significant

changes in the rate of responding during the sessions. Thus, the

lesion produced a selective attenuation of the discriminative stimulus

functions of the drug.

194
FIGURE 3. Generalization gradients indicating the effects of

consecutive doses of cocaine

NOTE: Data presented indicate the percent of sessions during which the rats

completed their first ratio on the drug lever. The data collected from each

rat are indicated by the individual points, and the lines indicate the mean of

data collected from all subjects. The top left panel shows the gradient

obtained before the lesion. Data presented in the middle panel were collected

after a vehicle lesion, and the bottom panel shows the effects of the neurotoxin lesion.

195
CONCLUSION

The dopaminergic system innervating the nucleus accumbens is clearly

involved in modulating both the reinforcing and discriminative stimulus functions of cocaine. Selective lesions of this system attenuate

cocaine self-administration while having little or no effect on food or

water intake. Furthermore, rates of responding maintained by food

and water presentations and the effects of cocaine on food- and

water-maintained responding are not affected by the lesion. Cocaine

self-administration involves the reinforcing, rate-modulating, and

discriminative stimulus effects of the drug. lt appears, however, that

the reinforcing effect of the drug is more a function of the stimulus

functions of the drug than its effects on ongoing rates of responding.

Schedule-controlled behavior, however, does provide an excellent

screen to evaluate the potential for a drug to attenuate selfadministration. Both chlorpromazine and benzodiazepines alter the

effects of cocaine on schedule-controlled behavior and attenuate

cocaine self-administration. Evaluations of the effects that phamacologic and neurobiologic perturbations have on the behavioral effects of cocaine increase our understanding of the behavioral and

neurobiological aspects of drug abuse.

REFERENCES

Aston-Jones, S.; Aston-Jones, G.; and Koob, J.F. Cocaine antagonizes

the anxiolytic effects of ethanol. Psychopharmacology (Berlin)

84:28-31, 1984.

Barrett, J.E. Effects of alcohol, chlordiazepoxide, cocaine and

pentobarbital on responding maintained under fixed-interval

schedules of food or shock presentation. J Pharmacol Exp Ther

196:605-615, 1976.

Brady, J.V., and Lukas, S.E., eds. Testing Drugs for Physical

Dependence Potential and Abuse Liability. National Institute on

Drug Abuse Research Monograph 52. DHHS Pub. No. (ADM) 84

1332. Washington, DC: Supt. of Docs., U.S. Govt. Print. Off., 1984.

147 pp.

Dworkin, S.I.; Goeders, N.E.; Grabowski, J.; and Smith, J.E. The

effects of 12-hour limited access to cocaine: Reduction in drug

intake and mortality. In: Harris, L.S., ed. Problems of Drug

Dependence 1986. National Institute on Drug Abuse Research

Monograph 76. DHHS Pub. No. (ADM) 87-1508. Washington, DC:

Supt. of Docs., U.S. Govt. Print. Off., 1987. pp. 221-225.

196
Ford, R.D.; Rech, R.H.; and Tobin, D. Interactions between stimulants

and depressants on schedule-controlled behavior. Pharmacol

Biochem Behav 12:657-661, 1980.

Gill, C.A.; Holz, W.C.; Zirkle, C.L.; and Hill, H. Pharmacological

modification of cocaine and apomorphine self-administration in the

squirrel monkey. In: Deniker, P.; Radouco-Thomas, C.; and

Villeneuve, A., eds. Proceedings of the Xth lnternational Congress

of the Collegium Internationale Neuro-Psychopharmacologicurn.

Oxford: Pergamon Press, 1977. pp. 1477-1484.

Gonzalez, F.A., and Goldberg, S.R. Effects of cocaine and damphetamine on behavior maintained by various schedules of food

presentation in squirrel monkeys. J Pharmacol Exp Ther 201:33-43,

1977.

Herling, S., and Woods, J.H. Chlorpromazine effects on cocainereinforced responding in rhesus monkeys: Reciprocal modification

of rate-altering effects of the drugs. J Pharmacol Exp Ther

214:354-361, 1980.

Iglauer, C., and Woods, J.H. Concurrent performances: Reinforcement by different doses of intravenous cocaine in rhesus monkeys.

J Exp Anal Behav 22:179-196, 1974.

Jarbe, T.U.C. Cocaine as a discriminative cue in rats: Interactions

with neuroleptics and other drugs. Psychopharmacology (Berlin)

59:183-187, 1978.

Johanson, C.E. Effects of intravenous cocaine, diethylpropion,

d-amphetamine and perphenazine on responding maintained by food

delivery and shock avoidance in rhesus monkeys. J Pharmacol Exp

Ther 204:118-129, 1978.

Johanson, C.E., and Schuster, C.R. A choice procedure for drug

reinforcers: Cocaine and methylphenidate in the rhesus monkey.

J Pharmacol Exp Ther 193:676-688, 1975.

Roberts, D.C.S., and Vickers, G. Atypical neuroleptics increase

self-administration of cocaine: An evaluation of a behavioral

screen for antipsychotic activity. Psychopharmacology (Berlin)

71:83-89, 1984.

Tessel, R.E., and Barrett, J.E. Antagonism of the behavioral effects

of cocaine and d-amphetamine by prazosin. Psychopharmacology

(Berlin) 90:436-440, 1986.

Thompson, T.; Dews, P.B.; and McKim, A. Advances in Behavioral

Pharmacology. Vol. 111. New York: Academic Press, 1981. 217

pp.

Wilson, M.C., and Schuster, C.R. Interactions between atropine,

chlorpromazine and cocaine on food reinforced behavior. Pharmac

Biochem Behav 3:363-375, 1975.

197
Woods, J.H.; Winger, G.D.; and France, C.P. Reinforcing and

discriminative stimulus effects of cocaine: Analysis of pharmacological mechanisms. In: Fisher, S.; Raskin, A.; and Uhlenhuth,

E.H., eds. Cocaine: Clinical and Biobehavioral Aspects. New York

Oxford University Press, 1987. pp. 21-65.

ACKNOWLEDGMENTS

This research was supported in part by U.S. Public Health Service

grants DA 03631 and DA 03628 from the National Institute on Drug

Abuse.

AUTHORS

Steven I. Dworkin, Ph.D.

James E. Smith, Ph.D.

Psychiatry Research Unit

Departments of Psychiatry and Pharmacology

Louisiana State University

School of Medicine in Shreveport

P.O. Box 33932

Shreveport, LA 71130-3932

198
Intracranial Cocaine SelfAdministration

Nick E. Goeders

INTRODUCTlON

The nonmedical use of cocaine has continued to increase during the

last decade, while the use of other illicit drugs has steadily declined

since 1979. While the neurobiological mechanisms potentially involved

in the complex processes mediating cocaine reinforcement have been

under vigorous examination, they have not yet been conclusively

elucidated. The intravenous self-administration paradigm has been the

most frequently employed laboratory tool used to investigate cocaine

reinforcement, since this route most closely parallels human selfadministration. Neuronal pathways and circuits that may be involved

in cocaine reinforcement processes can be identified by assessing the

effects of specific neurotransmitter antagonists or neurotoxin lesions

on self-administration as described elsewhere in this volume. However, the distinct brain sites responsible for the initiation of neuronal

activity in these reinforcement-related pathways and circuits

following the response-contingent administration of cocaine may be

difficult if not impossible to identify following systemic delivery.

With systemic self-administration, it is difficult to conclude accurately whether pharmacological or neurobiological treatments specifically

affect the reinforcing properties of the drug or whether they affect

other processes such as rate of responding, conditioned responses to

the drug, ability to discriminate the presence of the drug, or changes

in heart rate, blood flow, or drug metabolism and elimination. These

and other factors can nonspecifically influence self-administration and

may result in potentially erroneous conclusions. Intracranial selfadministration (ICSA) offers an exciting alternative procedure for

providing valuable information that can be used to complement

intravenous self-administration data for a better understanding of the

neurobiological mechanisms of drug reinforcement. With ICSA, the

199
completion of the appropriate response results in the delivery of the

drug directly into a discrete brain region. If interactions of the drug

with receptors localized within the injection locus result in the

activation of neuronal systems associated with reinforcement, then

these intracranial microinjections should maintain responding and,

therefore, circumvent many of the problems inherent in systemic drug

delivery.

Although the full potential for ICSA has only recently been realized,

the idea of the direct self-administration of a drug into brain tissue

dates back almost 30 years (Olds and Olds 1958; Olds 1962). Intraventricular self-administration of drugs (Gustafson and Pickens 1975;

Amit et al. 1976; Belluzi and Stein 1977; Tortella and Moreton 1980;

Brown et al. 1979) has been demonstrated as has the direct intracranial self-administration of opioids, psychomotor stimulants, and

putative neurotransmitters. Among the opioids, morphine is selfadministered into the lateral hypothalamus (Stein and Olds 1977; Olds

1979), the ventral tegmental area (Bozarth and Wise 1980; Bozarth

and Wise 1981), and the nucleus accumbens (Olds 1982), while D-ala2-

methionine enkephalinamide, a stable enkephalin analog, maintains

ICSA into the lateral hypothalamus (Olds and Williams 1980). and

methionine-enkephalin, an endogenous ligand for delta opiate

receptors, is self-administered into the nucleus accumbens (Goeders et

al. 1984). With respect to putative neurohumors, neurotensin supports

ICSA into the ventral tegmental area (Glimcher et al. 1983) while

cholecystokinin maintains ICSA into the nucleus accumbens (Hoebel

and Aulisi 1984). Dopamine is self-administered into the nucleus

accumbens (Guerin et al. 1984; Dworkin et al. 1986) and preliminary

data suggest that glutamate maintains ICSA into the ventral pallidum

and preoptic area (Goeders et al. 1987). The psychomotor stimulants

amphetamine and cocaine are self-administered into the mesolimbic/

mesocortical dopaminergic system. Cocaine maintains ICSA into the

medial prefrontal cortex (Goeders and Smith 1983), while amphetamine

is self-administered into the orbitoprefrontal cortex of rhesus

monkeys (Phillips and Rolls 1981) and the nucleus accumbens of rats

(Monaco et al. 1981; Hoebel et al. 1983). The remainder of this

chapter will focus on what has been learned about the central

mechanisms of cocaine reinforcement using the intracranial selfadministration approach.

DRUG DELIVERY SYSTEMS

ICSA is an elegant methodology for demonstrating that a discrete

brain region is involved in the initiation of reinforcing neuronal

200
activity following the response-contingent administration of a drug.

By restricting the delivery of the drug to potentially reinforcementrelevant receptor populations, this procedure may minimize many of

the peripheral effects resulting from systemic drug administration.

However, it is important to recognize some of the limitations that

may be involved in the use of this technique and in the interpretations of the data obtained. To engender and subsequently

maintain responding, the substance must be delivered directly to the

reinforcement-relevant receptor populations with minimal delay. An

ideal drug delivery system is one that rapidly dispenses a small,

reproducible volume under moderate pressure. Large injection

volumes (i.e., greater than 500 nl) can result in nonspecific local

tissue damage at the injection site and backflow of the injectate

(Olds 1962). Diffusion of the injectate is directly related not only to

the infusion volume but also to the rate of infusion (Myers 1974;

Myers and Hoch 1978; Routtenberg 1972). lf there is a significant

diffusion from the site of injection, the precise identification of the

reinforcement-relevant receptors may be severely compromised. To

avoid mechanical damage to tissue and significant diffusion from the

injection site, incredibly small injection volumes (i.e., less than

1/2,000 ml) must be accurately delivered.

Drug delivery systems similar to those used for intravenous selfadministration have also been utilized for ICSA. These systems

consist of a microsyringe controlled by a motor-driven syringe pump,

which is activated for a predetermined duration each time that the

animal completes the appropriate response. While these delivery

systems are relatively accurate when used with restrained animals,

their use for discrete, small-volume intracranial injections in

relatively unrestrained subjects may lead to inaccurate drug delivery

due to mechanical effects of the tubing used to connect the microsyringe to the injection cannula. Electrolytic microinfusion systems

using chemitrodes (Criswell 1977) circumvent many of the problems

experienced with small-volume drug delivery.

An electrolytic drug delivery system consists of an airtight drug

reservoir, containing two electrodes, which is attached to an injection

cannula that mounts directly on a guide cannula on the animal’s head.

Microinfusions are produced by passing a direct current between the

two electrodes, resulting in the evolution of hydrogen gas which, in

turn, increases the pressure in the reservoir and forces a reproducible

volume of drug solution out through the injection cannula. The

amount of solution infused is proportional to the current intensity

and duration. With the electrolytic systems, movement does not

201
affect drug delivery, and continuity between responding and reinforcer presentation is achieved. Electrolytic microinfusion systems were

used in all the cocaine intracranial self-administration experiments

described below.

The first series of experiments determined the accuracy and dependability of the electrolytic microinfusion system for cocaine ICSA

(Goeders et al. 1985) by using tracer amounts of radiolabeled cocaine

mixed with unlabeled cocaine. The injection cannula was placed into

fresh brain tissue, and varying intensities of either an injection

current or a quiescent current were passed between the electrodes.

The tissue was then solubilized, and the radioactivity was determined

with liquid scintillation spectrophotometry. This calibration demonstrated that an injection current of 200μA with a 6-μA quiescent

current delivered reliable 100-nl microinfusion volumes (± 7 percent)

with no leakage between injections.

Next, the extent of diffusion of the drug from the injection site was

determined. Rats received 4μl of 3H-cocaine in 40 infusions of 100

nl each delivered over 5.5 seconds on a random time, 4-minute

schedule (similar to the maximum rate of self-administration) into a

self-administration site in the medial prefrontal cortex (Goeders and

Smith 1985). The animals were sacrificed by total immersion in liquid

nitrogen 30 seconds following the final injection. Ninety-three

percent of the radioactivity was recovered within 1 mm3 of the

cannulae tips, suggesting that the reinforcing actions of cocaine are

probably localized within the target area in the medial prefrontal

cortex and not at distal sites as a result of drug diffusion. A cyclic

voltagram demonstrated that the cocaine was not electrochemically

altered by the current intensities produced by the electrolytic

microinfusion system. These initial studies demonstrated that the

electrolytic microinfusion system is the ideal drug delivery system for

investigating intracranial cocaine self-administration.

MAPPING INTRACRANIAL COCAINE SELF-ADMINISTRATION

Although the effects of cocaine on neurotransmission are complex,

the primary neurochemical action appears to be an inhibition of

biogenic amine neurotransmitter uptake into presynaptic nerve

endings. Results obtained from intravenous self-administration

experiments have demonstrated a primary role for the mesolimbic/

mesocortical dopaminergic neuronal system in cocaine reinforcement

(Koob and Hubner, this volume). Therefore, initial behavioral

experiments were designed to investigate whether intracranial

202
injections of cocaine would maintain self-administration into various

target sites located within the mesolimbic dopaminergic system

(Goeders and Smith 1983). Rats were stereotaxically implanted

unilaterally with guide cannulae into the nucleus accumbens, ventral

tegmental area, or medial prefrontal cortex. The animals were housed

on a reversed 12-hour light-dark cycle and tested for intracranial

self-administration at the start of their active cycle. Sessions were

terminated either after 8 hours or after 40 injections were delivered

to minimize potential cumulative cytotoxic effects. A red light above

the response lever was illuminated at the start of the session, and

each response in the presence of the red light resulted in a 5.5

second infusion of 100 nl of the drug. During the infusion, the red

light was extinguished, a white light was illuminated, and a tone was

presented. Each microinfusion was followed by a 30-second timeout

period during which both stimulus lights were extinguished, and

responses were monitored but had no scheduled consequences. Concentrations of 0 to 5,000 pmol of the drug dissolved in artificial

cerebrospinal fluid were tested in the three brain regions.

Microinjections of cocaine into the nucleus accumbens or ventral

tegmental area did not maintain responding at any concentration

tested, but microinfusions into the medial prefrontal cortex resulted

in a rapid acquisition of lever pressing. These results were surprising

in light of intravenous self-administration data, which suggest a

major role for the nucleus accumbens in cocaine reinforcement

processes. Therefore, a second group of animals implanted with

cannulae into the nucleus accumbens was trained to lever-press

before being retested for intracranial self-administration (Goeders and

Smith 1987). These rats were originally tested for intracranial selfadministration; they were then food-deprived to 80 percent of their

ad lib weights and allowed to respond on a variable interval,

30-second schedule of food reinforcement in the same experimental

chamber. When stable baselines of responding were observed

(approximately 1 week), the food magazine was removed from the

chamber, and the microinjection system was secured to the rat’s head

for ICSA testing. However, even after lever-pressing training, these

rats did not self-administer cocaine, suggesting that cocaine does not

initiate reinforcing neuronal activity by its response-contingent

application into the nucleus accumbens.

However, all rats with cannulae implanted into the medial prefrontal

cortex self-administered cocaine, with an average of six behavioral

sessions required to engender stable rates of self-administration at

the training dose of 50 pmol/100 nl infusion. When lever pressing

203
stabilized for at least three consecutive sessions at this dose, doseresponse curves were determined by increasing or decreasing the drug

concentrations.

Self-administration was dose related, with maximal rates of responding obtained with 50 to 90 pmol/infusion. Dose-response relationships

were unique for each animal, with responding maintained over either

narrow or wide ranges in different subjects. Large changes in drug

intake often occurred after increasing or decreasing the concentration

by as few as 5 or 10 pmol. Rates of self-administration declined as

the dose was either increased or decreased from optimum levels. An

empirical measure of the patterns of self-administration was therefore

employed to analyze the self-administration data (Goeders et al.

1986). The interinfusion intervals were counted separately in four

15-minute divisions (0 to 15 minutes, 15 to 30 minutes, 30 to 45

minutes, and 45 to 60 minutes), a frequency distribution was

generated for each session, and relative frequencies were calculated.

The significance of difference in the mean relative frequency distributions of the interinfusion intervals for the various treatment

conditions was evaluated with a two-factor randomized analysis of

variance followed by Tukey’s all pairwise comparisons among

treatment means.

A two-lever discrimination procedure was used to demonstrate that

self-administration resulted from the reinforcing properties of cocaine

and not from a nonspecific increase in motor activity (Goeders and

Smith 1983). For these lever-reversal experiments, an identical but

inactive lever was installed on the opposite wall of the chamber, with

responses being recorded but having no scheduled consequences.

When responding stabilized, active contingencies were alternated

between the right and left sides of the chamber. In this two-lever

choice procedure, responding rapidly increased on the active lever.

When the active and inactive levers were reversed, the rats switched

responding to the new active manipulandum. Extinction (i.e., the

substitution of vehicle for cocaine) resulted in a loss of this preference, and the animals responded at equally low rates on both levers.

During reconditioning, when cocaine microinfusions were again made

available, the animals rapidly increased responding primarily on the

active lever. Since responding occurred significantly more often on

the active manipulandum during several reversals, these data suggest

that the self-administration did not result from a generalized,

nonspecific behavioral stimulant action of cocaine. Furthermore, in

some animals, the response requirement was gradually increased from

204
1 to 10 lever presses per microinfusion, and the animals correspondingly increased their rates of responding to maintain similar interinfusion intervals and patterns of self-administration. These data

demonstrate that the response-contingent delivery of cocaine into the

medial prefrontal cortex does serve as a reinforcer.

NEUROPHARMACOLOGY

In addition to being classified as a psychomotor stimulant, cocaine

also possesses local anesthetic properties. Therefore, the next series

of experiments tested whether the local anesthetic properties of

cocaine were responsible for the ICSA behavior (Goeders et al. 1986).

Rats were trained to self-administer cocaine into the medial prefrontal cortex, and dose-response curves were determined for each animal. Equimolar concentrations of lidocaine HCI and nine times

equimolar concentrations of lidocaine HCI were then substituted for

the optimum dose of cocaine (the dose resulting in the highest and

most stable drug intake). Lidocaine was used in these experiments

because, unlike other local anesthetics (e.g., procaine, chloroprocaine), it does not appear to have reinforcing properties. However,

this substitution resulted in rates of drug intake similar to vehicle,

with patterns of responding resembling those seen with either a very

low dose of cocaine or the vehicle. These data suggest that the

reinforcing properties of cocaine are unrelated to its local anesthetic

effect.

The neurotransmitter receptors involved in the intracranial selfadministration of cocaine were investigated using specific receptor

antagonists. Rats were trained to self-administer cocaine into the

medial prefrontal cortex, and dose-response curves were determined

for each animal as described above. Equimolar concentrations of the

D-2 dopaminergic receptor antagonist sulpiride, the D-1 dopaminergic

receptor antagonist SCH 23390, the beta-noradrenergic receptor

antagonist dl-propranolol hydrochloride, or the muscarinic cholinergic

receptor antagonist atropine sulfate were coadministered in a random

order with the optimum dose of cocaine. If an attenuation of selfadministration was observed, the concentration of cocaine was

doubled, and the effects of the antagonist were again determined. If

no change in intake occurred, then the concentration of the antagonist was doubled and the effects again determined. Sulpiride significantly decreased drug intake to 47 percent of baseline levels and

generated patterns of self-administration (i.e., mean relative frequency

distributions) consistent with those seen when the concentration of

cocaine was decreased from optimum levels. This effect persisted for

205
two sessions, but drug intake returned to baseline by the third

posttreatment session. Doubling the concentration of cocaine did not

alter the sulpiride effect, suggesting a noncompetitive action at

separate receptor sites. The administration of the other antagonists

did not alter drug intake directly. Atropine had no effect on rates

or patterns of self-administration during the session it was delivered

but significantly decreased drug intake during the first posttreatment

session, with patterns of responding consistent with those seen with

higher doses of cocaine. These effects dissipated by the second

posttreatment session and were not altered by doubling the concentration of atropine. Similar results were observed with equimolar

and twice-equimolar concentrations of SCH 23390. Propranolol,

however, did not affect the rates or patterns of self-administration at

any concentration tested.

These results suggest that D-2 dopaminergic receptors in the medial

prefrontal cortex appear to be excitatory to cocaine reinforcement

processes, since equimolar concentrations of sulpiride reduced selfadministration to vehicle levels with patterns of responding consistent

with decreasing the cocaine dose. Cocaine and sulpiride may act at

different receptor sites, since the attenuation of self-administration

after D-2 receptor blockade was not reversed by increasing the cocaine dose. On the other hand, the D-1 dopaminergic receptor

antagonist SCH 23390 and the muscarinic cholinergic receptor

antagonist atropine had either no effect or minimal effects on

intracranial self-administration. However, the blockade of these

receptors resulted in a delayed attenuation of drug intake, with

patterns of responding resembling those seen with an increased dose

of cocaine (i.e., a potentiation of cocaine-initiated neuronal activity).

Beta-noradrenergic systems in the medial prefrontal cortex may not

be involved in cocaine reinforcement.

6-HYDROXYDOPAMINE LESIONS

These data suggest that the initiation of cocaine reinforcement

processes results, in part, from direct or indirect interactions of the

drug with D-2 dopaminergic receptors in the medial prefrontal cortex.

To determine whether the reinforcement-relevant receptors are

localized on presynaptic dopaminergic terminals or on postsynaptic

neurons, the self-administration site was lesioned with the neurotoxin

6-hydroxydopamine (Goeders and Smith 1986). Rats were implanted

with guide cannulae into the medial prefrontal cortex and trained to

intracranially self-administer cocaine as described above. When dose

effect analyses were completed and stable baselines of drug intake

206
obtained, animals were pretreated with desmethylimipramine to inhibit

uptake into noradrenergic neurons followed by unilateral 6-hydroxydopamine injections into the medial prefrontal cortex at the selfadministration site. In postlesion self-administration sessions, various

concentrations of cocaine were tested for self-administration in each

rat, and the rates and patterns of intake at each concentration were

compared with those obtained prelesion.

The lesion resulted in a 45 percent decrease in the content of

dopamine in the medial prefrontal cortex, while the levels of the

other biogenic amine neurotransmitters serotonin and norepinephrine

were not affected. However, there may have been a hyperactivity of

serotonergic neuronal activity on the nonlesion side, since the

content of serotonin was increased 85 percent compared to control

animals. No changes were observed in neurotransmitter content in

the nucleus accumbens, suggesting that the lesion was localized to

neurons in the medial prefrontal cortex.

The 6-hydroxydopamine lesion at the site of self-administration

reduced drug intake to vehicle levels and flattened the dose-response

curves for all animals. Prelesion rates of self-administration did not

return during the 6 to 8 weeks postlesion that the behavior was

monitored, and responding could not be reinitiated by either

increasing or decreasing the cocaine concentration. However, the

substitution of dopamine (300 pmol) for cocaine resulted in a rapid

reacquisition of lever pressing almost to prelesion baseline levels,

with rates and patterns of responding similar to those maintained by

the high or optimum dose of cocaine prior to the lesion. This

dopamine self-administration was probably mediated through postsynaptic D-2 dopaminergic receptors, since blockade with the

coinfusion of sulpiride reduced intake to vehicle levels. Serotonergic

receptors do not appear to be involved, since the substitution of 300

pmol of this neurotransmitter would not maintain responding in the

lesioned animals. The involvement of serotonin cannot be ruled out,

however, since different concentrations of the neurotransmitter were

not tested in these animals. In summary, these experiments

demonstrated that dopaminergic innervations are necessary for ICSA

of cocaine in the medial prefrontal cortex. The 6-hydroxydopamine

lesions that damaged presynaptic dopaminergic terminals eliminated

the primary mechanism for cocaine-induced initiation of reinforcing

neuronal activity. Postlesion response-contingent microinfusions of

dopamine appeared to activate postsynaptic neuronal systems in a

manner analogous to the delivery of cocaine prelesion.

207
NEUROTRANSMITTER TURNOVER

A final set of experiments used neurotransmitter turnover procedures

to investigate whether the response-contingent delivery of cocaine

into the medial prefrontal cortex results in the activation of neuronal

activity mediated through other brain regions. Rats were implanted

unilaterally with cannulae into the right medial prefrontal cortex.

One group of rats was allowed to self-administer cocaine, while

another group served as a yoked-vehicle control. Depression of the

lever by a rat in the self-administration group simultaneously

delivered a microinfusion of artificial cerebrospinal fluid to the

yoked-vehicle rat contained in a chamber similar to the selfadministration chamber. Depressions of the lever by the yokedvehicle animals were counted but had no scheduled consequences,

although all stimulus changes were identical. After stable baselines

of responding were observed at the optimum concentration of cocaine

in the self-administration group, all rats were implanted with jugular

catheters using previously reported methods (Weeks 1962; Weeks

1972). The catheter was sealed at the end and enclosed in a lightweight aluminum backpack (Lane et al. 1982). Following a 1-week

recovery from surgery, the rats were allowed to self-administer

cocaine at the optimum concentration until stable rates of drug

intake were obtained. Two days prior to sacrifice, the patency of

each jugular catheter was determined by injecting 100μl of

methohexital. An immediate behavioral depression indicated that the

catheter was functional. The catheters were resealed, and the

animals were returned to their home cages.

Immediately prior to the start of the experimental session, one of the

self-administration animals and its yoked-vehicle control received 0.5

mCi of L-[U-3H] tryptophan and 1.0 mCi of L-[2,6-3H]-tyrosine

injected in 100μl of heparinized saline through the jugular

catheters. Two timepoints (60 and 90 minutes) were used to determine turnover following the administration of the radioactive

precursors. These timepoints were chosen because they have been

demonstrated to be on the linear portion of the decay in radioactivity

curves for the biogenic amine neurotransmitters (Smith and Lane

1983). Self-administration rats and yoked-vehicle rats were simultaneously sacrificed at the appropriate pulse times by total immersion

in liquid nitrogen in combination operant conditioning/liquid nitrogen

immersion chambers (Smith et al. 1977). The heads were warmed in

-18 °C cryostat, and the brains were removed and cut in the coronal

plane into 500- μm serial sections. The right and left sides of the

medial prefrontal cortex and nucleus accumbens were microdissected

208
from these coronal sections. The contents of norepinephrine, dopamine, serotonin, tyrosine, and 3,4-dihydroxyphenylacetic acid (DOPAC)

were measured using high-pressure liquid chromatography with electrochemical detection, and specific radioactivity was determined using

liquid scintillation spectrophotometry (Co et al. 1982).

Turnover rates were calculated (Smith et al. 1982) and expressed as

pmol mg protein-1 hr-1.

No significant differences in neurotransmitter content were observed

between treatment groups in either brain region (tables 1 and 2).

Increased levels of tyrosine and DOPAC were seen in the right medial

prefrontal cortex compared to the left in both self-administration rats

and yoked-vehicle rats (table 1), suggesting that dopamine utilization

is greater on the right side of the brain compared to the left.

Significant changes in neurotransmitter turnover were observed in

both medial prefrontal cortex and nucleus accumbens as a result of

the response-contingent intracranial delivery of cocaine, but only on

the side of the brain containing the injection cannula. At the site of

self-administration in the right medial prefrontal cortex, decreases in

the utilization of dopamine (-87 percent) and serotonin (-90 percent)

and increases in the utilization of norepinephrine (435 percent) were

found (table 3). In contrast, the turnover rate for dopamine was

increased (128 percent), norepinephrine decreased (-69 percent), and

serotonin unchanged in the right nucleus accumbens (table 4).

The turnover rates for dopamine in the left nucleus accumbens of

both treatment groups and the right nucleus accumbens of the yokedvehicle rats were consistent with the values reported from other

investigations (Smith et al. 1982). However, the utilization of

dopamine was significantly increased in the right nucleus accumbens

of the self-administration rats, suggesting that dopaminergic activity

in this structure was enhanced. The medial prefrontal cortex sends

efferent connections to a number of subcortical structures including

the ventral tegmental area (Leonard 1969) and nucleus accumbens

(Beckstead 1979; Walaas 1981). Cocaine administration into the medial

prefrontal cortex may result in the activation of an ipsilateral

descending neuronal system projecting to one or more of these brain

areas. This, in turn, could result in the activation of the dopaminergic innervations of the nucleus accumbens with increased

synthesis and release of the neurotransmitter. These data are in

agreement with investigations of the effects of 6-hydroxydopamine

lesions of the nucleus accumbens on intravenous cocaine selfadministration, which suggest that the dopaminergic innervations of

209
TABLE 1. Content of dopamine (DA), norepinephrine (NE), serotonin

(5-HT), tyrosine (TYR), and dihydroxyphenylacetic acid

(DOPAC) in right and left medial prefrontal cortex of

rats self-administering cocaine into the medial prefrontal

cortex and in yoked-vehicle rats

Medial

Prefrontal Treatment

Cortex Group DA NE 5-HT TYR DOPAC

Right Side ICSA

Yoke

Left Side ICSA

Yoke

17.6±5.2 14.4±4.2 29.6±5.6 570.7±242 16.5±9.0

16.2±7.0 16.9±5.1 27.9±3.3 602.9±243 18.6±13.3

15.1±1.8 16.6±2.4 32.4±7.5 350.7±80 5.3±2.5

16.3±6.2 17.0±3.1 30.6±5.8 344.1±77 5.4±2.1

NOTE: Values are means ± standard deviations for n=8 in each treatment group,

measured in pmol mg protein-1. Significant differences between the right and

left sides for each treatment group deterrnined with Student’s t-tests were:

p<0.05; +p<0.01.

TABLE 2. Content of dopamine (DA), norepinephrine (NE), serotonin

(5-HT), tyrosine (TYR), and dihydroxyphenylacetic acid

(DOPAC) in right and left nucleus accumbens of rats

self-administering cocaine into the medial prefrontal

cortex and in yoked-vehicle rats

Nucleus Treatment

Accumbens Group DA NE 5-HT TYR DOPAC

Right Side ICSA 321.0±75.9 14.7±4.8 40.0±8 488.8±135 54.0±10.5

Yoke 393.0±104.8 14.9±4.1 43.7±8.5 471.3±131 77.4±24

Left Side ICSA 277.6±44 20.3±8.2 36.6±10.4 455.0±122 44.3±11.1

Yoke 297.3±72 12.3±4.7 38.6±9.8 488.0±146 59.2±15.5

NOTE: Values are means ± standard deviations for n=8 in each treatment group,

measured in pmol mg protein-1.

210
TABLE 3. Turnover rates of dopemine (DA), norepinephrine (NE), and

serotonin (5HT) concurrently measured in right and left

medial prefrontal cortex of rats self-administering

cocaine into the medial prefrontal cortex and in yokedvehicle rats

Medial

Prefrontal

Cortex

Treatment

Group DA NE 5-HT

Right Side ICSA 4.2±3.0+

30.1±8.2+6.2±4.0+

Yoke 31.7±6.4 1.4±0.3 61.0±21.4

Left Side ICSA

Yoke

9.4±9.1 8.2±7.5 31.9±8.1

10.1±5.3 18.7±8.2 16.3±11.8

NOTE: Values are means ± standard deviations for n=8 in each treatment group,

measured in pmol mg protein-1 hr-1 . Significance of the differences determined with Student’s t-tests were: +

p<0.01.

TABLE 4. Turnover rates of dopamine (DA), norepinephrine (NE), and

serotonin (5-HT) concurrently measured in right and left

nucleus accumbens of rats self-administering cocaine into

the medial prefrontal cortex and in yoked-vehicle rats

Nucleus Treatment

Accumbens Group DA NE 5-HT

Right Side ICSA

Yoke

318.0±44 3.1±1.1* 10.1±1.6

139.7±43 10.2±0.3 19.6±15.2

Left Side ICSA

Yoke

152.5±77 ND 13.2±2.1

122.5±52 ND 19.6±12.2

NOTE: Values are means ± standard deviations for n=8 in each treatment group,

measured in pmol mg protein-1 hr-1 . Significance of the differences determined with Student’s t-tests were: *p<0.02; +

p<0.01. ND, data not collected.

211
this structure are involved in cocaine reinforcement (Roberts et al.

1980).

CONCLUSIONS

The response-contingent delivery of cocaine into the medial prefrontal cortex initiates reinforcing neuronal activity directly through the

presynaptic inhibition of the reuptake of dopamine and indirectly

through the postsynaptic activation of dopaminergic receptors by

increased synaptic concentration of the neurotransmitter.

These effects may result in the downregulation of dopamine utilization in the medial prefrontal cortex and the initiation of neuronal

activity in endogenous reinforcement circuits. The various cell bodies

and fiber connections within these pathways may each be involved in

the mediation of cocaine reinforcement processes, with a loss of

function at any point in the circuit detrimental to the full expression

of these effects. While the nucleus accumbens and ventral tegmental

area did not support intracranial self-administration, these data do

not exclude the possibility that other neurotransmitter systems in

other brain regions might maintain this behavior, especially since the

intravenous administration of cocaine results in a much more potent

reinforcing stimulus than is produced through the intracranial route.

It would be naive to assume that the magnitude of reinforcement

produced by the systemic delivery of the drug results from interactions in a single brain area, especially when the intracranial

administration of the drug can, at best, result in a small stimulus

change in the organism. The actions of cocaine at multiple initiation

sites are probably responsible for the degree of reinforcement observed following intravenous presentation. A more thorough understanding of cocaine reinforcement processes can be obtained through

the identification of these loci and the integration of these data with

those obtained using intravenous self-administration procedures.

REFERENCES

Amit, Z.; Brown, Z.W.; and Sklar, L.S. lntraventricular selfadministration of morphine in naive laboratory rats.

Psychopharmacology 48:292-294, 1976.

Beckstead, R.M. An autoradiographic examination of corticocortical

and subcortical projections of the mediodorsal-projection (prefrontal) cortex in the rat. J Comp Neural 184:43-62, 1979.

Belluzi, J., and Stein, L. Enkephalin- and morphine-induced facilitation of long-term memory. Abstr Soc Neurosci 3:230, 1977.

212
Bozarth, M.A., and Wise, R.A. Electrolytic microinfusion transducer

system: An alternative method of intracranial drug application.

J Neurosci Methods 2:273-275, 1980.

Bozarth, M.A., and Wise, R.A. Intracranial self-administration of

morphine into the ventral tegmental area in rats. Life Sci 28:551-

555, 1981.

Brown, Z.W.; Amit, Z.; and Rockman, G.E. lntraventricular selfadministration of acetaldehyde, but not ethanol, in naive laboratory

rats. Psychopharmacology 64:271-276, 1979.

Co, C.; Smith, J.E.; and Lane, J.D. Use of a single compartment

LCEC cell in the determinations of biogenic amine content and

turnover. Pharmacol Biochem Behav 16:641-646, 1982.

Criswell, H.W. A simple chronic microinjection system for use with

chemitrodes. Pharmacol Biochem Behav 6:237-238, 1977.

Dworkin, S.I.; Goeders, N.E.; and Smith, J.E. The reinforcing and

rate effects of intracranial dopamine administration. In: Harris,

L.S., ed. Problems of Drug Dependence, 1985. National Institute

on Drug Abuse Research Monograph 67. DHHS Pub. No. (ADM) 86-

1448. Rockville, MD: the Institute, 1986. pp. 242-248.

Glimcher, P.W.; Giovino, A.A.; and Hoebel, B.G. Self-injection of

neurotensin into the ventral tegmental area (VTA). Abstr Soc

Neurosci 9:120, 1983.

Goeders, N.E.; Dworkin, S.I.; and Smith, J.E. Neuropharmacological

assessment of cocaine self-administration into the medial prefrontal

cortex. Pharmacol Biochem Behav 24:1429-1440, 1986.

Goeders, N.E.; Guerin, G.F.; Dworkin, S.I.; and Smith, J.E. Reinforcing stimulus properties of endocoids. In: Lal, H.; Labella, F.;

and Lane, J., eds. Endocoids. New York: Alan R. Liss, 1985. pp.

63-69.

Goeders, N.E.; Lane, J.D.; and Smith, J.E. Intracranial self-administration of methionine enkephalin into the nucleus accumbens.

Pharmacol Biochem Behav 20:451-455, 1984.

Goeders, N.E., and Smith, J.E. Cortical dopaminergic involvement in

cocaine reinforcement. Science 221:773-775, 1963.

Goeders, N.E., and Smith, J.E. Parameters of intracranial selfadministration of cocaine into the medial prefrontal cortex. In:

Harris, L.S., ed. Problems of Drug Dependence, 7984. National

Institute on Drug Abuse Research Monograph 55. DHHS Pub. No.

(ADM) 85-1393. Rockville, MD: the Institute, 1985. pp. 132-137.

Goeders, N.E., and Smith, J.E. Reinforcing properties of cocaine in

the medial prefrontal cortex: Primary action of presynaptic dopaminergic terminals. Pharmacol Biochem Behav 25: 191-199, 1986.

Goeders, N.E., and Smith, J.E. Intracranial self-administration

methodologies. Neurosci Biobehav Rev 11:319-329, 1987.

213
Goeders, N.E.; Van Osdell, C.; McAllister, K.H.; Dworkin, S.I.; and

Smith, J.E. Responding maintained by intracranial delivery of Iglutamate. Fed Proc 46:548, 1987.

Guerin, G.F.; Goeders, N.E.; Dworkin, S.I.; and Smith, J.E. Intracranial self-administration of dopamine into the nucleus accumbens.

Abstr Soc Neurosci 10: 1072, 1984.

Gustafson, L,K., and Pickens, P. lntraventricular amphetamine selfadministration in rats. Fed Proc 34:780, 1975.

Hoebel, B.G., and Aulisi, E. Cholecystokinin self-injection in the

nucleus accumbens and block with proglumide. Abstr Soc Neurosci

10:694, 1984.

Hoebel, B.G.; Monaco, A.P.; Hernandez, L.; Aulisi, E.F.; Stanley, B.G.;

and Lenard, L. Self-injection of amphetamine directly into the

brain. Psychopharmacology 81:158-163, 1983.

Lane, J.D.; Sands, M.P.; Co, C.; Cherek, D.R.; and Smith, J.E. Biogenie monoamine utilization in discrete rat brain regions is

correlated with conditioned emotional response and its conditioning

history. Brain Res 240:94-108, 1982.

Leonard, C.M. The prefrontal cortex of the rat: I. Cortical projections of the mediodorsal nucleus. II. Efferent connections. Brain

Res 12:321-343, 1969.

Monaco, A.P.; Hernandez, L.; and Hoebel, B.G. Nucleus accumbens:

Site of amphetamine self-injection: Comparison with the lateral

ventricle. In: Chronister, R.B., and DeFrance, J.F., eds. The

Neurobiology of the Nucleus Accumbens. Brunswick, ME: Haer

Institute for Electrophysiology, 1981. pp. 338-342.

Myers, R.D. Handbook of Drug and Chemical Stimulation of the

Brain. New York: Van Nostrand and Reinhold, 1974.

Myers, R.D., and Hoch, D.B. 14C-dopamine microinjected into the

brainstem of the rat: Dispersion kinetics, site content and functional dose. Brain Res Bull 3:601-609, 1978.

Olds, J. Hypothalamic substrates of reward. Physiol Rev 42:554-604,

1962.

Olds, J., and Olds, M.E. Positive reinforcement produced by stimulating hypothalamus with iproniazid and other compounds. Science

127:1175, 1958.

Olds, M.E. Hypothalamic substrate for the positive reinforcing prop

erties of morphine in the rat. Brain Res 168:351-60, 1979.

Olds., M.E. Reinforcing effects of morphine in the nucleus accumbens. Brain Res 237:429-440, 1982.

Olds, M.E., and Williams, K.N. Self-administration of D-Ala2-Metenkephalinamide at hypothalamic self-stimulation sites. Brain Res

194: 155-170, 1980.

214
Phillips, A.G., and Rolls, E.T. lntracerebral self-administration of

amphetamine by rhesus monkeys. Neurosci Lett 24:81-86, 1981.

Roberts, D.C.S.; Koob, G.F.; Klonoff, P.; and Fibiger, H.C. Extinction

and recovery of cocaine self-administration following 6-hydroxydopamine lesions of the nucleus accumbens. Pharmacol Biochem

Behav 12:781-787, 1980.

Routtenberg, A. Intracranial chemical injection and behavior: A

critical review. Behavioral Biology 7:601-642, 1972.

Smith, J.E.; Co, C.; Freeman, M.E.; and Lane, J.D. Brain neurotransmitter turnover correlated with morphine-seeking behavior of rats.

Pharmacol Biochem Behav 16:509-519, 1982.

Smith, J.E., and Lane, J.D. Brain neurotransmitter turnover correlated with morphine self-administration. In: Smith, J.E., and Lane

J.D., eds. The Neurobiology of Opiate Reward Processes.

Amsterdam: Elsevier, 1983. pp. 361-402.

Smith, J.E.; Leckrone, W.R.; and Co, C. Combination operant

conditioning-liquid nitrogen immersion chamber for studying

neurotransmitters and behavior. Pharmacol Biochem Behav 7:167-

172, 1977.

Stein, E.A., and Olds, J. Direct intracerebral self-administration of

opiates in the rat. Neurosci Abstr 3:302, 1977.

Tortella, F.C., and Moreton, J.F. D-Ala2-methionine-enkephalinamide

self-administration in the morphine-dependent rat.

Psychopharmacology 69:143-147, 1980.

Walaas, I. Biochemical evidence for overlapping neocortical and

allocortical glutamate projections to the nucleus accumbens and

rostral caudateoputamen in the rat brain. Neuroscience 6:399-405,

1981.

Weeks, J.R. Experimental morphine addiction: Method for automatic

intravenous injections in unrestrained rats. Science 138:143-144,

1962.

Weeks, J.R. Long-term intravenous infusion. In: Myers, R.D., ed.

Methods in Psychobiology. New York: Academic Press, 1972. pp.

155-168.

ACKNOWLEDGMENTS

This work was supported in part by U.S. Public Health Service grants

DA 04293, DA 03832, and DA 03628 from the National Institute on

Drug Abuse.

215
AUTHOR

Nick E. Goeders, Ph.D.

Assistant Professor

Departments of Pharmacology and Therapeutics

and Psychiatry

Louisiana State University Medical Center

P.O. Box 33932

Shreveport, IA 71130

216
Psychomotor Stimulant vs. Local

Anesthetic Effects of Cocaine:

Role of Behavioral Sensitization

and Kindling

Robert M. Post and Susan R.B. Weiss

lNTRODUCTlON

This paper focuses on behavioral sensitization and pharmacological

kindling, two distinct effects of cocaine, which appear to be related

to cocaine’s separate effects as a psychomotor stimulant and local

anesthetic. Both of these effects increase in intensity following

repeated, intermittent administration of a constant drug dose.

Behavioral sensitization, increased responsivity to the locomotorstimulating or stereotypy-inducing effects of cocaine, can be produced

by relatively low doses of cocaine. Pharmacological kindling,

repeated administration of subconvulsive doses of cocaine eventually

resulting in the development of full-blown seizures, occurs with

higher doses and is similar to that produced by other local anesthetics such as lidocaine. Thus, it would appear that the unique

properties of cocaine as both a psychomotor stimulant and a local

anesthetic carry with them the liability of inducing, upon repetition,

increased behavioral pathology and convulsive responsivity. These

two mechanisms, demonstrated in a variety of preclinical animal

laboratory studies, may have clinical relevance for the human cocaine

user who tends to become involved with repeated administration of

either low, high, or escalating doses. It is possible that the psychomotor stimulant properties of cocaine, which are associated with

behavioral sensitization, may play a role in the evolution and

development of cocaine-induced dysphorias and paranoid psychoses

like those reported for amphetamine, while the local anesthetic properties of cocaine may be more closely associated with the late

development of cocaine-induced panic attacks, seizures, and the

attendant potential lethality.

217
We will also emphasize an important role for environmental context

and conditioning in the development of cocaine-induced behavioral

sensitization. Initial studies of the possible neural substrates

mediating this effect will be presented. In some paradigms, cocaineinduced behavioral sensitization may be attributable to conditioning

variables, while, as doses are increased and repeated more

frequently, a context-independent behavioral sensitization can also

become evident; i.e., sensitization will occur independent of

conditioning variables. At the highest doses, pharmacological kindling

can ensue, an effect that may also be independent of conditioning

mechanisms. lf the present analysis is relevant to the human situation, it would suggest that there are inherent toxicities to repeated

cocaine use that can become increasingly manifest even if the abuser

of cocaine were never to escalate his cocaine dose.

COCAINE-INDUCED BEHAVlORAL SENSITIZATION

Elsewhere, we have reviewed some of the characteristics of cocaineinduced behavioral sensitization in detail (Post and Contel 1981).

Briefly, there appears to be a long-lasting change in behavioral

responsivity, which is thought to involve alterations in dopaminergic

function, perhaps in the mesolimbic system. Behavioral sensitization

is dose-dependent (Shuster et al. 1977), occurs at lower doses in

females than in males, and may be, in part, dependent upon intact

vasopressin function (Post and Contel 1981). Cocaine-induced

behavioral sensitization tends to show cross-sensitization to other

psychomotor stimulants, especially amphetamine, and also to show

cross-sensitization to a variety of environmental stressors (Antelman

et al. 1980; Kalivas et al. 1986; Kalivas et al. 1988; Duffy and

Kalivas, in press; DuMars et al. 1988). The interval between

injections appears important, with greater time between injections

associated with more robust sensitization up to a certain point (which

has not been adequately delineated).

A particular focus of this manuscript will be the role of environmental context and conditioning in cocaine-induced behavioral

sensitization. As previously demonstrated (Post et al. 1981), repeated

administration of cocaine in the same environment leads to marked

increases in locomotor hyperactivity and stereotypy, while pretreatment in one environment and testing in a dissimilar environment

leads to lesser degrees of sensitization. We have recently attempted

to reproduce the environmental context finding in a more expeditious

fashion. We have adopted a 1-day pretreatment paradigm in which

animals are injected with cocaine (40 mg/kg IP) or saline on day 1

218
and rechallenged with cocaine (10 mg/kg IP) on day 2 (Weiss et al.

1988). In this paradigm, as illustrated in figure 1 (left panel),

animals show a substantial cocaine-induced hyperactivity in response

to the 40 mg/kg injection, while saline-pretreated controls show only

minimal degrees of activity. A third group of animals was also used

in this study; they received saline in the test chamber and the same

dose of cocaine (40 mg/kg) in their home cage following the experimental session. In response to the challenge dose of cocaine (10

mg/kg) on day 2, only those animals that received cocaine in the

context of the test cage showed notable degrees of behavioral sensitization, and animals that received equal doses of cocaine in the

context of the home cage did not differ from saline-pretreated

controls (figure 1, right panel). Thus, it would appear with this

single high-dose paradigm that all of the cocaine-induced sensitization

is related to a conditioned process.

The effects are dose related. Pretreatment with cocaine (10 mg/kg

IP) on day 1 does not lead to behavioral sensitization to rechallenge

on day 2. However, if cocaine (10 mg/kg IP) is repeated for several

days, significant degrees of sensitization are evident by 4 to 7 days

(Post et al. 1981). In this paradigm, the behavioral sensitization also

appears to be environmental-context-dependent. With this repeateddose paradigm, significant increases in response to a saline challenge

are also evident in sensitized animals, while this is not apparent with

the single-dose sensitization method.

If high-dose cocaine pretreatment (40 mg/kg IP) is repeated for 3

days, cocaine-induced behavioral sensitization shifts from a predominance of locomotor hyperactivity as an endpoint to more

predominant stereotypy. With this shift, there is also a change from

sensitization involving exclusively a conditioning component to one in

which the stereotypic behavior demonstrates a sensitization in a

fashion that is independent of environmental context. These doserepetition relationships are summarized in table 1. We would suggest

the possibility that this shift might involve a change in neural sub

strates with the nucleus accumbens and amygdala being involved in

the environmental-contextdependent sensitization and striatal and

other extrapyramidal dopaminergic substrates, more typically associated with stereotypic behaviors, mediating the context-independent

cocaine-induced sensitization. Finally, as illustrated in table 1, with

high doses, cocaine’s local-anesthetic mechanisms are brought “on

line” and pharmacological kindling results, perhaps requiring additional

neuronal pathways (Post et al. 1984a).

219
FIGURE 1. Cocaine-induced behavioral sensitization depends on environmental context

NOTE: Left panel: Horizontal locomotar activity is illustrated on the ordinate and time on the abscissa for three groups of rats

(n=10/group) receiving the following day-i pretreatments: cocaine in the test cage and saline in the home cage filled

circles); saline in the test cage and cocaine in the home cage (Wed squares); saline in both cages (open circles). Right

panel: Horizontal activity is illustrated for these three groups following a cocaine challenge (10 mg/kg). Only the rats that
TABLE 1. Long-term effects of cocaine: Function of dose and

number of repetitions

NOTE: Size and number of arrows indicate dose of cocaine and number of administrations. COC = cocaine and dose in mg/kg administered once daily IP.

except in the study of Gale, when doses were subcutaneous. 0 = no effect;

± = equivocal; + = moderate effect: ++ = marked or definite effect. Effects of

cocaine increase and are more persistent with dose and number of repetitions.

They also become less dependent on environmental context and conditioning.

The highest doses may be associated wtth kindling of seizures and their

associated lethality. Thus, effects shift from behavioral sensitization to

kindling as the dose is increased.

In the single-day cocaine-induced behavioral sensitization paradigm,

we have attempted to further elucidate variables related to the

conditioning phenomenon. We have demonstrated that the degree of

221
similarity of the pretreatment and test environments is a critical

factor in determining the degree of behavioral sensitization (Weiss

et al. 1988). Animals pretreated and tested in the same environment

show the most robust degree of behavioral sensitization, although

animals pretreated in a Plexiglas cage that is very similar to the

activity-monitored cage also show significant degrees of behavioral

sensitization. When the initial place where animals display their

cocaine-induced hyperactivity is very different from that of the test

environment, such as one involving a wire cage in a different room,

there is no significant cocaine-induced behavioral sensitization.

Manifestation of cocaine-induced hyperactivity or stereotypy on day 1

is required for behavioral sensitization to become evident on rechallenge on day 2. That is, when we pretreated animals with neuroleptics or high doses of benzodiazepines in order to block day 1

cocaine-induced hyperactivity, no sensitization was evident upon

rechallenge on day 2 (Weiss et al. 1988). This effect was evident at

both moderate (0.2 mg/kg) and high (0.5 mg/kg) doses of haloperidol.

However, while these doses were sufficient to block the development

of sensitization when haloperidol was administered prior to the day-l

pretreatment, they were not sufficient to block the expression of

sensitization when they were administered only prior to the day-2

test dose (figure 2). That is, once cocaine-induced hyperactivity had

been manifest on day 1, moderate to high doses of neuroleptics were

insufficient to block the differential effects of this prior cocaine

experience on subsequent cocaine-induced hyperactivity. These data

replicate and extend those of Beninger and Herz (1986) Beninger and

Hahn (1983), and Tadokoro and Kuribara (1986).

Elsewhere (Post and Weiss 1988), we have discussed in detail the

interpretation of these results with respect to neuroleptic nonresponsiveness in some psychotic states. Basically, once the sensitized

behaviors have become established, neuroleptics may no longer be

capable of reversing these behaviors. To the extent that behavioral

sensitization accounts for some aspects of cocaine-induced syndromes,

one might postulate that these components would be less responsive

to neuroleptics than in instances where acute cocaine-induced

behavioral pathology is induced.

In contrast to neuroleptics, specific 6-hydroxdopamine lesions (with

desmethylimipramine pretreatment) lesions of the nucleus accumbens

and amygdala appear capable of blocking cocaine-induced sensitization

while leaving the degree of day-l cocaine-induced hyperactivity intact

222
FIGURE 2. Cocaine-induced behavioral sensitization: Halaperidol does not Mock its expression

NOTE: Haloperidol does not block the expression of sensitiztion to cocaine once it is induced. Group means and standard errors

for horizontal activit are plotted over time in animals challenged with saline prior to cocaine (10 mg/kg) (left panel) or

haloperidol (0.2 mg/kg) prior to cocaine (10 mg/kg) on day 2 (right panel). Animals that received cocaine (40 mg/kg

pretreatment on day 1 are indicated in the filled symbols: those given saline, open symbols. The left panel illustrates the

magnitude of cocaine-induced sensitization without haloperidol the right panel shows the persistent sesitization effeucts with

haloperidol
(Post et al. 1987b). While nucleus accumbens dopamine depletion of

60 to 65 percent was not sufficient to block cocaine-induced hyperactivity in response to the day-l injection with cocaine (40 mg/kg IP),

these animals were not different in their response on day 2 from

saline-pretreated controls.

Electrolytic lesions of the amygdala also blocked the development of

cocaine-induced behavioral sensitization, while lesions of the dorsal

hippocampus increased baseline hyperactivity but left the differential

effect of prior cocaine intact. We have subsequently replicated the

effect of amygdala lesions, demonstrating that both electrolytic

lesions and selective depletion of dopamine in the amygdala (achieved

by bhydroxydopamine and desmethylimipramine cotreatment) were

sufficient to block cocaine-induced behavioral sensitization (Post

et al. 1987b). Lesions of the ventral hippocampus or cerebellum were

not effective in blocking sensitization in this paradigm.

Thus, preliminary data would suggest that dopaminergic pathways

involving the nucleus accumbens and amygdala may be involved in the

mediation of the conditioned cocaine sensitization, possibly affecting

the coding of the experience of the prior cocaine-induced hyperactivity and/or its impact on subsequent behavior. Gold and associates (in press) have also suggested that the nucleus accumbens is

involved in the conditioned component of amphetamine-induced

behavioral sensitization. However, in their studies, more complete

lesions of the nucleus accumbens were achieved, which also blocked

the unconditioned amphetamine-induced behavior. Clearly, further

studies are needed in order to elucidate the precise anatomical

pathways and neurochemical substances involved in the conditioned

component of cocaine-induced behavioral sensitization.

PHARMACOLOGICAL KlNDLING

Goddard and associates (1969) first named the process “kindling,” in

which repeated electrical stimulation of the brain at subthreshold

currents eventually led to the development of major motor seizures.

A critical requirement for the development of kindling is intermittent

stimulation, since continuous stimulation or stimulation at intervals of

only several minutes never leads to the production of seizures. With

repeated, intermittent electrical stimulation, there is increasing

duration, spread, and complexity of after discharges, which culminate

in the appearance of major motor seizures. Once these have been

achieved, there seems to be a permanent change in excitability lasting

for 6 months to 1 year or longer. Moreover, if kindled seizures are

224
induced a sufficient number of times, a condition of spontaneity

occurs in which seizures occur in the absence of exogenous electrophysiological stimulation (Pinel 1981; Pinel 1983). Early evidence

suggested that a parallel process of “pharmacological kindling”

attributable to cocaine’s local anesthetic properties might occur with

repeated high-dose drug administration (Post et al. 1975; Post and

Kopanda 1976; Post 1977).

In order to more systematically assess this possibility, we administered the pure local anesthetic lidocaine and observed a kindlinglike

timecourse at doses of 60 to 65 mg/kg daily. In this paradigm,

approximately 40 percent of animals eventually developed seizures to

what was previously a subconvulsant dose of drug. Moreover, the

seizures resembled those achieved by amygdala kindling, with

involvement of head, trunk, and forepaws, with rearing and falling.

Electrophysiological spiking was also evident in the amygdala.

Deoxyglucose studies of regional metabolic activity revealed prominent

increases in glucose utilization in hippocampus and amygdala or

perirhinal structures achieved during lidocaine-induced seizures (Post

et al. 1984a). Following sufficient repetitions of lidocaine-induced

seizures, spontaneous seizures developed parallel to the phenomenon

of spontaneity in electrical kindling.

As reported by Eidelberg and associates (1963), cocaine also induces

prominent spiking in related limbic structures. When a dose of

cocaine (65 mg/kg) that is equipotent to lidocaine’s local anesthetic

effects is administered on a once-daily basis, more than 50 percent of

the animals show seizures on day 1. Moreover, all of the animals

administered this dose of cocaine experienced seizures and died by

day 3 of the study, even though a majority of animals tolerated repeated lidocains-induced seizures for many weeks or months. Thus, it

would appear that cocaine’s unique combination of local anesthetic

and psychomotor stimulant properties make the local anesthetic

seizures more lethal than those achieved by the pure local anesthetic

lidocaine.

We have observed several interesting effects of the anticonvulsant

carbamazepine on local anesthetic kindling. Chronic carbamazepine

treatment, using a diet containing 5.0 g carbamazepine/kg food,

almost completely blocks the development of lidocaine-kindled

seizures, but is without effect on completed kindled or high-dose

lidocaine seizures, although acute administration of the benzodiazepine

diazepam blocks kindling development and completed seizures.

Similarly, chronic carbamazepine is highly potent in inhibiting the

225
development of cocaine-kindled seizures (figure 3). In contrast to its

effects on high-dose lidocaine seizures, carbamazepine decreases the

incidence of highdose cocaine seizures and their associated lethality.

This effect is even more robust if a lower dose of cocaine is used to

more clearly demonstrate pharmacological kindling. A roughly similar

timecourse of seizure evolution can be achieved by cocaine

(40 mg/kg) compared with lidocaine (65 mg/kg), such that none of

the animals demonstrates seizures or dies on day 1, although some 80

percent of the animals experience seizures by 2 weeks of treatment.

In this instance, chronic carbamazepine remarkably decreases cocaineinduced seizure development (from 80 to 25 percent) and decreases

lethality from 50 to 5 percent by day 16.

Notably, however, acute carbamazepine in doses ranging from 15 to

50 mg/kg is without effect on either lidocaine- or cocaine-kindled or

high-dose seizures (figure 4). Furthermore, pretreatment with

repeated acute injections of carbamazepine (15 mg/kg IP) is also

without effect on the development of cocaine or lidocaine seizures

and may actually increase the lethality associated with the local

anesthetic seizures (figure 5). In contrast, acute IP administration of

the benzodiazepine diazepam will decrease both pharmacological

kindling development to the local anesthetic lidocaine as well as

completed lidocaine-kindled seizures. Thus, chronic but not acute

carbamazepine treatment appears to produce specific biochemical

effects that may be of use in the prevention of seizure development.

EFFECTS OF PURE LOCAL ANESTHETICS IN MAN

It is thus apparent that repeated administration of high doses of local

anesthetics are capable of inducing pharmacological kindling in

animals. Before further considering the possible implications of this

phenomenon in humans, should high enough doses of cocaine be selfadministered, it is important to consider what is known about the

subjective physiological effects of more moderate doses of pure local

anesthetics administered to man. Although in one study the effects

of nasally insufflated lidocaine were mistaken for those of cocaine in

experienced cocaine users (Van Dyke et al. 1979), lidocaine is generally adequately discriminated from cocaine (Fischman et al. 1983a),

and this local anesthetic is not self-administered by animals (Balster,

this volume). In contrast, procaine is self-administered by animals,

and its effects are more often confused with those of cocaine in

clinical experiments with sophisticated stimulant abusers (Fischman et

al. 1983b).

226
FIGURE 3. Chronic cerbemezepine prevents the development of cocaine-kindled seizures and mortality

NOTE: Rats were placed on a control or carbamazepine-containing diet for 4 days prior to the studies and were kept on these diets

throughout the period of cocaine injections. Cocaine was administered in doses of 40, 50, or 65 mg/kg. Carbamezapine was

effective in inhibiting the cocaine-induced seizures and lethality.
FlGURE 4. Lack of effect of acute carbamazepine on cocaine-induced

seizures and lethality

NOTE: Carbarnazepine was administered IP, 15 minutes before cocaine, in doses of 0,

15.25, or 50 mg/kg in an attempt to block acute high-dose (65 mg/kg) cocaine

seizures and lethality. No anticonvulsant effect was seen at any of these

doses. Cocaineinduced lethality either was not affected or was exacerbated

by these acute doses.

228
FIGURE 5 Pretreatment with chronic carbamazepine in diet (not

repeated injection) is required to block cocaine seizures

and lethality

NOTE: Rats were treated with chronic carbamazepine (diet), IP-injected carbamazepine

(15 mg/kg), or vehicle for 4 days prior to cocaine and IP carbamazepine (15

mg/kg) throughout the study. This regime of repeated IP carbamazepine

treatment did not decrease the response to high-dose cocaine, in contrast to

the chronic oral dosing.

229
We administered ascending doses of procaine to naive normal

volunteers and psychiatrically ill patients with diagnoses of primary

affective illness or borderline personality disorder (Kellner et al.

1987). Procaine has about one-quarter the local anesthetic potency

of cocaine or lidocaine, and was used because of its rapid metabolism

and safety in man. In doses ranging from .46 to 2.30 mg/kg IV, we

found dose-related effects of IV procaine on psychosensory and

cognitive distortions in a variety of sensory modalities. In contrast,

there were marked variations in the degree of affective response,

ranging from euphoria to profound dysphoria. Increases in fast EEG

frequencies in the beta range from 26 to 45 Hz were evident selectively over the temporal cortex. The degree of activation of these

fast frequencies was correlated with the degree of dysphorfc reaction

in the first 21 subjects studied. Procaine was also associated with an

endocrine profile of increases in plasma ACTH and cortisol as well as

prolactin, but not growth hormone. lt is of interest that preclinical

studies have indicated that cocaine, procaine, and lidocaine all induce

release of corticotropin releasing factor (CRF) in hypothalamic

preparations in vitro and that carbamazepine blocks this cocaineinduced release of CRF (Calogero et al. 1987).

CLINICAL IMPLICATlONS

Based on the procaine data, we would suggest that some of the acute

effects of cocaine normally associated with its psychomotor stimulant

properties could, in fact, be related to its local anesthetic effects.

These might include mood lability and, in some instances, euphoria,

profound anxiety and dysphoria, tinnitus, sensory distortions,

hallucinatory-like phenomena, and even seizures should the dose be

high enough. Based on the extrapolation from the pharmacological

kindling effects that have been demonstrated in preclinical studies

with repeated administration, we would suggest that some of the

chronic effects of cocaine could also be attributed to its local

anesthetic properties, including sensitization to bizarre behavior,

seizure sensitization (pharmacological kindling), interictal irritability

and aggression (particularly in subjects who have shown local

anesthetic-induced seizures), cognitive impairment, and even panic

attacks. It was strikingly unexpected to find that some 50 percent of

the first 500 patients calling the Cocaine Hotline to report adverse

psychological effects reported experiencing cocaine-induced panic

attacks (Washton and Gold 1984).

Based on the previous clinical observations of local anesthetic

dysphoria and preclinical data on local anesthetic kindling, we would

230
suggest that this extraordinarily high incidence of cocaine-induced

panic attacks might be related to cocaine’s local anesthetic rather

than its psychomotor stimulant properties. We are not aware of a

similar high incidence of amphetamine-induced panic attacks being

reported by any of the many investigators reporting epidemiological

data on the psychological consequences of amphetamine or by the

laboratories reporting psychological effects of amphetamine

administered under more controlled conditions. These background

data, along with our data (Kellner et al. 1987) show that many

patients experience the local anesthetic procaine, even in low doses,

as extremely dysphoric, and do so in proportion to the degree of EEG

activation over the temporal lobe, further support the contention that

the local anesthetic effects of cocaine might be related to these

dysphoric experiences. lf we further factor in the possible pharmacological kindlinglike effects occurring at a subconvulsant level, it is

possible that repeated cocaine-induced activation of temporal lobe

structures, as has been documented to occur in man (Ervin et al.

1969), may be sufficient to induce a kindlinglike increase in physiological and behavioral responsivity eventually leading to the

development of panic attacks.

The temporal characteristics of such a proposed course are evident in

a case observed by Uhde, Geraci, and colleagues in a clinic for

panic-anxious patients at the National Institute of Mental Health

(Post et al. 1987a). A 23-year-old single white male reported 3 years

of essentially continuous daily use of cocaine by the intranasal route

prior to his experience of his first cocaine-induced panic attack,

immediately following cocaine insufflation. Over the next 5 months,

he continued to experience cocaine-related panic attacks (approximately five times weekly); in each instance, these occurred in

immediate association with cocaine use. However, after these many

repetitions of cocaine-induced panic attacks, he developed a

spontaneous panic attack and assumed that his cocaine-related panic

attacks had led to this autonomous occurrence. He thus gave up his

daily cocaine use, but despite this, continued to experience intermittent and unpredictable spontaneous panic attacks. These became

associated with high levels of generalized anxiety and increasing

agoraphobia, leading to his eventually becoming housebound. These

spontaneous panic attacks improved with treatment with not only

such traditional agents as imipramine, but also with the anticonvulsant carbamazepine and, later, alprazolam.

Thus, it would appear that these panic attacks followed a kindlinglike timecourse in that they did not occur with the first of many

231
repetitions of cocaine administration, but then began to occur on a

regular basis with cocaine use, and finally, like the stage of spontaneity observed in both electrical and pharmacological kindling,

began to occur in the absence of local anesthetic administration. It

is also possible that various aspects of behavioral sensitization and

conditioning, demonstrated with subconvulsant doses in animals, are

relevant to the progressive development of such symptomatology.

Moreover, it is difficult to assess the exact dose of a drug that is

administered in uncontrolled street-use settings, as reported in this

case study, and whether a relatively constant dose was used over

time.

The likelihood of adverse consequences related to behavioral

sensitization and kindlinglike effects may be further enhanced because

of gradual escalations in actual cocaine doses over time. Since there

is ample evidence that tolerance or tachyphylaxis can develop to

cocaine’s euphoria-inducing properties (Fischman et al. 1976),

increasingly high doses might be employed by the user in an attempt

to recapture and/or maintain the euphoria or positive affective state.

This may, in fact, be a pharmacological impossibility, as suggested by

the recent studies of Sherer and colleagues (1986) indicating that,

even with maintenance of high steady-state cocaine levels with

continuous IV infusion in volunteers, increasingly dysphoric elements

and paranoia begin to emerge. In the face of dose escalation, one

might eventually achieve blood levels of cocaine high enough to

induce toxic local anesthetic effects. Then, with repeated

administration of even the same subconvulsant doses, the habitual

user might, in addition, be at increased risk for the induction of

seizures.

If the findings from the preclinical studies reviewed are relevant to

the clinical situation, it would appear that cocaine-induced behavioral

and psychological toxicities may, in fact, grow with repetition of the

same dose over time, just as they do in cocaine-induced behavioral

sensitization. Moreover, in some circumstances, powerful conditioned

phenomena appear to be engaged by cocaine, although at higher doses

and with greater numbers of repetitions, context-independent behavioral sensitization also occurs. Our conditioning findings in rodents

are of greater interest now that a variety of conditioned phenomena

have been reported in man. For example, greater degrees of paranoia

are experienced when cocaine is used in the place where it is

habitually administered rather than when it is used in novel environments (D.L., personal communication, 1986). Gawin and associates

(1986) reported numerous vignettes of conditioned cues inducing

232
relapses, cocaine craving, or withdrawal syndromes. Similarly,

Childress et al. (1987) repotted that pictures of cocaine-related

paraphernalia were powerful conditioned cues in producing cocaine

craving in abstinent former cocaine users, similar to that described

with opiates. Craving could be systematically extinguished using

principles of learning theory. These findings are also of interest in

relationship to the report of Hinson and Poulos (1981) that cocaineinduced behavioral sensitization in rats could, likewise, be

deconditioned with repeated injections of saline.

It is also noteworthy that Stein and Belluzzi (1986) have reported

conditioning with cocaine at the level of the single cell. The cell

was “rewarded” with cocaine whenever it increased its firing rate.

Compared with pseudoconditioned controls, which were administered

cocaine at random intervals, the cocaine-conditioned cells “learned” to

increase their firing rates to a remarkable and significant degree.

Thus, cocaine-induced conditioned phenomena appear to be robust at

the level of the single cell (Stein and Belluzzi 1986), the rodent (Post

et al. 1981; Hinson and Poulos 1981), the cat (Ellinwood and Kilbey

1980), and, finally, man (Gawin et al. 1988; Childress et al. 1987).

Perhaps specific therapeutic strategies aimed at the powerful nature

of cocaine-induced behavioral sensitization and its associated

conditioned cues may be helpful in the long-term treatment of

cocaine users. Preclinical studies also suggest the need for

considerable caution in relationship to potential clinical utility of

pharmacological interventions. For example, neuroleptics may or may

not be effective depending on the phase of development of cocaineinduced symptoms that the user is experiencing. Not only might the

neuroleptic nonresponsiveness of cocaine-induced behavioral sensitization provide a model for neuroleptic refractoriness in some

psychotic states clinically, but it should also lead to a search for

pharmacological interventions that may be effective on this phase.

The local anesthetic effects of cocaine and their associated liability

for pharmacological kindling appear to add a danger beyond that

experienced by users of the pure psychomotor stimulants such as

amphetamine, methamphetamine, methylphenidate, and related compounds. Moreover, pharmacological interventions, such as acute

carbamazepine, do not reverse cocaine-induced seizures (figures 4

and 5), limiting any potential clinical utility of an acute approach

strategy. Carbamazepine also does not block the euphoria of stimulants such as methylphenidate (Meyendorff et al. 1985). However,

based on our preclinical analysis of local anesthetic-related

233
kindlinglike effects, we would predict that chronic carbamazepine

might be capable of blocking the development of cocaine-induced

panic attacks and seizures clinically. Since carbamazepine does not

block cocaine-induced hyperactivity acutely or the development of

behavioral sensitization (Post et al., in press), evidence of the

efficacy of this compound in the development of cocaine-induced

panic would clearly suggest that cocaine-induced panic attacks were

more closely related to a kindlinglike rather than a sensitizationlike

mechanism.

Thus, it is likely that repeated cocaine administration may carry the

danger of increased behavioral and physiological toxicities, even with

use of the same dose over time. Behavioral sensitization (figure 1)

and kindling paradigms (figure 3) that focus on possible mechanisms

for this effect may help direct us to the neuroanatomical substrates

involved (e.g., the nucleus accumbens and amygdala), and eventually

lead to the possibility of new psychological and pharmacological

treatment interventions. Further study of the evolution of cocainerelated syndromes not only may be of value in developing new treatments for cocaine-related behavioral and physiological toxicities in

man, but also may provide important clinical and preclinical models

for a variety of endogenous psychiatric syndromes and their evolution

in man, including affective disorders, panic attacks, and paranoid

schizophreniform states. It is also increasingly important for the

potential and often naive user of cocaine to be made aware of the

large variety of extremely adverse psychological and physiological

consequences of acute and, especially, chronic use of this drug,

effects that may not be at all evident early in the course of cocaine

use. Since cocaine has such powerful reinforcing effects, which are

extraordinarily difficult for many users to resist, it is critical to

educate the public regarding potential extreme adverse consequences

of repeated cocaine use.

REFERENCES

Antelman, S.M.; Eichler, A.J.; Black, C.A.; and Kocan, D. Interchangeability of stress and amphetamine in sensitization. Science

207:329-331, 1980.

Beninger, R.J., and Hahn, B.L. Pimozide blocks establishment but not

expression of amphetamine-produced environment-specific

conditioning. Science 220:1304-1306, 1983.

Beninger, R.J., and Herz, R.S. Pimozide blocks establishment but not

expression of cocaine-produced environment-specific conditioning.

Life Sci 38:1425-1431, 1986.

234
Calogero, A.E.; Kling, M.A.; Gallucci, W.T.; Saoutis, C.; Post, R.;

Chrousos, G.P.; and Gold, P.W. Local anaesthetics procaine and

lidocaine stimulate corticotropin releasing hormone secretion in

vitro: Clinical implications. Abstr Soc Neurosci 319.2:1, 163, 1987.

Childress, A.R.; McLellan, A.T.; Ehrman, R.N.; and O’Brien, C.P.

Extinction of conditioned responses in abstinent cocaine or opioid

users. In: Harris, L.S., ed. Problems of Drug Dependence, 1986.

Proceedings of the 48th Annual Scientific Meeting, the Committee

on Problems of Drug Dependence, Inc. National Institute on Drug

Abuse Research Monograph 76. DHHS Pub. No. (ADM)87-1508.

Washington, DC: Supt. of Docs., U.S. Govt. Print. Off., 1987. pp.

189-195.

Duffy, P., and Kalivas, P.W. Decreased release of endogenous

dopamine in the A10 region in rats sensitized to cocaine and

morphine. Ann NY Acad Sci, in press.

DuMars, L.A.; Rodger, L.D.; and Kalivas, P.W. Behavioral crosssensitization between cocaine and enkephalin in the A10 dopamine

region. Behav Brain Res 27:87-91, 1988.

Eidelberg, E.; Lesse, H.; and Gault, R.P. An experimental model of

temporal lobe epilepsy: Studies of the convulsant properties of

cocaine. In: Glaser, G.H., ed. EEG and Behavior. New York:

Basic Books, 1963. pp. 272-283.

Ellinwood, E.H., and Kilbey, M.M. Fundamental mechanisms

underlying altered behavior following chronic administration of

psychomotor stimulants. Biol Psychiatr 15:749-757, 1980.

Ervin, F.R.; Mark, V.; and Stevens, J. Behavioral and affective

responses to brain stimulation in man. Proc Annu Meet Am

Psychopathol Assoc 58:54-65, 1989.

Fischman, M.W.; Schuster, C.R.; and Hatano, Y. A comparison of the

subjective and cardiovascular effects of cocaine and lidocaine in

humans. Pharmacol Biochem Behav 18:123-127, 1983a.

Fischman, M.W.; Schuster, C.R.; and Rajfer, S. A comparison of the

subjective and cardiovascular effects of cocaine and procaine in

humans. Pharmacol Biochem Behav 18:711-716, 1983b.

Fischman, M.W.; Schuster, C.R.; Resnekov, L; Schick, J.F.E.;

Krasnegor, N.A.; Fennell, W.; and Freedman, D.X. Cardiovascular

and subjective effects of intravenous cocaine administration in

humans. Arch Gen Psychiatry 33:983-989, 1976.

Gawin, F.H.; Byck, F.; and Kleber, D. Desipramine augmentation of

cocaine abstinence: Initial results. Clin Neuropharmacol 9(4):202-

204, 1986.

235
Goddard, G.V.; McIntyre, D.C.; and Leech, C.K. A permanent change

in brain function resulting from daily electrical stimulation. Exp

Neurol 25:295-330, 1969.

Gold, L.H.; Swerdlow, N.R.; and Koob, G.F. 1986. The role of

mesolimbic dopamine in conditioned locomotion produced by

amphetamine. Behav Neurosci, in press.

Hinson, R.E., and Poulos, C.X. Sensitization to the behavioral effects

of cocaine: Modification by Pavlovian conditioning. Pharmacol

Biochem Behav 15:559-562, 1981.

Kalivas, P.W.; Monguio, I.; and Duffy, P. Neurochemical sensitization

to daily stress and cross-sensitization with cocaine. Brain Res,

submitted for publication, 1988.

Kalivas, P.W.; Richardson-Carlson, R.; and Van Orden, G. Crosssensitization between foot shock stress and enkephalininduced

motor activity. Bid Psychiatry 21:939-950, 1986.

Kellner, C.H.; Post, R.M.; Putnam, F.; Cowdry, R.; Gardner, D.; Kling,

M.A.; Minichiello, M.D.; Trettau, J.R.; and Coppola, R. Intravenous

procaine as a probe of limbic system activity in psychiatric patients

and normal controls. Biol Psychiatry 22:1107-1126, 1987.

Meyendorff, E.; Lerer, B.; Moore, N.C.; Bow, J.; and Gershon, S.

Methylphenidate infusion in euthymic bipolars: Effect of

carbamazepine pretreatment. Psychiatry Res 16:303-308, 1985.

Pinel, J.P. Effects of diazepam and diphenylhydantoin on elicited and

spontaneous seizures in kindled rats: A double dissociation.

Pharmacol Biochem Behav 18:61-63, 1983.

Pinel, J.P.H. Kindling-induced experimental epilepsy in rats: Cortical

stimulation. Exp Neural 72:559-569, 1981.

Post, R.M. Progressive changes in behavior and seizures following

chronic cocaine administration: Relationship to kindling and

psychosis. In: Ellinwood, E.H., and Kilbey, M.M., eds. Advances

in Behavioral Biology. Vol. 21. Cocaine and Other Stimulants.

New York: Plenum Press, 1977. pp. 353-372.

Post, R.M., and Contel, N.R. Cocaine-induced behavioral sensitization:

A model for recurrent manic illness. In: Perris, C.; Struwe, G.;

and Jansson, B., eds. Biological Psychiatry. Amsterdam: Elsevier,

1981. pp. 746-749.

Post, R.M.; Kennedy, C.; Shinohara, M.; Squillace, K.; Miyaoka, M.;

Suda, S.; Ingvar, D.H.; and Sokoloff, L. Metabolic and behavioral

consequences of lidocaine-kindled seizures. Brain Res 324:295-303,

1984a.

Post, R.M., and Kopanda, R.T. Cocaine, kindling, and psychosis. Am

J Psychiatry 133:627-634, 1976.

236
Post, R.M.; Kopanda, R.T.; and Lee, A. Progressive behavioral

changes during chronic lidocaine administration: Relationship to

kindling. Life Sci 17:943-950, 1975.

Post, R.M.; Lockfeld, A.; Squillace, K.M.; and Contel, N.R. Drugenvironment interaction: Context dependency of cocaine-induced

behavioral sensitization. Life Sci 28:755-760, 1981.

Post, R.M.; Rubinow, D.R.; and Ballenger, J.C. Conditioning,

sensitization, and kindling: Implications for the course of affective

illness. In: Post, R.M., and Ballenger, J.C., eds. Neurobiology of

Mood Disorders. Baltimore: Williams & Wilkins, 1984b. pp. 432-

466.

Post, R.M., and Weiss, S.R.B. Commentary to the paper by R.J.

Beninger. In: Simon, P.; Soubrie, P.; and Widlocher, D., eds.

Animal Models of Psychiatric Disorders. Vol. 1. New York:

Karger Press, 1988. pp. 52-60.

Post, R.M.; Weiss, S.R.B.; and Pert, A. Cocaine-induced behavioral

sensitization and kindling: Implications for the emergence of

psychopathology and seizures. Ann NY Acad Sci, in press.

Post, R.M.; Weiss, S.R.B.; Pert, A.; and Uhde, T.W. Chronic cocaine

administration: Sensitization and kindling effects. In: Fisher, S.;

Raskin, A.; and Uhlenhuth, E.H., eds. Cocaine: Clinical and

Biobehavioral Aspects. New York: Oxford University Press, 1987a.

pp. 109-173.

Post, R.M.; Weiss, S.R.B.; Smith, J., Jr.; Sullivan, T.L.; and Pert, A.

Amygdala and nucleus accumbens lesions impair cocaine

sensitization. Abstr Soc Neurosci 185.7:661, 1987b.

Sherer, M.; Kumar, K; Golden, R.; and Jaffe, J. Continuous infusion

of cocaine-a model for cocaine psychosis? Society for Biological

Psychiatry, Washington, DC, May 1986, Abst. #160, p. 212.

Shuster, L; Yu, G.; and Bates, A. Sensitization to cocaine

stimulation in mice. Psychopharmacology (Berlin) 52:185-190, 1977.

Stein, L., and Belluzzi, J. Second messengers, natural rewards, and

drugs of abuse. Clin Neuropharmacol 9:S205-S207, 1986.

Tadokoro, S., and Kuribara, H. Reverse tolerance to the ambulationincreasing effect of methamphetamine in mice as an animal model

of amphetamine psychosis. Psychopharmacol Bull 22:757-762, 1986.

Van Dyke, C.; Jatlow, P.; Ungerer, J.; Barash, P.; and Byck, R.

Cocaine and lidocaine have similar psychological effects after

intranasal application. Life Sci 24:271-274, 1979.

Washton, A.M., and Gold, M.S. Chronic cocaine abuse: Evidence for

adverse effects on health and functioning. Psychiatric Annals

14:733-743, 1984.

237
Weiss, S.R.B.; Post, R.M.; Pert, A.; Woodward, R.; and Murman, D.

Role of conditioning in cocaine-induced behavioral sensitization:

Differential effects of haloperidol. Neuropsychopharmacology,

submitted for publication, 1988.

AUTHORS

Robert M. Post, M.D.

Susan R.B. Weiss, Ph.D.

Biological Psychiatry Branch

National Institute of Mental Health

Bethesda, MD 20892

238
Genetic Differences in Responses

to Cocaine

Frank R. George and Steven R. Goldberg

INTRODUCTION

Cocaine is an effective reinforcer and at present a widely used drug

of abuse. It produces marked physiological effects (e.g., changes in

heart rate and blood pressure) as well as behavioral effects, including

euphoria and increased locomotor activity. There is, however,

significant individual variability in cocaine uptake and metabolism, as

well as in behavioral responses to cocaine. This suggests that

genetic factors may play an important role in determining the extent

of responses to cocaine. Although the pharmacology and biochemistry

of this drug have been widely studied, little is known about the

contribution of genotype in determining response to this increasingly

abused substance.

Studies with other abused drugs such as ethanol and, to a lesser

extent, opiates have repeatedly demonstrated that genetic factors play

an important role in regulating the effects of these drugs (Broadhurst

1979; Crabbe and Belknap 1980; McClearn and Rogers 1961). One

widely used measure of drug effects in animal models is locomotor

activity. Genetic differences in locomotor response to ethanol and

opiates, as well as to xanthines and barbiturates, have been shown

(Ritz et al. 1981; Shuster et al. 1975; Oliverio and Castellano 1974;

Logan et al. 1986; Suzuki et al. 1987).

In addition to locomotor and other acute responses to drug challenge,

genetic factors could also play a role in regulating drug-seeking

behavior. The use of inbred strains and selectively bred lines in

studies on ethanol (McClearn and Rodgers 1961; Rodgers and

McClearn 1962; Eriksson 1968; Li et al. 1979; Eriksson and Rusi 1981;

Tampier et al. 1981; Elmer et al. 1986; Ritz et al. 1986; George 1987),

239
has provided valuable information about the contribution of genetic

factors to ethanol drinking.

There are very few reports of studies on genetic differences in

intake of nonethanol drugs. The existing data, all of which involve

studies on home cage drinking of opioid solutions, do suggest that

large population differences may exist with regard to drug-seeking

behavior (Nichols and Hsiao 1967; Carroll et al. 1986; George and

Meisch 1984). A different approach to the analysis of drug-seeking

behavior involves self-administration studies using operant techniques.

Under certain conditions, drugs from several pharmacological classes

can act as positive reinforcers (Deneau et al. 1969; Weeks 1962;

Schuster and Thompson 1969; Goldberg 1976; Spealman and Goldberg

1978; Griffiths et al. 1979; Meisch 1984; Young and Herling 1986).

Recently, several reports have shown significant genetic influences on

ethanol self-administration (Ritz et al. 1986; Elmer et al. 1986; Elmer

et al. 1987; George 1987; George, in press). The results obtained in

these studies and in other recent work by Li and coworkers (Li and

Lumeng 1984; Waller et al. 1984) indicate that genetic analyses are

important in studies related to the understanding of drug-reinforced

behavior. To the best of our knowledge, however, there are no

reports on genetic factors in nonethanol operant drug self-administration.

Self-administration procedures utilizing the operant paradigm combine

several key criteria necessary for an effective animal model of

substance abuse. However, the major focus in self-administration

studies has been to determine the environmental conditions important

in the initiation and persistence of drug-seeking behaviors. These

experiments generally have used a limited number of genetically

undefined subjects, and experimental conditions are often varied

independently across subjects. Individual differences found in these

studies have generally been attributed to differences in training and

subject history, or often ignored. It is possible that genetic differences in sensitivity, metabolism, or the reinforcing effects of drugs

may have given rise to this variability.

The integration of behavioral genetic and operant methodologies has

great potential for increasing our understanding of the contributions

and interactions of genetic and environmental factors in determining

behavior. The objectives of this integrative approach to pharmacological research would be to identify and study at several levels of

scientific analysis, including molecular, cellular, and behavioral, those

factors that mediate responses to drugs. The methodology and

240
principles of operant conditioning and pharmacogenetic analysis can

be effectively combined to ascertain the relationship between one

operant behavior, such as drug self-administration under fixed-ratio

schedules and other operant behaviors, such as food-reinforced

behavior, as well as nonoperant behaviors, such as locomotor activity.

These comparisons can be used to determine the degree of common

genetic control among these factors as well as to establish the extent

to which one factor, such as neurosensitivity, covaries with another

factor, such as self-administration. These and other issues, such as

commonality of self-administration behavior across drugs, can be most

effectively addressed by using genetically defined animals within selfadministration paradigms.

The purpose of this paper is to (1) summarize findings to date on

genetic factors in drug self-administration; (2) present preliminary

data from our laboratory showing genetic differences in locomotor

stimulant and lethal effects of cocaine: and (3) present preliminary

data on genetic differences in cocaine self-administration obtained

from a home cage drinking procedure derived from operant selfadministration experimental designs.

MATERIALS AND METHODS

Animals

Adult (10 to 14 weeks old at start of testing) LEW (LEW/CRLBR) and

F344 (CDF(F-344)CRLBR) male rats were used. All animals were

experimentally naive, housed individually (self-administration studies)

or in groups of same-sex littermates (locomotor studies) in a

temperature-controlled room 26 °C) with a 12-hour light/dark cycle

(0700 to 1900 lights on), and given free access to Purina laboratory

chow and tapwater prior to initiation of the experiments.

Locomotor Activity Studies

Cocaine, when administered to rats, is known to produce dose-related

increases in spontaneous locomotor behavior followed by death at

very high doses. In order to examine the role of genetic factors in

response to cocaine, we studied the dose-dependent effects of acute

cocaine administration in LEW and F344 inbred rats.

Procedure. Six each LEW and F344 rats were individually placed in

Digiscan Animal Activity monitors for an acclimation period of 20

minutes, then injected IP with one of several doses of cocaine HCI in

241
0.9 percent sterile saline expressed as mg/kg freebase. Doses

were administered in a volume of 1.0 ml/kg. Rats were immediately

returned to the activity monitor, and measures of activity were

accumulated electronically and summed every 10 minutes for 60

minutes.

Home Cage Drinking Studies

Apparatus The animal’s plastic home cage served as the experimental chamber. Each cage was equipped with a stainless steel grid

top. Standard 100-ml drinking bottles with sipper tubes were used.

Two additional cages with bottles but no subjects were used to

control for evaporation and spillage.

Procedure. Three each F344 and LEW rats weighing approximately

230 g and 300 g, respectively, at the start of their training were

used. Sessions were run 5 days a week between 900 and 1200 hours.

The cocaine concentrations (expressed in mg/ml) were prepared using

cocaine HCI in tapwater. The stock solutions were prepared, sealed,

and stored in a refrigerator. Solutions were periodically checked to

control for possible degradation of cocaine. The volumes consumed

were measured at the end of each session by weighing the bottles.

The animals were weighed daily.

Establishment of water drinking during the testing period. Water

bottles were removed from the rats’ home cages for 22 hours/day

and, to increase further the probability of drinking, the daily

feedings of Purina laboratory chow were placed in the cage 60

minutes prior to the beginning of the session.

Induction of cocaine drinking. Presession feedings continued for a

series of daily 1-hour sessions. During the first five sessions,

0 mg/ml cocaine (water) was the available liquid, then .05, .1, .2, .4,

and .57 mg/ml cocaine for five sessions each. The presession

feedings were then gradually reduced, and eventually all food was

given to the rats following each session.

Cocaine drinking after the termination of food-induced drinking.

After each 1-hour session the rats now had free access to water and

their daily food allotment in their home cages. As an initial test of

whether cocaine was functioning as a reinforcer, the rats had access

to .57 mg/ml cocaine for 10 sessions, followed by 0 mg/ml cocaine

(i.e., water) for 10 sessions.

242
RESULTS

Locomotor Activity Studies

The results are summarized in figure 1. A potency difference was

found, with LEW rats being more sensitive and F344 rats less

sensitive to the locomotor stimulant effects of cocaine. Conversely,

while a difference in lethality response to cocaine was also found,

the direction was opposite. All F344 rats but no LEW rats died at a

cocaine dose of 60 mg/kg.

FIGURE 1. Locomotor activity as a function of cocaine dose in

LEW and F344 rats

Home Cage Drinking Studies

Figure 2 shows that both LEW and F344 rats ingested substantial

amounts of cocaine when food induced. Intakes averaged 17.3 and

11.5 mg/kg cocaine and 29.1 and 19.8μl/g liquid per 1-hour exposure

for LEW and F344 rats, respectively. In the absence of food

inducement, liquid intake of cocaine solutions was higher in the LEW

rats relative to the F344 rats (14.1 vs. 4.8 μI/g and 8.0 vs. 2.7

mg/kg respectively) and exceeded intake of vehicle (8.8 μI/g) in the

243
LEW animals). Conversely, intake of cocaine solutions was not higher

than intake of vehicle (5.4 μI/g) in the F344 rats. This suggests

that orally delivered cocaine was serving as a reinforcer only for

LEW rats. While not quantified, observations of the LEW rats showed

that most intake of cocaine solutions occurred during the initial

portion of the test session.

FIGURE 2. Self-administration of .57 mg/ml cocaine in LEW

end F344 male rats

244
DISCUSSION

The results of these preliminary studies suggest that significant

genetic differences may exist in response to cocaine. In addition, the

stimulant, reinforcing, and lethal effects of cocaine may not be highly

correlated. Striatal dopaminergic pathways appear to be important in

mediating the locomotor-stimulant effects of cocaine (Heikkila et al.

1979), and these pathways have also been shown to be critical for the

initiation of reinforcement from cocaine (Ritz et al. 1987). This

suggests the possibility that innate differences in striatal dopaminergic pathways may exist between the genotypes used in this study.

Although not quantitatively or pathologically determined, death from

cocaine in this study was apparently due to effects on cardiovascular

tissue. Since sensitivity to the lethal and locomotor stimulant effects

does not appear to be correlated, this suggests that the lethal effects

of cocaine may not be the result of actions on dopaminergic

pathways. Ritz and coworkers (1987) have recently implicated heart

muscarinic sites as potent sites for cocaine binding. It may be that

the LEW and F344 rats differ with regard to these sites, thus

contributing to the difference seen in lethal response to cocaine

between these strains.

These are important questions with regard to substance abuse that

can be ideally addressed using behavioral genetic methods, particularly

genetic correlations. Genetically defined animals can be readily

characterized for a given behavioral phenotype and can then be used

to test mechanistic hypotheses by studying the correlations between

the behavioral phenotype and other measures hypothesized to be

related in some manner. A lack of correlation allows for the

conclusion that the measures studied are not mechanistically related.

A strong positive correlation, especially a perfect rank order

correlation between measures, provides strong supportive evidence

that the measures are causally related. While pharmacological

correlations can be effective in identifying a specific site through

which a drug initiates its effects, this site may be necessary but not

sufficient for a drug action, and establishing the extent of genetic

correlation is a necessary step in completely identifying the biochemical system(s) that mediate a particular drug response.

Behavioral genetics is not only the study of genetic influences upon

behavior, but is also, and importantly, the study of environmental

influences upon behavior. This latter facet results from the fact that

not only can genotype be manipulated as an independent variable, it

can also be controlled. When genotype is completely controlled or

245
held constant, then the resulting variability among subjects is due to

environmental factors. When specific environmental variables are

being systematically varied, as in most operant studies, the inclusion

of complete genetic control through the use of highly inbred subjects

can provide a very powerful and repeatable test for the significance

of specific environmental factors on behavior.

The results of the present studies are important for several reasons:

(1) they illustrate the importance of control for genetic variability in

pharmacological research; (2) they provide initial data that can be

used in correlational studies dealing with mechanistic hypotheses

regarding cocaine’s sites of action; and (3) they suggest the existence

of populations “at risk” for substance abuse.

REFERENCES

Brase, D.A.; Loh, H.H.; and Way, E.L. Comparison of the effects of

morphine on locomotor activity and analgesia in six mouse strains.

J Pharmacol Exp Ther 201:368-374, 1977.

Broadhurst, P.L. Drugs and the Inheritance of Behavior: A Survey

of Comparative Pharmacogenetics. New York and London: Plenum

Press, 1979. 206 pp.

Carroll, M.E.; Peterson, M.C.; and Harrison, R.G. Food deprivation

reveals strain differences in opiate intake of Sprague-Dawley and

Wistar rats. Pharmacol Biochem Behav 24:1095-1099, 1986.

Crabbe, J.C., and Belknap, J.K. Pharmacogenetic tools in the study of

drug tolerance and dependence. Subst Alcohol Actions Misuse

1:385-413, 1980.

Deneau, G.; Yanagita, T.; and Seevers, M.H. Self-administration of

psychoactive substances by the monkey. Psychopharmacologia

16:30-48, 1969.

Elmer, G.I.; Meisch, R.A.; and George, F.R. Oral ethanol reinforced

behavior in inbred mice. Pharmacol Biochem Behav 24:1417-1421,

1986.

Elmer, G.I.; Meisch, R.A.; and George, F.R. Differential concentration-response curves for oral ethanol self-administration in

C57BL/6J and BALB/cJ mice. Alcohol 4:63-68, 1987.

Eriksson, K. Genetic selection for voluntary alcohol consumption in

the albino rat. Science 159:739-741, 1968.

246
Eriksson, K., and Rusi, M. Finnish selection studies on alcoholrelated behaviors: General outline. In: McCleam, G.E.; Deitrich,

R.A.; and Erwin, V.G., eds. Development of Animal Models as

Pharmacogenetic Tools. National Institute on Alcohol Abuse and

Alcoholism Research Monograph 6. DHHS Pub. No. (ADM)81-1133.

Rockville, MD: U.S. Department of Health, Education, and Welfare,

1981. pp. 87-117.

George, F.R. Genetic and environmental factors in ethanol selfadministration. Pharmacol Biochem Behav 27:379-364, 1987.

George, F.R. The use of genetic tools in the study of substance

abuse. Alcoholism: Clin Exp Res, in press.

George, F.R., and Meisch, R.A. Oral narcotic intake as a reinforcer:

Genotype x environment interactions. Behav Genet 14:603, 1984.

Goldberg, S.R. The behavioral analysis of drug addiction. In: Glick,

S.D., and Goldfarb, J., eds. Behavioral Pharmacology.

St. Louis: C.V. Mosby, 1976. pp. 283-316.

Griffiths, R.R.; Brady, J.V.; and Bradford, L.D. Predicting the abuse

liability of drugs with animal drug self-administration procedures:

Psychomotor stimulants and hallucinogens. In: Thompson, T., and

Dews, P.B., eds. Advances in Behavioral Pharmacology. Vol. 2.

New York: Academic Press, 1979. pp. 163-208.

Heikkila, R.E.; Cabbat, F.S.; Manzino, L.; and Duvoison, R.C.

Rotational behavior induced by cocaine analogs in rats with

unilateral 6-hydroxydopamine lesions of the substantia nigra:

Dependence upon dopamine uptake inhibition. J Pharmacol Exp

Ther 211:189-194, 1979.

Li, T.-K., and Lumeng, L. Alcohol preference and voluntary alcohol

intakes of inbred rat strains and the National Institutes of Health

heterogeneous stock of rats. Alcoholism: Clin Exp Res 8:485-486,

1984.

Li, T.-K.; Lumeng, L.; McBride, W.J.; and Waller, M.B. Progress

toward a voluntary oral consumption model of alcoholism. Drug

Alcohol Depend 4:45-60, 1979.

Logan, L.; Seale, T.W.; and Carney, J.M. Inherent differences in

sensitivity to methylxanthines among inbred mice. Pharmacol

Biochem Behav 24:1281-1286, 1986.

McClearn, G.E., and Rodgers, D.A. Genetic factors in alcohol

preference of laboratory mice. J Comp Physiol Psychol 54:116-119,

1961.

Meisch, R.A. Alcohol self-administration by experimental animals.

In: Smart, R.G.; Cappell, H.D.; Glaser, F.B.; Israel, Y.; Kalant, H.;

Popham, R.E.; Schmidt, W.; and Sellers, E.M., eds. Research

Advances in Alcohol and Drug Problems. Vol. 8. New York:

Plenum Press, 1984. pp. 23-45.

247
Nichols, J.R., and Hsiao, S. Addiction liability of albino rats:

Breeding for quantitative differences in morphine drinking. Science

157:561-563, 1967.

Oliverio, A., and Castellano, C. Genotype dependent sensitivity to

morphine and heroin: Dissociation between opiate-induced running

and analgesia in the mouse. Psychopharmacologia 39:13-22, 1974.

Ritz, M.C.; George, F.R.; and Collins, A.C. lndomethacin antagonizes

ethanol-induced but not pentobarbital-induced behavioral activation.

Subst Alcohol Actions Misuse 2:289-299, 1981.

Ritz, M.C.; George, F.R.; deFiebre, C.M.; and Meisch, R.A. Genetic

differences in the establishment of ethanol as a reinforcer.

Pharmacol Biochem Behav 24:1089-1094, 1986.

Ritz, MC.; Lamb, R.J.; Goldberg, S.R.; and Kuhar, M.J. Cocaine

receptors on dopamine transporters are related to self-administration of cocaine. Science 237:1219-1223, 1987.

Ritz, M.C.; Sharkey, J.; and Kuhar, M.J. Cocaine inhibition of 3HQNB binding to muscarinic receptors in rat brain and heart. Fed

Proc 13(1):144, 1987.

Rodgers, D.A., and McClearn, G.E. Mouse strain differences in

preference for various concentrations of alcohol. Q J Stud Alcohol

23:26-33, 1962.

Schuster, C.R., and Thompson, T. Self-administration of and

behavioral dependence on drugs. Annu Rev Pharmacol 9:483-502,

1969.

Shuster, L; Webster, G.W.; Yu, G.; and Eleftheriou, B.E. Genetic

analysis of the response to morphine in mice: Analgesia and

running. Psychopharmacology (Berlin) 42:249-254, 1975.

Spealman, R.D., and Goldberg, S.R. Drug self-administration by

laboratory animals: Control by schedules of reinforcement. Annu

Rev Pharmacol Toxicol 18:313-339, 1978.

Suzuki, T.; Koike, Y.; Yanaura, S.; George, F.R.; and Meisch, R.A.

Genetic differences in the development of physical dependence on

pentobarbital in four inbred strains of rats. Jpn J Pharmacol

45:479-486, 1987.

Tampier, L.; Quintanilla, M.E.; and Mardones, J. Genetic differences

in tolerance to ethanol: A study in UChA and UChB rats.

Pharmacol Biochem Behav 14:165-168, 1981.

Young, A.M., and Herling, S. Drugs as reinforcers: Studies in

laboratory animals. In: Goldberg, S.R., and Stolerman, I.P., eds.

Behavioral Analysis of Drug Dependence. Orlando, FL: Academic

Press, 1986. pp. 967.

Waller, M.B.; McBride, W.J.; Gatto, G.J.; Lumeng, L.; and Li, T.-K.

lntragastric self-infusion of ethanol by ethanol-preferring and

-nonpreferring lines of rats. Science 225:78-80, 1984.

248
Weeks, J.R. Experimental morphine addiction: Method for automatic

intravenous injections in unrestrained rats. Science 138:143-144,

1962.

ACKNOWLEDGMENTS

Mary Rauschenberg, M.S., National Institute on Drug Abuse Addiction

Research Center, provided technical laboratory support and data

evaluation.

AUTHORS

Frank R. George, Ph.D.

Steven R. Goldberg, Ph.D., Chief

Preclinical Pharmacology Branch

National Institute on Drug Abuse

Addiction Research Center

Baltimore, MD 21224

249
Hepatotoxicity of Cocaine

Louis Shuster, Christine A. Garhart, James Powers,

Yona Grunfeld, and Gary Kanel

INTRODUCTION

Fatty liver necrosis in mice that had been fed with cocaine was first

described by Paul Ehrlich (1890). This phenomenon was rediscovered

independently 87 years later in two laboratories in the United States.

Evans et al. (1977) observed that pretreatment with phenobarbital for

4 days decreased the acute toxicity of high doses of cocaine, that is,

the incidence of death from convulsions within 3 hours. This

pretreatment also produced a marked increase in delayed lethality,

that is, deaths from hepatic necrosis during the 7 days after injection

of cocaine.

We encountered cocaine-induced hepatotoxicity while studying

sensitization to the stimulant effects of cocaine. The surface of

livers from mice that had received four or five daily injections of 20

mg/kg cocaine had an unusual pitted or roughened appearance. After

other possible causes such as infection had been ruled out, it was

found that a single injection of 50 mg/kg cocaine could produce

severe fatty necrosis of the liver (Shuster et al. 1977).

THE NATURE OF THE DAMAGE

Histologic examination of cocaine-damaged livers revealed hemorrhagic

fatty necrosis that was mainly midzonal and perivenular, with sparing

of periportal regions (Shuster et al. 1977; Kanel et al. 1979). This

pattern is different from the necrosis produced by carbon tetrachloride and acetaminophen, which is mainly centrilobular. Pretreatment with phenobarbital did cause extension of cocaine-induced

damage to periportal areas, in agreement with the findings of Evans

and Harbison (1978) and Jordan and Franklin (1978).

250
Liver damage from cocaine produced a massive release of transaminases into the circulation. Peak levels of serum glutamateoxaloacetate transaminase (SGOT) were attained at about 16 hours

after the injection of cocaine. There was indirect evidence for

damage to the smooth endoplasmic reticulum in the form of prolonged

sleeptime after the administration of short-acting barbiturates

(Shuster et al. 1977). Other workers have reported a diminution of

the rate of antipyrine metabolism in vivo (Kloss et al. 1983), correlated with a loss of cytochrome P450 (Rauckman 1982a). In addition, electron microscopy by Gottfried et al. (1986) has revealed

damage to mitochrondria and peroxisomes.

The dose-response curve for elevation of SGOT showed a threshold of

about 40 mg/kg (figure 1). The large standard errors reflect marked

individual variability, such as was also encountered when separate

mice were tested at various times after cocaine (figure 2). This

variability is believed to result from the large number of factors that

can affect the hepatotoxicity of cocaine.

STRUCTURAL REQUIREMENTS FOR DAMAGE

Even minor changes in the structure of the cocaine molecule can

abolish hepatotoxicity (table 1). The products of deesterification, i.e.,

benzoyl ecgonine, ecgonine methyl ester, and ecgonine were all

ineffective at a dose of 100 mg/kg (Thompson et al. 1979). Removal

of the carboxymethyl ester residue at position 2, as in tropacocaine,

or even a change in its configuration from axial to equatorial, as in

pseudococaine, also abolished hepatotoxic activity (Thompson et al.

1979).

The phenyltropane analog of cocaine, in which a benzene ring is

attached directly to the tropane ring (Clarke et al. 1973), was

inactive. However, when there was a fluorine on the benzene ring

and an isopropyl ester at position 2, the resulting compound was very

toxic, as determined by SGOT levels (Thompson et al. 1979). Other

tropane analogs such as tropine, atropine, scopolamine, and tropineN-oxide were inactive (Thompson et al. 1979; Freeman and Harbison

1981b).

THE ROLE OF COCAINE METABOLISM

Metabolites that resulted from oxidation of the N-methyl position by

the cytochrome P450 system, that is, norcocaine and N-hydroxynorcocaine, were more potent hepatotoxins than cocaine (Thompson et al.

251
FIGURE 1. Response of SGOT to different doses of cocaine HCI

NOTE: SGOT was determined according to Shuster et al. (1978). Blood samples were

taken at 24 hours after Injection. Each point represents the mean ± SEM for

a different group of 10 mice.

SOURCE: Shuster et al. 1977. Copyright 1977, Pergamon Press.

1979). These observations, together with others, suggest that, in

order to produce liver damage, cocaine may have to be activated by

oxidative metabolism, as is the case with carbon tetrachloride and

acetaminophen.

Evans and Harbison (1978) had used phenobarbital, an inducer of the

cytochrome P450 system, in order to make mice susceptible to liver

damage from cocaine. Such induction did not appear to be necessary

in our experiments. However, it turned out that our mice were

exposed to a different inducer, that is, softwood bedding (Ferguson

252
FIGURE 2. Time course of changes in SGOT after the injection of

50 mg/kg cocaine HCI

NOTE: Each curve represents an individual mouse.

SOURCE: Shuster et al. 1977, Copyright 1977, Pergamon Press.

253
TABLE 1. Structure of cocaine, some metabolites, and related

compounds

Compound R R' R''

Cocaine

Norcocaine

Benzoylecgonine

Ecgonine methyl ester

Ecgonine

N-hydroxynorcocaine

Tropacocaine

Phenyltropane 2a

Phenyltropane 22

C H 3

H

CH3

CH3

CH3

OH

CH3

CH3

CH3

COOCH3

COOCH3

COOH

CH3

COOH

COOCH3

H

COOCH3

COO-i-pr

OOCC6H5

OOCC6H5

OOCC6H5

COOCH3OH

OH

OOC6H5

OOC6H5

C6H5

C6H4F

1966). Beta ionone, an essential oil found in cedarwood, markedly

increased susceptibility to liver damage from cocaine. Methyl

cholanthrene, an inducer of cytochrome P448, was ineffective

(Thompson et al. 1979). As reported by Smith et al. (1981), cocaine

is also hepatotoxic when injected into mice that have been induced

by pretreatment with ethanol. Liver damage from cocaine could be

obtained in the absence of any pretreatment if the cocaine was

administered chronically, that is, in the form of two or three daily

injections (Freeman and Harbison 1981a; Thompson et al. 1984). This

finding raises the possibility that cocaine itself might act as an

inducer of the mixed-function oxidase system of the smooth endoplasmic reticulum. Puma and Ramos-Aliaga (1981) have presented evidence that chronic administration of cocaine increases the amount of

microsomal protein and cocaine N-demethylase in rat livers.

Inhibitors of the cytochrome P450 system, such as SKF525A,

iproniazid, and chloramphenicol, prevented liver damage when injected

before cocaine, presumably because they prevent the formation of

toxic metabolites (Thompson et al. 1979).

254
The oxidation of cocaine by the cytochrome P450 system is believed

to account for only 10 or 15 percent of overall cocaine metabolism.

The remainder is rapidly hydrolyzed by plasma and tissue esterases

(Stewart et al. 1978). It is therefore not surprising that the

hepatotoxicity of cocaine is markedly potentiated by pretreatment

with esterase inhibitors such as diazinon (Thompson et al. 1979) and

tri-o-tolyl phosphate (Freeman and Harbison 1981 b) even when the

cytochrome P450 system has not been induced.

The postulated pathway for the metabolic activation of cocaine is

depicted in figure 3. There is good evidence for the in vitro

formation of norcocaine (Stewart et al. 1978; Evans and Harbison

1978) and N-hydroxynorcocaine (Shuster et al. 1983). These

metabolites can also be detected in vivo, especially in mice that have

been pretreated with an esterase inhibitor (Shuster et al. 1983; Evans

1983; Benuck et al., in press). Our results suggest that the formation

of N-hydroxynorcocaine is carried out primarily by the cytochrome

P450 system (Shuster et al. 1983). Others claim that the flavinlinked monooxygenase system is responsible for this step (Kloss et al.

1982).

The cytochrome P450-catalyzed formation of norcocaine nitroxide is

detectable in vitro by electron spin resonance (ESR) spectroscopy

(Evans and Johnson 1981; Evans 1982; Rauckman et al. 1982b). This

conversion seems to be a necessary step for hepatotoxicity, because

inhibitors of the cytochrome P450 system prevent liver damage from

N-hydroxynorcocaine (Thompson et al. 1979).

THE ULTIMATE HEPATOTOXIN

Norcocaine nitroxide, being a free radical, was initially believed to be

the metabolite that actually damaged hepatocytes, either directly, by

reacting with membranes, or indirectly, by depleting glutathione. The

loss of glutathione could then permit the buildup of toxic peroxides

that are formed in the course of normal metabolism (Evans and

Johnson 1981; Rauckman et al. 1982a).

However, it soon became apparent that norcocaine nitroxide is a

stable free radical resembling substituted piperidine nitroxides that

are used as spin labels. Norcocaine nitroxide is stable in solution

and does not react spontaneously with either glutathione or proteins

(Rauckman et al. 1982b).

255
FIGURE 3. Postulated pathway for the metabolic activation of

cocaine to a hepatotoxic free radical

256
Rauckman et al. (1982a) and Kloss et al. (1984) have suggested that

enzymatically catalyzed cycling between N-hydroxynorcocaine and

norcocaine nitroxide may lead to liver damage by depleting NADPH

and eventually reduced the amount of glutathione. However, 2,2,6,6-

tetramethylpiperidinoxyl (TEMPO) undergoes such cycling (Rosen and

Rauckman 1977) (figure 4), and this compound is not hepatotoxic

when injected in a dose of 100 mg/kg (Shuster and Powers, unpublished).

FIGURE 4. The reversible reduction of 2,2,6,6-tetramethylpiperidinoxyl

(TEMPO) to 1-hydroxy-2,2,6,6-tetramethylpiperidine (NOH-TEMPO)

NOTE: According to Rosen and Rauckman (1977), the reduction is carried out by

cytochrome P450 and NADPH. while the oxidation Is catalyzed by a flavin

monooxygenase.

We have proposed a different mechanism (Charkoudian and Shuster

1985). The one-electron oxidation of nitroxides such as TEMPO gives

rise to extremely reactive nitrosonium ions (Golubev et al. 1965).

The nitrosonium ion of norcocaine has been generated by chemical

and electrochemical oxidation of norcocaine nitroxide. Such ions

react readily with glutathione, and can produce hemolysis of red

blood cells in vitro (figure 5). Biochemical formation of nitrosonium

ions could be catalyzed by cytochrome P450 acting as a peroxidase

together with either hydrogen peroxide or a lipid hydroperoxide

(Blake and Coon 1981). Semmelhack et al. (1983) and Semmelhack

and Schmid (1983) have reported that electrochemically generated

nitrosonium ions are very effective oxidizing agents, and can convert

alcohols and amines to aldehydes and ketones in high yield. The

257
FIGURE 5. Hemolysis of mouse etythrocytes by the nitrosonium

ion of TEMPOL

SOURCE: Charkoudiin and Shuster 1985, Copyright 1985, Academic Press.

resulting N-hydroxy derivative is reoxidized to the nitrosonium ion so

that it acts as a catalyst. We have found that nitrosonium ions

readily oxidize aspartate to oxaloacetate in aqueous solution. They

also oxidize lipids to produce malondialdehyde.

THE POSSIBLE ROLE OF GLUTATHlONE DEPLETION

The hepatotoxicity of cocaine is potentiated when mice are pretreated

with diethyl maleate in order to deplete the liver of glutathione

(Evans and Harbison 1978). Sulfhydryl compounds such as cysteine

and cysteamine protect against liver damage from cocaine (Evans and

Harbison 1978; Thompson et al. 1979). These observations suggest

that, as in the case of liver damage from acetaminophen, the

depletion of glutathione may be an important contributing factor in

cocaine hepatotoxicity.

258
However, liver damage in acetaminophen-injected animals is associated

with a decrease of 80 percent or more in the level of reduced

glutathione (Mitchell et al. 1973). The extent of glutathione

depletion encountered by us (Thompson et al. 1979) and by others

(Evans and Harbison 1978) in the livers of cocaine-injected mice was

in the range of 20 to 40 percent. A recent abstract by Boyer et al.

(1987) reported a decrease of 50 to 65 percent after 60 mg/kg

cocaine. Considering that an overnight fast can deplete glutathione

levels in mouse liver by 50 percent without causing any liver damage

(Strubelt et al. 1981), it appears unlikely that such depletion is by

itself the cause of cocaine-induced hepatotoxicity. Even in the case

of acetaminophen, it has been found that pretreatment with cimetidine will prevent the liver damage in the face of a marked drop in

glutathione levels (Peterson et al. 1983).

The depletion of glutathione from the livers of cocaine-injected mice

may be a secondary effect of a primary lesion. Mitochondrial

disruption 2 hours after the injection of cocaine has been described

(Gottfried et al. 1986). A primary loss of mitochondrial function,

particularly oxidative phosphorylation, may be responsible for the 90

percent depletion of ATP after 2 hours, as well as an 85 percent loss

of NADPH at 12 hours (Boyer et al. 1987).

LIPID PEROXIDATION

Whether lipid peroxidation is a cause or an effect of liver damage

from cocaine is still unresolved. Cornette et al. (1985) did not

observe any marked increase in thiobarbiturate-reactive material in

the livers of mice that had been injected with cocaine. According to

Kloss et al. (1984), the oxidation of cocaine in the liver can lead to

the formation of superoxide radicals, hydrogen peroxide, and lipid

peroxides. These active forms of oxygen, rather than any freeradical metabolites of cocaine, may be responsible for damage to liver

membranes (Rauckman et al. 1982; Rosen et al. 1982). However, this

explanation does not account for the covalent binding of labeled

cocaine metabolites to liver proteins. This binding correlated with

liver damage, and was not seen in rats. More labeled N-hydroxynorcocaine than labeled cocaine was bound (Evans 1983).

THE ROLE OF CATECHOLAMINES

Recent observations from our laboratory suggest that catecholamines

play an important role in liver damage from cocaine.

259
(1) Adrenalectomy prevents cocaine-induced hepatotoxicity. The

effect of adrenalectomy is reversed by subcutaneous injection of

epinephrine in oil (Shuster and Thompson 1985).

(2) Pretreatment with alpha blockers such as prazocin or dibenzyline, or depletion of catecholamines by reserpine, also

prevents liver damage from cocaine (figure 6). James et al.

(1987) have reported that the alpha-adrenergic antagonists

FIGURE 6. Effect of adrenalectomy and pretreatment with reserpine

or alpha blockers on liver damage from cocaine

NOTE: Sham-operated or adrenalectomized B6AF1

/J male mice were maintained on 1

percent NaCl in their drinking water for 1 week before being injected with

cocaine HCI, 50 mg per kg IP. Pretreatments were: reserpine, 5 mg/kg IP 5

days before cocaine; prazocin, 1 mg/kg 30 minutes before cocaine: dibenzyline.

25 mg/kg 5 hours before cocaine. Each group consisted of four to six mice.

Blood samples for SGOT determinations were taken 18 hours after cocaine

injection.

260
phentolamine and yohimbine decreased both liver damage and

glutathione depletion by cocaine.

(3) High doses of epinephrine in oil (between 2 and 7 mg/kg) can

by themselves produce fatty liver damage and elevation of SGOT

levels (figure 7). Other workers have reported that epinephrine

can also lower liver glutathione levels (Register and Bartlett

1954; James et al. 1983).

FIGURE 7. Effect of epinephrine on SGOT levels

NOTE: Separate groups of four to six male B6AF1

/J male mice were injected subcutaneously with different doses of L-epinephrine bitartarate in peanut oil.

Eighteen hours later, the mice were bled for the determination of SGOT.

261
(4) The injection of high doses of cocaine increases the levels of

circulating epinephrine and norepinephrine twofold to fourfold

(figure 8). Similar findings have been made in rats (Chiueh

and Kopin 1978).

lt is not unreasonable to suggest that catecholamines may, by

stimulating hormone-sensitive lipases, cause the release of free fatty

acids and increased formation of lipid hydroperoxides. These

hydroperoxides, in addition to causing damage directly, could also

FIGURE 8. Effect of cocaine on plasma levels of norepinephrine,

epinephrine, and serotonin

NOTE: At each time, a separate group of 10 to 20 male B6AF1/J mice were bled from

the orbital sinus. Plasma monoemines were determined by HPLC with

electrochemical detection.

262
serve as cofactors for the oxidation of norcocaine nitroxide to the

nitrosonium ion.

Those cocaine analogs and metabolites that do not elevate catecholamine should not be hepatotoxic. For example, pseudococaine, which

appears to be metabolized like cocaine (Misra and Pontani 1977),

even to the formation of a nitroxide (Charkoudian and Shuster,

unpublished), is not hepatotoxic. lt is also much less effective than

cocaine in blocking the reuptake of monoamines (Williams et al. 1977),

although it does possess comparable local anesthetic activity

(Matthews and Collins 1983.

EARLY CHANGES IN LIVER FUNCTION

The clearance of bromsulfophthalein (BSP) is inhibited by cocaine.

An injection of 10 mg/kg, which is not hepatotoxic, will decrease BSP

clearance, as demonstrated by elevated plasma levels of the dye,

within 1 hour (figure 9). Part of the effect of higher doses of

cocaine may be attributed to elevated plasma levels of epinephrine

and serotonin, because these substances also interfere with clearance

(figure 10).

BSP is extensively conjugated to glutathione before it is excreted

into bile. lndocyanine green (ICG) is removed from the liver without

prior metabolism. Cocaine did not affect ICG clearance at 1 hour in

doses below 60 mg/kg (figure 9). A similar distinction between BSP

and ICG clearance was obtained with hepatotoxic doses of carbon

tetrachloride. These findings suggest that cocaine may be acting to

prevent the conjugation of BSP, perhaps by inhibiting or damaging

glutathione transferase.

We have also found that cocaine can alter liver perfusion and bile

secretion. When rat livers were subjected to nonrecirculating

perfusion with cocaine, there was a dose-related increase in perfusion

pressure, an indication that cocaine produces vasoconstriction. Low

doses of cocaine increased bile flow, but high doses produced drastic

inhibition (figure 11). Similar concentrations of tropacocaine, a

nonstimulating analog with local anesthetic activity, and other local

anesthetics such as procaine and lidocaine, had little effect on either

perfusion pressure or bile flow.

263
FIGURE 9. Effect of cocaine on dye uptake

NOTE: Separate groups of B6AF1/J male mice (8 to 10 for BSP, 5 for ICG) were

injected with varying doses of cocaine. After 1 hour, they were injected with

dye and then bled from the orbital sinus. Plasma dye concentrations were

determined according to Hurwitz et al. (1985).

264
FIGURE 10. Effect of monoamine injections on dye uptake

NOTE: The monoamines were injected subcutaneously in peanut oil 1 hour before dye

injection. From 6 to 11 male B6AF1/J mice per group were used to measure

BSP uptake and from 6 to 9 for ICG uptake.

The hepatic vasoconstriction produced by cocaine was not blocked by

concentrations of phentolamine that prevented completely the vasoconstriction caused by phenylephrine. This observation suggests that

the cocaine is not acting through an adrenergic mechanism.

THE POSSIBLE ROLE OF CALCIUM IONS

Thor et al. (1985) have suggested that oxidative damage to hepatocytes may require the mobilization of calcium ions secondary to a

depletion of glutathione. The infusion of norepinephrine into rabbits

at a rate of 2 μg/kg/min for 90 minutes produces massive hepatic

necrosis and calcium mineralization. These changes are prevented by

prazocin (Lee et al. 1986).

265
FIGURE 11. Effects of cocaine on perfused isolated rat liver

NOTE: A single-pass perfusion was set up according to Anwer and Hegner (1983).

266
We have added cocaine to isolated rat hepatocytes that had been

loaded with the fluorogen Quin 2. In millimolar concentrations,

cocaine by itself produced little change in free calcium levels.

However, it did inhibit strongly the release of intracellular calcium

ions produced by adding epinephrine. There was also inhibition of

epinephrine-induced glycogenolysis. Whether these changes are

related to the hepatotoxicity of cocaine is unclear, because they can

also be produced by somewhat higher concentrations of lidocaine and

other local anesthetics. In this case, cocaine is not acting like

epinephrine (McConnell et al. 1987).

The direct effects of cocaine on the liver, such as those described

here, may involve the specific binding sites for cocaine that have

been described by Sershen et al. (1982) and Calligaro and Eldefrawi

(1987).

CCCAINE-INDUCED HEPATOXICITY IN SPECIES OTHER THAN

MICE

Species differences are a striking feature of liver damage from

cocaine. They may be determined in part by differences in esterase

activity and in the inducibility of mixed function oxidases. These

variables appear to play a role in the susceptibility of different

mouse strains and sexes (Thompson et al. 1984). At least one strain

of mice, DBA/2, shows appreciable damage without preinduction of

the cytochrome P450 system (Kloss et al. 1982).

Evans (1978) did not find liver damage from cocaine in rats, guinea

pigs, or rabbits. We have produced some damage, as indicated by a

moderate elevation of SGOT levels, in rats and Syrian hamsters. The

increases are much less than those seen in mice, and there does not

seem to be any potentiation when the animals are pretreated with

phenobarbital. In both species, the response was greater in females

than in males (figure 12). Watanabe et al. (1987) have reported that

cocaine produces more liver damage in spontaneously hypertensive

rats than in Wistar-Kyoto rats.

COCAINE HEPATOTOXICITY IN HUMAN ADDICTS

Until recently, the evidence for cocaine-induced liver damage in

humans was limited to a report by Marks and Chapple (1967). These

authors described elevated levels of serum transaminases in 80

percent of 89 addicts who were taking both cocaine and heroin.

267
FIGURE 12. Effect of cocaine on SGOT levels in rats and

Syrian hamsters

NOTE: Pretreatment consisted of four daily injections of either 0.15 M NaCl (S) or

phenobarbital, 50 mg/kg (P). On the 5th day, Sprague-Dawley rats were

injected with 75 mg/kg cocaine HCI IP (C), and hamsters were injected with 50

mg/kg. Blood was collected 18 hours after cocaine. Each group contained

four animals.

Values returned to normal when the drugs were stopped, and went

up again upon resumption of the addiction.

268
This year, Perino et al. (1987) described the case of a 32-year-old

man who was abusing a variety of drugs. He was admitted to the

hospital exhibiting bizarre behavior after snorting cocaine. His blood

tested positive for ethanol, phenobarbital, secobarbital, and methaqualone. Following respiratory arrest, resuscitation, and a downhill

course, he died 72 hours after admission. His SGOT level was 620

units/l upon admission, and 10,700 at 28 hours. His serum creatine

phosphokinase activity (CPK) upon admission was 7,490 units/l.

At autopsy, there was extensive hepatonecrosis. The necrosis was

periportal and midzonal. Centrilobular hepatocytes were intact.

Midzonal hepatocytes displayed fatty changes and mitotic regeneration. Perino et al. (1987) emphasized the resemblance of these

changes to those observed in cocaine-injected mice.

Five similar cases have now been encountered at the University of

Southern California Liver Unit. One was a 20-year-old male who

presented with hepatic necrosis, serum glutamate-pyruvate transaminase and SGOT levels in the range of 2,000 to 4,000 units/l,

rhabdomyolysis (CPK levels ranging from 14,000 to 28,000 units/l),

and renal failure. Death was attributed to submassive hepatic

necrosis with pneumonia. The pattern of necrosis was primarily

perivenular and midzonal. Periportal hepatocytes showed some fatty

changes.

Considering the large number of people who abuse cocaine, it is

surprising that such cases are so rare. There is a general tendency

to attribute any liver damage in drug addicts to viral infections.

Instances of cocaine-induced liver damage in the absence of any viral

hepatitis are likely to be uncommon unless one or more of the

following precipitating factors are also present:

(1)

(2)

(3)

Pretreatment with inducers of the cytochrome P450 system, such

as phenobarbital, phenytoin, or ethanol.

Exposure to esterase inhibitors, such as organophosphate

insecticides.

A genetic deficiency of pseudocholinesterase. About 1 person in

2,500 has an atypical level that is 50 percent of normal values.

However, this level may still be sufficient for rapid hydrolysis

of cocaine (La Du 1971).

269
(4) Coadministration of cocaine with an opiate such as heroin.

Several investigators have shown that narcotic drugs can

produce fatty liver infiltration and elevated serum transaminase

levels in mice (Thureson-Klein et al. 1978; Needham et al. 1981).

We have observed synergistic hepatotoxicity in mice injected

with cocaine plus morphine.

Additional information about the requirements for liver damage from

cocaine in primates may come from experiments with monkeys. Liver

damage has shown up in rhesus monkeys that were injected repeatedly with 32 mg/kg cocaine (Kleven and Woolverton, personal

communication, 1987).

CONCLUSION

The oxidative pathway of cocaine metabolism, while relatively minor

in comparison to hydrolysis by esterases, gives rise to active

metabolites capable of producing severe hepatic necrosis. In addition,

cocaine can act directly on the liver to produce vasoconstriction and

inhibition of bile secretion. Susceptibility to liver damage from

cocaine is influenced by many different factors, so that frank hepatotoxicity is not likely to become a common feature of cocaine addiction in humans. Studies of this phenomenon have contributed to our

understanding of cocaine metabolism and its sequelae.

REFERENCES

Anwer, M.S., and Hegner, D. Sodium and chloride dependency of

dibucaine- and procaine-induced choleresis in isolated perfused rat

liver. J Pharmacol Exp Ther 225:284-290, 1983.

Benuck, M.; Reith, M.E.A.; and Lajtha, A. Presence of the toxic

metabolite N-hydroxynorcocaine in brain and liver of the mouse.

Biochem Pharmacol 37:1169-1172, 1988.

Blake, R.C. II, and Coon, M.J. On the mechanism of action of

cytochrome P-450. Evaluation of homolytic and heterolytic

mechanisms of oxygen-oxygen bond cleavage during substrate

hydroxylation by peroxides. J Biol Chem 256:12127-12133, 1981.

Boyer, C.S.; Ross, D.; and Peterson, D.R. In vivo depletion of hepatic

glutathione, NADPH and ATP following acute cocaine administration

in DBA mice. Hepatology 7:1046, 1987.

Calligaro, D.O., and Eldefrawi, M.E. Central and peripheral cocaine

receptors. J Pharmacol Exp Ther 243:61-68, 1987.

270
Charkoudian, J.C., and Shuster, L. Electrochemistry of norcocaine

nitroxide and related compounds: Implications for cocaine

hepatotoxicity. Biochem Biophys Res Commun 130:1044-1051, 1985.

Chiueh, C.C., and Kopin, I.J. Centrally mediated release by cocaine

of endogenous epinephrine from the sympathoadrenal medullary

system of unanesthetized rats. J Pharmacol Exp Ther 205:148-154,

1978.

Clarke, R.L.; Daum, S.J.; Gambino, A.J.; Aceto, M.D.; Pearl, J.; Levitt,

M.; Cumiskey, W.R.; and Bogado, E.F. Compounds affecting the

central nervous system. 4,3 beta-phenyltropane-2-carboxylic esters

and analogs. J Med Chem 18:1260-1267, 1973.

Cornette, C.; Serrar, D.; Lallemant, A.; and Thevenin, M. Modifications de I’hepato-toxicite de la cocaine chez la Souris Swiss sous

I’influence de divers inhibiteurs. C R Soc Biol 179:320-326, 1985.

Ehrlich, P. Studien in der cocainreihe. Deutsch Med Wochenschr

16:717-719, 1890.

Evans, M.A. Role of metabolism in cocaine-induced hepatic necrosis.

The Pharmecologist 20:182, 1978.

Evans, M.A. Microsomal activation of N-hydroxy norcocaine to a

reactive nitroxide. Toxicologist 1 :1 , 1982.

Evans, M.A. Role of protein binding in cocaine-induced hepatic

necrosis. J Pharmacol Exp Ther 224:73-79, 1983.

Evans, M.A.; Dwivedi, C.; and Harbison, R.D. Enhancement of

cocaine-induced lethality by phenobarbital. In: Ellinwood, E.H.,

Jr., and Kilbey, M.M., eds. Cocaine and other stimulants.

Advances in Behavioral Biology. New York: Plenum Press, 1977.

pp. 253-267.

Evans, M.A., and Harbison, R.D. Cocaine-induced hepatotoxicity in

mice. Toxicol Appl Pharmacol 45:739-754, 1978.

Evans, M.A., and Johnson, M.E. The role of a reactive nitroxide

radical in cocaine-induced hepatic necrosis. Fed Proc 40:638, 1981.

Ferguson, H.C. Effect of cedar chip bedding on hexobarbital and

pentobarbital sleep time. J Pharm Sci 55:1142-1143, 1966.

Freeman, R.W., and Harbison, R.D. Hepatic periportal necrosis

induced by chronic administration of cocaine. Biochem Pharmacol

30:777-783, 1981a.

Freeman, R.W., and Harbison, R.D. The role of benzoylmethyl

ecgonine in cocaine-induced hepatotoxicity. J Pharmacol Exp Ther

218:558-567, 1981b.

Garhart, C.A.; Anwer, M.S.; and Shuster, L. Cocaine directly induces

vasoconstriction in the isolated perfused rat liver. FASEB J

2:A1803, 1988.

271
Golubev, V.A.; Rozantsev, G.; and Neiman, M.B. Some reactions of

free imminoxyl radicals with the participation of the paired

electron. Bull Acad Sci USSR: 1927-1936, 1965.

Gottfried, M.R.; Kloss, M.W.; Graham, D.; Rauckman, E.J.; and Rosen,

G.M. Ultrastructure of experimental cocaine hepatotoxicity.

Hepatology 6:299-304, 1986.

Hurwitz, A.; Fischer, H.R.; Innis, J.D.; Rousse, S.; and Ben-Zvi, Z.

Opioid effects in hepatic disposition of dyes in mice. J Pharmacol

Exp Ther 232:617-623, 1985.

James, R.C.; Roberts, S.M.; and Harbison, R.D. The perturbation of

hepatic glutathione by alpha 2-adrenergic agonists. Fundam Appl

Toxicol 3:303-308, 1983.

James, R.C.; Schiefer, M.A.; Roberts, S.M.; and Harbison, R.D.

Antagonism of cocaine-induced hepatotoxicity by the alpha

adrenergic antagonists phentolamine and yohimbine. J Pharmacol

Exp Ther 242:726-732, 1987.

Jordan, R.A., and Franklin, M.R. Cocaine hepatotoxicity in mice.

The Pharmacologist 20:258, 1978.

Kanel, G.C.; Engler, S.J.; Thompson, M.L.; and Shuster, L. Cocaineinduced hepatic necrosis in mice. Gastroenterology 77:A20, 1979.

Kleven, M.S., and Woolverton, W.L. Personal communication, 1987.

Kloss, M.W.; Rosen, G.M.; and Rauckman, E.J. Acute cocaine-induced

hepatotoxicity in DBA/2H male mice. Toxicol Appl Pharmacol

65:75-83, 1982.

Kloss, M.W.; Rosen, G.M.; and Rauckman, E.J. N-demethylation of

cocaine to norcocaine: Evidence for participation by cytochrome P450 and FAD containing monooxygenase. Mol Pharmacol 23:482-485,

1983.

Kloss, M.W.; Rosen, G.M.; and Rauckman, E.J. Cocaine-mediated

hepatotoxicity. A critical review. Biochem Pharmacol 33:169-173,

1984.

La Du, B.N. Genetic factors modifying drug metabolism and drug

response. In: La Du, B.N.; Mandel, H.G.; and Way, E.L., eds.

Fundamentals of Drug Metabolism and Drug Disposition. Baltimore:

Williams and Wilkins, 1971. pp. 308-327.

Lee, J.C.; Saunders, G.K.; and Sponenberg, D.P. Norepinephrine

induces hepatic necrosis in rabbits. The Pharmacologist 28:107,

1986.

Marks, V., and Chapple, P.A.L. Hepatic dysfunction in heroin and

cocaine users. Br J Addict 62:189-195, 1967.

Matthews, J.C., and Collins, A. Interactions of cocaine and cocaine

congeners with sodium channels. Biochem Pharmacol 32:455-460,

1983.

272
McConnell, J.; Anwer, M.S.; Engelking, L.R.; Nolan, K.; and Shuster,

L. Cocaine and lidocaine inhibit epinephrine-induced changes in

cytosolic free Ca++ of rat hepatocytes. Fed Proc 46:1231, 1987.

Misra, A.L., and Pontani, R.B. Disposition and metabolism of pseudacocaine (dextro-cocaine) in the rat. Drug Metab Dispos 5:556-563,

1977.

Mitchell, J.R.; Jollow, D.J.; Potter, W.Z.; Gillette, J.R.; and Brodie,

B.B. Acetaminophen-induced hepatic necrosis. IV. Protective role

of glutathione. J Pharmacol Exp Ther 187:211-217, 1973.

Needham, W.P.; Shuster, L.; Kanel, G.C.; and Thompson, M.L. Liver

damage from narcotics in mice. Toxicol Appl Pharmacol 58:157-170,

1981.

Perino, L.E.; Warren, G.H.; and Levine, J.S. Cocaine-induced

hepatotoxicity in humans. Gastroenteology 93:178-180, 1987.

Peterson, F.J.; Knodell, R.G.; Lindemann, N.J.; and Steele, N.M.

Prevention of acetaminophen and cocaine hepatotoxicity in mice by

cimetidine treatment. Gastroenterology 85:122-129, 1983.

Puma, G., and Ramos-Aliaga, R. Adaptive increases in liver cocaine

N-demethylation induced by the drug in rats receiving different

levels of utilizable protein. II. Evidences of new enzymatic

protein synthesis and adrenal gland participation. Res Commun

Subst Abuse 2:47-55, 1981.

Rauckman, E.J.; Kloss, M.W.; and Rosen, G.M. Involvement of

nitroxide radicals in cocaine-induced hepatotoxicity. Can J Chem

60:1614-1620, 1982a.

Rauckman, E.J.; Rosen, G.M.; and Cavagnaro, J. Norcocaine nitroxide.

A potential hepatotoxic metabolite of cocaine. Mol Pharmacol

21:458-463, 1982b.

Reed, D.J., and Fariss, M.W. Glutathione depletion and susceptibility.

Pharmacol Rev 36:255-335, 1984.

Register, U.D., and Bartlett, R.G., Jr. Relationship of adrenalin to

tissue sulfhydryl compounds. Science 120:109-110, 1954.

Rosen, G.M., and Rauckman, E.J. Formation and reduction of a

nitroxide radical by liver microsomes. Biochem Pharmacol 26:675-

678, 1977.

Rosen, G.M.; Kloss, M.W.; and Rauckman, E.J. Initiation of in vitro

lipid peroxidation by N-hydroxynorcocaine and norcocaine nitroxide.

Mol Pharmacol 22:529-531, 1982.

Semmelhack, M.F.; Chou, C.S.; and Cortes, D.A. Nitroxyl-mediated

electrooxidation of alcohols to aldehydes and ketones. J Am Chem

Soc 105:4492-4494, 1983.

Semmelhack, M.F., and Schmid, C.R. Nitroxyl-mediated electrooxidation of amines to nitriles and carbonyl compounds. J Am

Chem Soc 105:6732-6734, 1983.

273
Sershen, H.; Reith, M.E.A.; and Lajtha, A. Comparison of central and

peripheral binding sites for cocaine. Neuropharmacology 21:469-474,

1982.

Shuster, L.; Bates, A.; and Hirsch, C.A. A sensitive radiochemical

assay for serum glutamic-oxaloacetic transaminase. Anal Biochem

86:648-654, 1978.

Shuster, L., and Thompson, M.L. Epinephrine and liver damage from

cocaine. The Pharmacologist 27:200, 1985.

Shuster, L; Casey, E.; and Welankiwar, S.S. Metabolism of cocaine

and norcocaine to N-hydroxynorcocaine. Biochem Pharmacol

32:3045-3051, 1983.

Shuster, L.; Quimby, F.; Bates, A.; and Thompson, M.L. Liver damage

from cocaine in mice. Life Sci 20:1035-1042, 1977.

Smith, A.C.; Freeman, R.W.; and Harbison, R.D. Ethanol enhancement

of cocaine-induced hepatotoxicity. Biochem Pharmacol 30:453-458,

1981.

Stewart, D.J.; Inaba, T.; and Kalow, W. N-demethylation of cocaine

in the rat and in isolated rat hepatocytes: Comparison with

aminopyrine demethylation. J Pharmacol Exp Ther 207:171-177,

1978.

Stewart, D.J.; Inaba, T.; Lucassen, M.; and Kalow, W. Cocaine

metabolism: Cocaine and norcocaine hydrolysis by liver and serum

esterases. Clin Pharmacol Ther 25:464-468, 1979.

Strubelt, O.; Dost-Kempf, E.; Siegers, C.P.; Younes, M.; Volpel, M.;

Preuss, U.; and Dreckmann, J.G. The influence of fasting on the

susceptibility of mice to hepatotoxic injury. Toxicol Appl

Pharmacol 60:66-77, 1981.

Thompson, M.L.; Shuster, L.; and Shaw, K. Cocaine-induced hepatic

necrosis in mice. The role of cocaine metabolism. Biochem

Pharmacol 28:2389-2395, 1979.

Thompson, M.L.; Shuster, L.; Casey, E.; and Kanel, G.C. Sex and

strain differences in response to cocaine. Biochem Pharmacol

33:1299-1307, 1984.

Thor, H.; HartzelI, P.; Svensson, S.; Orrenius, S.; Mirabelli, F.;

Marinoni, V.; and Bellomo, G. On the role of thiol groups in the

inhibition of liver microsomal Ca2+ sequestration by toxic agents.

Biochem Pharmacol 34:3717-3723, 1985.

Thureson-Klein, A.; Wang-Yang, J.; and Ho, I.K. Lipid accumulation

in mouse hepatocytes after morphine exposure. Experientia 34:773-

774, 1978.

Watanabe, H.K.; Hoskins, B.; and Ho, I.K. Sensitivity difference to

hepatotoxicity of cocaine in spontaneously hypertensive and WistarKyoto rats. Alcohol Drug Res 7:363-370, 1987.

274
Watanabe, H.K.; Hoskins, B.; and Ho, I.K. Effects of subacute

treatment with cocaine on activities of N-demethylase, UDPglucuronyltransferase and sulfotransferase in WKY and SHR rat

liver-sex and strain differences. Life Sci 42:79-86, 1988.

Williams, N.; Clouet, D.H.; Misra, A.L.; and Mule, S. Cocaine and

metabolites: Relationship between pharmacological activity and

inhibitory action on dopamine uptake into striatal synaptosomes.

Prog Neuro-Psychopharmacol 1:265-269, 1977.

ACKNOWLEDGMENTS

The work described here was supported by research grant DA-01885

from the National Institute on Drug Abuse.

AUTHORS

Louis Shuster, Ph.D.

Christine A. Garhart, D.V.M.

James Powers, B.S.

Yona Grunfeld, D.V.M.

Department of Pharmacology

Tufts University School of Medicine

Boston, MA 02111

Gary Kanel, M.D.

Department of Pathology

University of Southern California

School of Medicine

Los Angeles, CA 90033

275
Lack of Toxic Effects of Cocaine

on Dopamine or Serotonin

Neurons in the Rat Brain

Lewis S. Seiden and Mark S. Kleven

INTRODUCTlON

Due to their euphoric effects, methamphetamine (MA) and related

psychomotor stimulants have been abused, at times in epidemic

proportions, in various countries including Japan, Great Britain,

Sweden, and the United States. Abuse of MA and related drugs often

induces short-lasting behavioral episodes that closely resemble

paranoid schizophrenia (Snyder 1973). Because of its abuse potential

and related behavioral toxicity, there has been considerable interest

in assessing possible neurotoxic properties of MA and related

compounds.

This chapter will review data indicating that MA and other psychomotor stimulants, as well structurally related compounds, are toxic to

dopamine (DA)- and/or serotonin (5-hydroxytryptamine; 5-HT)containing neurons in brain. This chapter will present data showing that

cocaine, a psychomotor stimulant like amphetamine, does not cause

neurotoxicity to DA or 5-HT neurons in the central nervous system.

The lack of toxic effects of cocaine is similar to that found with

methylphenidate (Wagner et al. 1980a). Both drugs are psychomotor

stimulants, but neither appears to be toxic to DA or 5-HT cells in

the brain. We will also review pharmacological similarities and

differences between MA-like compounds and cocaine with regard to

neurotoxicity. This difference between cocaine and drugs related to

MA that cause neurotoxicity may be due to the intracellular storage

pools from which cocaine and MA release monoamines, as well as to

the fact that cocaine seems to be a potent DA reuptake blocker.

The difference in toxicity between MA and cocaine may thus be

related to the differences in the mode of action of the two drugs at

the cellular level in spite of similar behavioral effects. Early studies

276
in rhesus monkeys, rats, cats, and guinea pigs have shown that MA,

amphetamine (AMPH), and related phenethylamines are toxic to DA

and/or 5-HT neurons (tables 1 and 2). The degree of drug-induced

neurotoxicity depends on the duration of drug administration and the

dose. Generally, the neurotoxic dose is severalfold (40 to 50 times)

higher than the dose required for pharmacological effects such as

psychomotor stimulation, anorexia, or stereotypic behaviors, but with

some drugs the toxic dose is only slightly (2 to 3 times) higher than

the behaviorally effective dose.

TABLE 1. Neurochemical effects of chronic methamphetamine

Treatment Survival Effect Reference

3.0-6.5 mg/kg/inj 3-6

8 inj/day, months

3-6 months

50 mg/kg/day x 4 2-8

weeks

15 mg/kg/6 hr x 4

15 mg/kg/ 6 hr x 5

6, 11

days

30

days

50 mg/kg/ 8 hr x 3 2-3

weeks

4 mg/day SC inf x 3 2

weeks

striatal DA

striatal DA

3H-DA uptake

neuronal

degeneration

tryptophan

hydroxylase

tyrosine

hydroxylase

DA metabolism

striatal DA

neuronal

degeneration

Seiden et al.

1975

Wagner et al.

1980b

Lorez 1981

Morgan and

Gibb 1980

Ricaurte et al.

1983

Ricaurte et al.

1984

KEY: increased; decreased.

Early studies with MA have shown that, when this drug is given in

relatively high doses for periods of time ranging from 4 to 180 days,

277
TABLE 2. Neurochemical effects of chronic 3,4-methylene

dioxyamphetamine (MDA) and 3,4-methylene

dioxymethamphetamine (MDMA)

Treatment Survival Effect Reference

MDA

10-40 mg/kg SC 2

bid x 4 days weeks

10 mg/kg/ 6 hr x 5 2

weeks

MDMA

10 mg/kg SC x 1 7 5-HT, 5-HIAA

days striatum

10-40 mg/kg SC bid 2,8

x 4 days weeks

5-HT, 5-HIAA

3H-5-HT uptake

neuronal

degeneration

20 mg/kg SC x 1 7

days

10 mg/kg/ 6 hr x 5 2

weeks

5-HT

3H-5-HT

uptake hippocampus

neuronal

degeneration

5-HT, 5-HIAA

DA, DOPAC,

or HVA

tryptophan

hydroxylase

3H-5-HT uptake

5-HT, 5-HIAA

tryptophan

hydroxylase

Ricaurte et al.

1985

Stone et al.

1987

Schmidt et al.

1986

Commins et al.

1987

Schmidt 1987

Stone et al.

1987

KEY: increased; decreased; 5-HIAA 5-hydroxyindoleacetic acid;

DOPAC dihydroxyphenyiacetic acid; HVA homovanillic acid.

278
levels of DA and 5-HT are depleted in the brain even several months

after the drug is discontinued (table 1). In addition, the DA

depletions appear permanent. The number of uptake sites in the

high-affinity uptake pump are reduced, suggesting that nerve endings

have been lost after MA administration; the affinity constant (Km)

for reuptake did not change, but the maximum number of uptake sites

(Vmax) was lowered. The reduction of high-affinity uptake sites was

proportional to the reduction in amine levels. Morphological studies

also suggested that neuronal degeneration occurred. The FinkHeimer method revealed nerve cells in the striatum of rats killed

shortly after the MA injection in the process of degeneration

(Ricaurte et al. 1982; Ricaurte et al. 1984). The decrease in levels of

DA, the loss of reuptake sites, and the morphological evidence

indicated that MA had toxic effects on DA neurons. Neurotoxicity

has also been reported to occur following repeated administration of

the hallucinogenic amphetamines MDA and MDMA (Ricaurte et al.

1985; Commins et al. 1987). That is, long-lasting decreases in 5-HT

levels, loss of 5-HT reuptake sites, and signs of neuronal degeneration result from repeated or even single injections of these amphetamine congeners. However, 5-HT neurotoxicity is primarily

observed with DA neurons spared under conditions that produce

relatively large effects on 5-HT neurons. We conclude from these

and other studies (tables 1 and 2) that MA, AMPH, MDA, and MDMA

are toxic to DA and/or 5-HT neurons.

In contrast to the prolonged effects reported for amphetaminelike

drugs, most previous studies have reported neurochemical effects

within 24 hours of the last injection of cocaine (table 3). Taylor and

Ho (1976) reported that 45 days of single injections of cocaine

(10 mg/kg) caused an increase in particulate tryptophan hydroxylase

activity measured 24 hours after the last injection. Since this

enzyme is associated with nerve endings, these results suggest that

5-HT metabolism and utilization increased due to the repeated cocaine

regimen. However, the same authors later reported that striatal 5-HT

levels were slightly decreased (to 86 percent of control), and

cytosolic tryptophan hydroxylase activity was reduced as a result of 5

days of single cocaine injections at 10 mg/kg/injection (Taylor and

Ho 1977). These effects were also examined 24 hours after the last

daily cocaine injection but were not consistent with the increase in

5-HT metabolism observed after 45 days of cocaine administration.

With regard to effects of repeated cocaine administration on DA

neurons, a similar regimen (10 mg/kg/day for 7 days) produced an

increase in striatal tyrosine hydroxylase activity 24 hours following

the last injection, with no effect on basal levels of DA or HVA

279
TABLE 3. Neurochemical effects of chronic cocaine

Treatment Survival Effect Reference

Neurochemical

10 mg/kg/day

IPx45

24

hours

10 mg/kg/day IP x 5 24

hours

160 mg/kg/day 10

SCx10 days

20 mg/kg/day x 10

Receptor

10 mg/kg/day

10 mg/kg/day

x7,14

60

days

15-20

minutes

24

hours

particulate

tryptophan

hydroxylase

activity

Taylor and

Ho 1976

NE Taylor and

striatal 5-HT, Ho 1977

5-HIAA

tyrosine hydroxylase activity

tryptophan hydroxylase

3H-GABA binding

striatum

GABA turnover

striatum

tyrosine

hydroxylase

striatum

tyrosine

hydroxylase

nucleus accumbens

Gale 1984

Trulson et al.

1986

Trulson et al.

1987

3H-sulpiride Goeders and

nucleus accumbens Kuhar 1987

3H-sulpiride

striatum

3H-spiroperidol striatum

Taylor et al.

1979

280
TABLE 3. (Continued)

Treatment Survival Effect Reference

Receptor (continued)

20 mg/kg/day x 10 60 3H-spiroperi- Trulson and

days dol striatum Ulissey 1987

KEY: increased; decreased.

(Taylor and Ho 1977). Roy et al. (1978) also found that acute

administration of cocaine no longer decreased DA levels in the

striatum after 30 days of twice-daily cocaine injections (15 mg/kg at

8-hour intervals). Although these results suggest a functional change

(tolerance) as a consequence of the repeated cocaine dosing, the

effect of the repeated administration on levels of DA or 5-HT was

not reported. From these previous studies, it is difficult to conclude

that cocaine has long-lasting toxic effects on DA and 5-HT. More

recent studies have examined longer-lasting effects. Trulson and

colleagues (1986) have shown that cocaine treatment in rats twice a

day for 10 days at 10 mg/kg/injection causes a reduction in tyrosine

hydroxylase immunohistofluorescence staining in striatum 60 days

after the last injection of MA. On this basis, they have suggested

that cocaine is toxic to the DA system. In addition, they have found

that the DA synthesis rate as measured by the accumulation of DOPA

was reduced by up to 50 percent in several brain regions of the rat

when animals were injected with cocaine on the 10-day regimen (i.e.,

10 mg/kg twice a day) (Trulson and Ulissey 1987).

EXPERIMENTAL PROCEDURES AND RESULTS

Experiment 1

Rats were injected twice a day, 12 hours apart, with 10 mg/kg (IP)

cocaine hydrochloride and sacrificed 3 or 60 days after the last

injection. Brains were analyzed for DA and 5-HT levels in the

striatum, hypothalamus, frontal cortex, and hippocampus. There were

no changes in the levels of DA or 5-HT either 3 or 60 days after the

last injection of cocaine (table 4).

281
Experiment 2

Cocaine was delivered intravenously with minipumps implanted under

the skin and attached to a catheter implanted in the exterior jugular

vein. Rats received 100 mg/kg of cocaine per day for 21 days and

were sacrificed 2 weeks after removal of the pump. There were no

differences in DA or 5-HT values between the cocaine-injected

animals and controls (table 5).

Experiment 3

Rats were given 8 IP (100 mg/kg/day) injections 1 hour apart for a

period of 10 days. At 24 hours and 2 weeks after the last injection,

the animals showed no differences in either DA or 5-HT levels in

those regions of the brain examined (figure 1).

TABLE 4. Effect of repeated daily administration of cocaine

(20 mg/kg/day x 10 days) on regional levels of

monoamines and metabolites

Cocaine

Group Cortex Hippocampus Hypothalamus Striatum

5-HT

3 days 106.1 ±6.7

60 days 110.8±5.3

5-HIAA

3 days 99.2±16.9

60 days 112.2±5.1

DA

3 days ND

60 days ND

DOPAC

3 days ND

60 days ND

84.8±5.3 100.1±13.4

97.1±3.3 104.7±6.0

86.9±16.3

102.2±2.7

ND

ND

ND

ND

68.3±13.4

117.7±5.3

108.4±17.3

96.9±6.7

ND

ND

110.2±8.3

NA

115.1±8.9

NA

94.3±6.9

105.3±3.7

77.8±9.3

87.7±7.3

KEY: NA=not available: ND=not determined.

NOTE: Measures represent mean ± SE, percent of control (n=8/group).

282
The results of these experiments indicate that cocaine does not have

long-lasting effects on DA or 5-HT levels in the brain. Since

compounds that have been found to be neurotoxic show decreases in

transmitter levels (tables 1 and 2), these results suggest that cocaine

is not toxic to DA or 5-HT neurons.

FIGURE 1. Effect of continuous administration of cocaine

(100 mg/kg/day x 21 days) on regional levels of

monoamines and metabolites

NOTE: Rats (n=4) were sacrificed 2 weeks after removal of osmotic minipumps

containing cocaine (0.58 g/ml). Values are mean ± SEM of data expressed as a

percentage of sham-treated rats (n=5).

283
DISCUSSION

The results of Trulson and colleagues (Trulson et al. 1986; Trulson et

al. 1987; Trulson and Ulissey 1987) are not consistent with the results

presented in this paper. We expect that, if levels or activity of the

rate-limiting enzyme were decreased, then one would find a corresponding decrease in the amount of transmitter. While our studies

are an indirect replication of those of Trulson et al., it would seem

that, if the tyrosine hydroxylase in DA-containing areas is reduced,

then the amount of DA in those cells should also be reduced. In

addition, Trulson and Ulissey (1987) have reported that the synthetic

capability of DA cells is reduced to the same extent as tyrosine

hydroxylase activity. In this case, one would also predict that the

amount of DA should also be reduced, and in the discussion of the

paper in 1987, Trulson and Ulissey mention the fact that there was a

TABLE 5. Effects of repeated daily administration of cocaine

(100 mg/kg/day x 10 days) on regional levels of

monoamines and metabolites

Cocaine

Group Cortex

Nucleus

Hippocampus Hypothalamus Striatum Accumbens

5-HT

24 hours

2 weeks

5-HIAA

24 hours

2 weeks

DA

24 hours

2 weeks

N D

ND

DOPAC

24 hours

2 weeks

HVA

24 hours

2 weeks

108.7±11.3

98.1± 13.7

120.4±12.4

128.6±9.0

NDb

ND

N D

N D

N D

ND

ND

ND

88.0±9.4

100.0±4.9

116.0±6.5

107.4±7.0

ND

ND

ND

ND

84.0±7.5

92.7±6.9

100.3±7.7

89.2±4.4

101.4±5.2

91.0±23.1

ND

ND

80.7±3.9 97.5±13.3

90.9±10.5 93.5±12.7

91.7±6.4

86.1±4.0

109.0±6.5

119.8±4.8

117.7±14.2

102.1±6.7

135.6±12.9

113.4±5.7

139.5±35.3

83.9±13.5

81.7±11.6

89.1 ±8.6

122.2±15.3

82.5±6.4

115.2±20.6

95.2±18.4

KEY: ND=not determined.

NOTE: Measures represent mean ± SE, percent of control (n=6/group).

284
reduction in DA levels. However, our results show no reduction in

DA levels following several dosing regimes.

The similarities between the phenethylamine psychomotor stimulants

and cocaine in terms of their effects on behavior and their abuse

liability would allow one to predict that cocaine, MA, and AMPH may

have similar profiles with respect to neurotoxic effects. However,

the psychomotor stimulant methylphenidate is also similar in its

neurobehavioral actions to MA and AMPH, but does not cause neurotoxicity to either dopaminergic or serotonergic cells (Wagner et al.

1980a). The reasons for the differences in neurotoxic effects of such

drugs with similar behavioral effects may be due to differences in

their biochemical pharmacology.

Due to the similarities in the behavioral pharmacology but differences

in neurotoxicity, it becomes relevant to compare and contrast the

effects of AMPH, MA, methylphenidate, and cocaine with regard to

their neurochemical mode of action. First, both MA and AMPH cause

release of DA from a cytoplasmically bound pool, but do not release

from the granular pool (Rateri et al. 1979). MDA and MDMA similarly cause release from the cytoplasmically bound pool of 5-HT and

are toxic to 5-HT cells. In contrast, methylphenidate and cocaine are

not potent releasers of cytoplasmically bound DA or 5-HT, and insofar as these compounds release DA, they appear to do so from the

granular bound pool (Scheel-Kruger 1977).

Evidence suggests that DA release from the granular pool is a

necessary factor in toxicity to DA neurons. We have observed that

alpha-methyl-para-tyrosine (AMT) blocks toxicity engendered by MA

(Wagner et al. 1983; Commins and Seiden 1986). It also blocks

behavioral effects of amphetamine, and these actions can be

attributed to diminishing the cytoplasmic pool, thereby reducing the

releasable amine. Reserpine, which depletes granular bound DA,

enhances the toxic effects of MA (Wagner et al. 1983) by shifting the

equilibrium between the granular pool in favor of the cytoplasmic

pool. In contrast to AMPH, methylphenidate’s behavioral effects are

blocked by reserpine and are not affected by AMT (Scheel-Kruger et

al. 1977), providing evidence that methylphenidate causes release from

the granular pool. Similar to methylphenidate, the locomotor

stimulant effects of cocaine are not blocked by AMT (Wilson and

Holbrook 1979), but are blocked by reserpine (Scheel-Kruger et al.

1972; Scheel-Kruger et al. 1977). Thus, there may be two classes of

stimulants: those that utilize the granular pool and those that use

the cytoplasmic pool of transmitter.

285
MA blocks reuptake of DA as do both methylphenidate and cocaine

(Heikkila 1975; Koe 1976; Randrup and Braestrup 1977; Taylor and Ho

1978; Church et al. 1987). A salient feature of these neurochemical

properties of MA, methylphenidate, and cocaine is the fact that the

drugs that show neurotoxic effects have strong releasing actions from

the cytoplasmically bound pool, and those that seem to be nontoxic

release from the granular pool. This may also explain why methylphenidate is not toxic, because ascorbic acid, which can prevent

neurotoxicity, is an antioxidant released with DA from the granule

and may protect against the oxidation of DA to 6-hydroxydopamine

(6-OHDA). Both ascorbic acid (Wagner et al. 1986) and the antioxidant cysteine (Steranka and Rhind 1987) protect cells from MA

toxicity.

The importance of release from different storage pools can be viewed

in the context of the properties of the drugs that cause neurotoxicity

and the way in which this neurotoxicity can be prevented. According

to recent theory (Seiden and Vosmer 1984), by causing release of DA

from the cytoplasmically bound pool, MA frees DA to become a substrate for oxidation. Among the oxidation products of DA are trihydroxyphenethylamine derivatives (one of which is GABA), which,

through a further oxidation step, can be converted to quinones; the

quinone can crosslink proteins, thus causing neurotoxicity within the

cell. The fact that 6-OHDA has been detected shortly after the

administration of high doses of MA supports this theory (Seiden and

Vosmer 1984). The granular pool contains equal molar amounts of DA

and ascorbic acid; therefore, release of DA from granules will cause

the simultaneous release of ascorbic acid, which protects DA from

being oxidized to form 6-OHDA. Since cocaine mainly blocks the

reuptake of DA and releases weakly, but only from the granular pool

, Koe 1976; Heikkila et al. 1975; Bagchi and Reilly 1983), it would not

be expected to be toxic to DA cells. Similarly, methylphenidate

releases from the granular pools and is not toxic to dopaminergic

cells. Cocaine, like methylphenidate and unlike MA, is not toxic to

DA neurons. Both data and theory are consistent with this point.

However, it should be noted that the lack of long-term DA depletions

does not preclude the possibility that other forms of neurotoxicity

might result from repeated cocaine exposure.

REFERENCES

Bagchi, S.P., and Reilly, M.A. lntraneuronal dopaminergic action of

cocaine and some of its metabolites and analogs.

Neuropharmacology 22:1289-1295, 1983.

286
Church, W.H.; Justice, J.B., Jr.; and Byrd, L.D. Extracellular dopamine

in rat striatum following uptake inhibition by cocaine, nomifensine

and benztropine. Eur J Phermacol 139:345-348, 1987.

Commins, D.L., and Seiden, L.S. Alpha-methyltyrosine Mocks

methylamphetamine-induced degeneration in the rat somatosensory

cortex. Brain Res 365:15-20, 1986.

Commins, D.L.; Vosmer, G.; Virus, R.M.; Woolverton, W.L., Schuster,

C.R.; and Seiden, L.S. Biochemical and histological evidence that

methylenedioxy-methyl-amphetamine (MDMA) is toxic to neurons in

the rat brain. J Pharmacol Exp Ther 241:338-345, 1987.

Gale, K. Catecholamine-independent behavioral and neurochemical

effects of cocaine in rats. In: Sharp, C.W., ed. Mechanisms of

Tolerance and Dependence. National Institute on Drug Abuse

Research Monograph 54. DHHS Pub. No. (ADM) 84-1330.

Washington, D.C: Supt. of Docs., U.S. Govt. Print. Off., 1984. pp.

323-332.

Goeders, N.E., and Kuhar, M.J. Chronic cocaine administration

induces opposite changes in dopamine receptors in the striatum and

nucleus accumbens. Alcohol Drug Res 7:207-216, 1987.

Heikkila, R.E.; Oriansky, H.; and Cohen, G. Studies on the distinction

between uptake inhibition and release of dopamine in rat brain

tissue slices. Biochem Pharmacol 24:847-852, 1975.

Koe, B.K. Molecular geometry of inhibitors of the uptake of

catecholamines and serotonin in synaptosomal preparations of rat

brain. J Pharmacol Exp Ther 199:649-661, 1976.

Lorez, H. Fluorescence histochemlstry indicates damage of striatal

dopamine nerve terminals in rats after multiple doses of methamphetamine. Life Sci 28:911-916, 1981.

Morgan, M.E., and Gibb, J.W. Short-term and long-term effects of

methamphetamine on biogenic amine metabolism in extra-striatal

dopaminergic nuclei. Neuropharmacology 19:989-995, 1980.

Raiteri, M.; Cerrito, F.; Cervoni, A.M.; and Levi, G. Dopamine can be

released by two mechanisms differentially affected by the dopamine

transport inhibitor nomifensine. J Pharmacol Exp Ther 208:195-202,

1979.

Randrup, A., and Braestrup, C. Update inhibition of biogenic amines

by newer antidepressant drugs: Relevance to the dopamine

hypothesis of depression. Psychophamracology (Berlin) 53:309-3 14,

1977.

Ricaurte, G.A.; Bryan, G.; Strauss, L.; Seiden, L.S.; and Schuster, C.R.

Hallucinogenic amphetamine selectively destroys brain serotonin

nerve terminals. Science 229:986-988, 1985.

287
Ricaurte, G.A.; Guillery, R.W.; Seiden, L.S.; Schuster, C.R.; and

Moore, R.Y. Dopamine nerve terminal degeneration produced by

high doses of methylamphetamine in the rat brain. Brain Res

235:93-103, 1982.

Ricaurte, G.A.; Schuster, C.R.; and Seiden, L.S. Long-term effects of

repeated methylamphetamine administration on dopamine and

serotonin neurons in the rat brain: A regional study. Brain Res

193:153-163, 1980.

Ricaurte, G.A.; Seiden, L.S.; and Schuster, C.R. Increased dopamine

metabolism in the rat neostriatum after toxic doses of d-methylamphetamine. Neuropharmacology 22:1383-1388, 1983.

Ricaurte, G.A.; Seiden, L.S.; and Schuster, C.R. Further evidence that

amphetamines produce long-lasting dopamine neurochemical deficits

by destroying dopamine nerve fibers. Brain Res 303:359-364, 1984.

Roy, S.N.; Bhattacharyya, A.K.; Pradhan, S.; and Pradhan, S.N.

Behavioural and neurochemical effects of repeated administration of

cocaine in rats. Neuropharmacology 17:559-564, 1978.

Scheel-Kruger, J. Comparative studies of various amphetamine

analogues demonstrating different interactions with the metabolism

of the catecholamines in the brain. Eur J Pharmacol 14:47-59,

1971.

Scheel-Kruger, J. Behavioral and biochemical comparison of

amphetamine derivatives, cocaine, benztropine, and tricyclic

antidepressant drugs. Eur J Pharmacol 18:63-73, 1972.

Scheel-Kruger, J.; Braestrup, C.; Neilson, M.; Golembiowska, K.; and

Mogilnicka, E. Cocaine: Discussion on the role of dopamine in the

biochemical mechanism of action. In: Ellinwood, E.H., Jr., and

Kilbey, M.M., eds. Cocaine and Other Stimulants. New York:

Plenum Press, 1977. pp. 373-407.

Schmidt, C.J. Neurotoxicity of the psychedelic amphetamine,

methylenedioxymethamphetamine. J Pharmacol Exp Ther 240:1-7,

1987.

Schmidt, C.J.; Wu, L.; and Lovenberg, W. Methylenedioxymethamphetamine: A potentially neurotoxic amphetamine analogue.

Eur J Pharmacol 124:175-178, 1986.

Seiden, L.S.; Fischman, M.W.; and Schuster, C.R. Long-term

methamphetamine induced changes in brain catecholamines in

tolerant rhesus monkeys. Drug Alcohol Depend 1:215-219,1975.

Seiden, L.S., and Vosmer, G. Formation of 6-hydroxydopamine in

caudate nucleus of the rat brain after a single large dose of

methylamphetamine. Pharmacol Biochem Behav 21:29-31, 1984.

Snyder, S.H. Amphetamine psychosis: A “model” schizophrenia

mediated by catecholamines. Am J Psychiatry 130:61-67, 1973.

288
Wilson, M.C., and Holbrook, J.M. Actometric effects of intravenous

cocaine in rats. Arch Int Pharmacodyn Ther 238:244-256, 1979.

AUTHORS

Lewis S. Seiden, Ph.D.

Mark S. Kleven, Ph.D.

Departments of Pharmacological and

Physiological Sciences and Psychiatry

The University of Chicago

947 East 58th Street

Chicago, IL 60637

289
Developmental Effects of Cocaine

Diana L. Dow-Edwards

CLINICAL STUDIES

With the increase in cocaine use among the general population in

recent years (Adams and Durell1984), adverse effects of the drug

have occurred in all social strata and age groups, including the

pregnant population. For example, approximately 300 to 400 babies

are born at the Kings County Hospital (Brooklyn) each year from

women admitting to cocaine use (Glass, personal communication,

1987). In 1985, the first well-controlled clinical study appeared,

describing the increased incidence of abruptio placentae in cocaine

users and neurobehavioral abnormalities in their offspring (Chasnoff

et al. 1985). These exposed infants appeared jittery and exhibited

decreased interactive behavior and poor organizational responses.

Chasnoff et al. (1986) soon reported a case of an infant whose

mother used 5 g of cocaine just prior to delivery and who presented

with multiple focal seizures, cerebral infarction, tachycardia, and

hypertension. Cocaine, or the metabolite benzoylecgonine, was

present in the urine of this infant for 4 days. A significant decrease

in birth weight and head circumference was found to be associated

with cocaine use during pregnancy in a sample of 70 exposed infants

and an equal number of matched controls (MacGregor et al. 1987).

Others have also reported that cocaine exposure is associated with

intrauterine growth retardation, increased stillbirth, and skull defects

(Bingol et al. 1987). The first report (LeBlanc et al. 1987) on the

effects of intrauterine exposure to the alkaloidal form of cocaine, or

“crack,” states that exposed infants show abnormal tremulousness,

irritability, and muscular rigidity, as well as a decrease in gestation

length and birth weight. The clinical literature, therefore, points to

neurobehavioral and growth effects on the infant following intrauterine exposure to cocaine.

290
TERATOLOGIC STUDIES

Even prior to the first clinical report on the effects of cocaine abuse

during pregnancy, the teratology of the compound had been described

by two independent groups. When mice were examined on gestation

day 18 (1 day prior to delivery), Mahalik et al. (1980) found that a

single exposure to cocaine at 60 mg/kg SC between days G6 and G12

caused soft tissue and skeletal abnormalities, increased resorptions,

and altered the sex ratio. However, these single doses had no effect

on maternal weight gain or mean fetal weights. Fantel and Macphail

(1982), however, found that repeated cocaine doses of 60 mg/kg IP

between days 7 and 16 of gestation in the mouse significantly reduced

fetal weights, though, in general, the fetuses appeared to be normal.

This same dose, when given daily to pregnant rats between days 8

and 12 of gestation, also caused a decrease in maternal and fetal

weights, an increase in the number of resorptions, and the presence

of edema in 13 fetuses of 9 litters examined. Higher doses (75

mg/kg) were lethal; lower doses (50 mg/kg) increased the resorption

rate. Since the animals were anesthetized with ether prior to the

daily injection, the results of this study were questionable. Recently,

a fairly complete dose-response curve for subcutaneous administration

of cocaine during pregnancy has been established for the rat.

Between 40 and 90 mg/kg of cocaine produced decreases in maternal

food and water consumption and maternal weight gain in a doss

response manner (Church et al. 1988). Fetal weights were, however,

only affected at the highest dose. This relative sparing of the

fetuses is perhaps due to the increased fetal fatality observed at all

doses of the drug, thereby providing a greater share of the nutrients

to the survivors. Fetal edema, abruptio placentae, and cephalic

hemorrhage were also identified in the fetuses. Together, the animal

data indicate that these consequences-also identified in the clinical

population-are effects of cocaine exposure per se and are not merely

the result of polydrug interactions, which are so difficult to control

in humans.

Spear et al. (1987) have now examined the neurobehavioral development of rats exposed to 40 mg/kg cocaine SC between gestational

days 8 and 20. The 40 mg/kg dose had no effect on reflex ontogeny.

However, the treated pups showed decreased learning and memory in

a test using appetitive conditioning measures as well as a decrease in

wall climbing and an increase in locomotor activity following mild

foot shock. This dose was found to have minimal effects on the

physical development of the fetus as a whole, and no effect on

number and weight of live offspring or on gestation length.

291
Apparently, prenatal cocaine exposure at a dose too small to produce

obvious malformations nevertheless does induce abnormal behavioral

development, which persists at least until the time of weaning.

BRAIN FUNCTIONAL STUDIES

The effects of cocaine on synaptic development of the brain have

been investigated in the rat for the last several years by our

laboratory. We have found that cocaine has potent and long-lasting

effects on the function of several important cerebral pathways (DowEdwards et al. 1986; Dow-Edwards et al. 1988). Our approach has

been to administer the drug during the early postnatal period of the

rat, when the synapses in the forebrain regions are forming. This is

a time approximately equivalent to the third trimester in utero for

humans. This period is characterized by rapid axonal and dendritic

expansion, and is highly sensitive to environmental influences

(Dobbing 1968), particularly from agents interacting with the synapse,

such as cocaine. As discussed by Balster (this volume), cocaine acts

to increase activity at dopamine, norepinephrine, and serotonin

synapses. These transmitter substances are important in the

functioning of the forebrain regions associated with the reinforcing

properties of cocaine (Koob and Hubner, this volume), and are

undergoing synaptogenesis during the early postnatal period (Levitt

and Moore 1979; Hartley and Seeman 1983; Fillion and Bauguen 1984;

Moon-Edley and Herkenham 1984; Murrin et al. 1985). Manipulation

of these transmitter substances with agonists or antagonists during

critical periods of development can result in long-term alterations in

the function of these regions and their associated pathways (Loizou

1972; Coyle and Henry 1973; Rosengarten and Friedhoff 1979;

Friedhoff and Miller 1983). Since reuptake of the catecholamines and

its inhibition (the presumed action of cocaine on the synapse) are

occurring during the early postnatal period (Coyle and Axelrod 1971;

Coyle and Campochiaro 1976; Deskin et al. 1981), the mechanisms by

which cocaine exerts its effect at the synapse may be operating in

the neonate in the same manner as in the adult, but perhaps at a

lower level. Therefore, as with other agonists and antagonists,

exposure to cocaine during this critical period may alter the function

of these synapses both immediately and permanently. The results of

our experiments support this idea.

In the first study, we administered 50 mg/kg cocaine HCI to half of

the pups in a litter and the vehicle (water) to the other half.

During the first 3 days of life, the dose was divided into two

portions and administered 8 hours apart. On days 4 through 10, the

292
injections were administered only once a day in the morning. The

litters were culled to eight and were weighed frequently. Reflex

ontogeny was assessed on a small sample of pups throughout this

early period and was found not to be affected (table 1). Growth of

the pups before and after weaning (day 21) was also not affected

(figure 1). At 60 days of age, there were no statistically significant

TABLE 1. Reflex ontogeny of control and cocaine-treated rat

pups (mean day of appearance of reflex)

Tail Hang

Bar Grasp

Creeping

Walking

Eye Opening

Control+ Neonatal Cocaine

(8) (8)

8.75±0.2 7.75±0.5

12.9±0.5 12.7±0.5

8.9±0.3 8.5±0.5

13.2±0.5 13.2±0.2

14.6±0.3 14.5±0.3

Significant at p<0.05 compared to the control.

+Controls include four untreated and four vehicle-treated rat pups.

differences between groups in mean arterial blood pressure, arterial

blood gases or pH, hematocrit, or plasma glucose (table 2). However,

the female treated rats showed a significantly different pattern from

the untreated females in brain functional activity as measured by

glucose metabolism (table 3). Brain glucose metabolism was determined using the deoxyglucose method (Sokoloff et al. 1977) as

previously described (Vingan et al. 1986). The female rats were all

examined during diestrus to minimize the differences in brain

metabolism that occur across days of the estrus cycle (Nehlig et al.

1985). Using the fully quantified method, we found that 40 percent

of the limbic structures examined were metabolically stimulated in the

treated adult females compared with the female controls. The motor

and sensory systems were less significantly affected (table 3).

293
FIGURE 1. Growth curves of both male (top) and female (bottom)

treated and control Sprague-Dawley rats

NOTE: The treatment consisted of 50 mg/kg cocaine SC between days 1 and 10 of

life. Each point represents approximately eight observations. There were no

statistically different comparisons between treated and control values.

294
TABLE 2. Physiologic parameters of rats treated postnatally

with cocaine

Factor

Female

Water Cocaine

n=6 n=6

Male

Water Cocaine

n=6 n=7

Weight (g)

Glucose (mg/ml)

Temperature (°C)

Pressure (mmHg)

Hematocrit (%)

Gases

O2 (mmHg)

CO2 (mmHg)

pH

Estrus Cycle

197±3

1.22±.02

37.9±.18

127±3

46.6±2.1

89.8±11.5 94.3±6.6

24.6±2.1 23.4±2.9

7.36±.04 7.38±.05

Diestrus Diestrus

204±7

1.22±.07

37.7±.07

121±2

49.2±1.4

347±16

1.19±.05

37.4±.08

120±3

45.7±1.5

85±5.3

29±2.1

7.41±.05

343±13

1.20±.06

37.4±.13

117±2

47.3±0.9

86.3±5.9

30.9±2.9

7.40±.05

NOTE: Values are means ± SEM for the number of animals indicated.

Most of the regions examined in the female rats were 10 to 15

percent more active in the treated animals. Every structure found to

be significantly altered in this study was also significantly altered by

acute cocaine exposure in the adult (Porrino et al. 1988; Porrino and

Kornetsky, this volume; London et al. 1986). This is in sharp

contrast to the findings with the male rats. Here, neonatal cocaine

treatment had little effect on brain glucose metabolism. Table 4

shows a sample of the 43 brain regions evaluated in the male rats.

As can be seen, with the exception of the auditory cortex, none of

the regions showed a greater than 6 percent difference between the

treated and the control groups. Therefore, one might conclude that

cocaine administration at 50 mg/kg during the first postnatal week

has no significant effect on brain glucose metabolism in male rats,

but that it has a large stimulatory effect in females.

Other workers in the field have described similar sex-related

differences in response to cocaine. Glick and coworkers (Glick et al.

1983; Glick et al. 1984) reported that cocaine induces a greater

amount of turning in females than in males. Behavioral sensitization

295
TABLE 3. Brain glucose metabolism in control and cocaine-treated

female fats

Control Cocaine-Treated

(n=6) (n=6)

Motor Structures

Motor cx

Caudate n

Globus pallidus

Thalamus

Ventral n

Substantia nigra

Red n

Pontine nuclei

Pontine reticular

formation

Vestibular n

Cerebellar cx

Corpus callosum

Limbic Structures

Mesolimbic forebrain

Accumbens

Bed n stria terminalis

Horizontal limbdiagonal

Septum

Medial

Lateral

Limbic Cx

Cingulate cx

Piriform cx

Insular cx

Hypothalamus

Medial preoptic n

Mammillary body

Hippocampus

CA1

Dentate gyrus

69±4

78±4

45±2

67±3

44±2

59±3

46±2

48±3

96±4

38±4

36±2

74±1

34±1

77±3

64±2

47±1

86±2

84±5

112±6

35±2

95±4

61±3

43±3

79±3

89±2*

47±1

80±2*

48±1

68±2*

51±1*

54±3

108±5

41±1

38±2

84±5

36±1

89±3*

70±4

48±1

102±4*

91±4

125±6

36±2

108±5

70±2*

47±1

296
TABLE 3. (Continued)

Control

(n=6)

Cocaine-Treated

(n=6)

Limbic Structures (Continued)

Amygdala

Lateral n

Central n

Cortical n

Habenula

Medial n

Lateral n

Thalamus

Mediodorsal n

Ventral tegmental area

lntetpeduncular n

Medial raphe n

Central gray

64±2

33±2

44±1

58±3

97±2

67±3

109±5*

88±4

51±2

83±5

82±3

51±3

Sensory Structures

Sensory cx

Head

Vibrissa

Association (parletal) cx

74±4

76±4

69±3

Primary olfactory cx 94±4

Occipital cx

Lateral geniculate n

Superior colliculus

77±4

69±3

63±3

Auditory cx 113±11

Medial geniculate n 103±8

Inferior colliculus 129±11

70±1*

37±1

48±2

106±6*

59±1*

100±5*

99±7*

57±1

83±2

83±4

76±3

106±3*

86±4

78±2*

73±5

130±6

123±4*

143±5

*Significant at p<0.05 from control by t-test.

NOTE: Measures in μMol/100 g tissue/min; mean ± SEM for the numbers of animals

in parentheses.

297
TABLE 4. Brain glucose metabolism in control and cocainetreated male rats

Sensory Structures

Sensory cx

Head

Vibrissa

Association (parietal) cx

Control Cocaine-Treated

(6) (6)

85±4 86±2

86±3 87±3

80±3 83±3

Primary olfactory cx

Occipital cx

Lateral geniculate n

Superior colliculus

117±7 116±5

96±9 91±5

81±2 83±3

79±5 76±3

Auditory cx

Medial geniculate n

Inferior colliculus

148±7 126±5*

124±3 125±3

153±8 152±1

*Significant value p< 0.05 from control by t-test.

NOTE: Measures in μMol/100 g tissue/min; mean ± SEM for the numbers of animals

in parentheses.

to cocaine is also more prevalent and longer lasting in females than

in males (Glick et al. 1983; Glick and Hinds 1984; Post 1981).

Therefore, it is not surprising that greater alterations in brain

function were seen in our experiment in the females exposed to

cocaine during synaptogenesis than in the males.

In the next experiment, we wanted to determine whether these

functional changes in the female brain were associated with altered

behavior in the adult. Therefore, we prepared another 60 females by

injecting half with cocaine at 50 mg/kg and the other half with the

vehicle (water), following the same injection schedule (postnatal days

1 through 10) and treatments as we had previously. However, this

time only the females were studied. Again, at 60 to 65 days of age,

the rats (all in the same phase of the estrus cycle) were examined

for baseline activity for 15 minutes in a Digiscan Activity Monitor.

298
This monitor has eight photosensors on each side, as well as a second

row of sensors placed 16 cm from the floor of the chamber. lt is

enclosed in a sound-insulating wood box with two 10-watt lights and

has a Par ventilator fan. A window in the center of the top of the

box allows the experimenter to observe the animals. Between 10

a.m. and noon, each animal was taken directly from the home cage

and placed in the monitor. The behaviors occurring during the 15

1-minute intervals were automatically recorded and grouped into

three 5-minute intervals.

As shown in figure 2, neonatal cocaine exposure in female rats

increased several types of behavior. Distance traveled per minute

(during the second two 5-minute periods), the number of vertical

movements per minute (during all time intervals), and the time spent

(in seconds per minute) in the center of the monitor (during the first

and last 5-minute periods) were all significantly greater in the

treated animals than in the controls (by t-test analyses). Although

all of these data have yet to be analyzed by repeated measures

analysis of variance, they do suggest that neonatal cocaine treatment

increases general activity levels. However, the increase in time spent

in the center of the monitor may be related to the presence of the

viewing window in the top of the sound-insulating box; it does not

necessarily suggest that the cocaine induced some sort of wall-phobia

in these animals.

We have also examined the behavioral responses to amphetamine

administration in the neonatally treated female rats, and have found

that cocaine appears to increase the sensitivity to amphetamine.

Although we have not yet fully analyzed these data, the treated rats

appear to show an increase in stereotypic behaviors and a decrease in

vertical activity following 1 and 5 mg/kg d-amphetamine sulfate IP

30 minutes prior to the behavioral recording. Previous studies have

shown that other developmental insults alter responsivity to amphetamine (Silbergeld and Goldberg 1974), and a body of clinical

literature indicates that hyperactive children show a paradoxical

response to the drug. Our present findings suggest that perinatal

exposure to cocaine induces hyperactivity and results in abnormal

behavioral responses to amphetamine.

CONCLUSION

Exposure to cocaine during a critical period of development has been

shown to produce lasting changes in the function of the brain. lf

the drug is given during the first 10 days of postnatal life in the rat,

299
FlGURE 2. Baseline activity collected with a Digiscan Activity

Monitor with vertical sensors 16 cm from the base of

the floor

NOTE: Values ± SD represent data obtained from 60 adult female rats receiving

vehide or 50 mg/kg cocaine between days 1 and 10 of life: data averaged

over 5 minutes.

*Significant difference from control by t-test.

is control and O Is treated. All animals were in estrus at the time of the

behavioral observation.

the animals are hyperactive as adults. Hyperactivity in humans is

well known to interfere with performance in school. Therefore,

although low to moderate exposure to cocaine during pregnancy does

not appear to induce obvious structural abnormalities in the offspring,

our data suggest that the drug may place exposed children at risk for

neurobehavioral abnormalities that may last into adulthood.

300
REFERENCES

Adams, E.H., and Durell, D. Cocaine: A growing public health

problem. In: Grabowski, J., ed. Cocaine: Pharmacology, Effects

and Treatment of Abuse. National Institute on Drug Abuse

Research Monograph 50. DHHS Pub. No. (ADM)84-1328.

Washington, DC: Supt. of Docs., U.S. Govt. Print. Off., 1984.

pp. 9-14.

Bingol, H.; Fuchs, M.; Diaz, V.; Stone, R.K.; and Gromish, D.S.

Teratogenicity of cocaine in humans. J Pediatr 110:93-98, 1987.

Chasnoff, I.J.; Burns, W.J.; Scholl, S.H.; and Burns, K.A. Cocaine use

in pregnancy. New Engl J Med 313:666-669, 1985.

Chasnoff, I.J.; Bussey, M.E.; Savich, R.; and Stack, C.M. Perinatal

cerebral infarction and maternal cocaine use. J Pediatr 108:456-459,

1986.

Church, M.W.; Dintcheff, B.A.; and Gessner, P.K. Dose-dependent

consequences of cocaine on pregnancy outcome in the Long-Evans

rat. Neurotox and Teratol 10:51-58, 1988.

Coyle, J.T., and Axelrod, J. Dopoamine- b-hydroxylase in the rat

brain: Developmental characteristics. J Neurochem 19:449-459,

1972.

Coyle, J.T., and Campochiaro, P. Ontogenesis of dopaminergiccholinergic interactions in rat striatum: A neurochemical study. J

Neurochem 27:673-678, 1978.

Coyle, J.T., and Henry, D. Catecholamines in fetal and newborn rat

brain. J Neurochem 21:61-67, 1973.

Deskin, R.; Seidler, F.J.; Whitmore, W.L.; and Slotkin, T.A. Development of a-noradrenergic and dopaminergic receptor systems

depends on maturation of their presynaptic nerve terminals in the

rat brain. J Neurochem 36(5):1683-1690, 1981.

Dobbing, J. Vulnerable periods in developing brain. In: Davison, A.,

and Dobbing, J., eds. Applied Neurochemistry. Philadelphia: Davis

Company, 1988. pp. 287-316.

Dow-Edwards, D.L.; Freed, LA.: and Milhorat, T.H. The effects of

cocaine on development. Abstr Soc Neurosci 12:59.12, 1986.

Dow-Edwards, D.L.; Freed, LA.; and Milhorat, T.H. Stimulation of

brain metabolism by perinatal cocaine exposure. Dev Brain Res

42:137-142, 1988.

Fantel, A.G., and Macphail, T. The teratogenicity of cocaine.

Teratology 26:17-19, 1982.

Fillion, G., and Bauguen, C. Postnatal development of 3H-5-HT

binding in the presence of GTP in rat brain cortex. Dev Pharmacol

Ther 7(Suppl):1-5, 1984.

301
Friedhoff, A.J., and Miller, J.C. Prenatal psychotrophic drug exposure

and the development of central dopaminergic and cholinergic

neurotransmitter systems. Monogr Neural Sci 9:91-98, 1983.

Glick, S.D., and Hinds, P.A. Sex differences in sensitization to

cocaine-induced rotation. Eur J Pharmacol 99:119-123, 1984.

Glick, S.D.; Hinds, P.A.; and Shapiro, R.M. Cocaine-induced rotation:

Sex-dependent differences between left- and right-sided rats.

Science 221:775-777, 1983.

Harden, T.K; Wolfe, B.B.; Sporn, J.R.; Perkins, J.P.; and Molinoff, P.B.

Ontogeny of -adrenergic receptors in rat cerebral cortex. Brain

Res 125:99-108, 1977.

Hartley, E.J., and Seeman, P. Development of receptors for dopamine

and noradrenaline in rat brain. Eur J Pharmacol 91:391-397, 1983.

LeBlanc, P.E.; Parekh, A.J.; Naso, B.; and Glass, L. Effects of

intrauterine exposure to alkaloidal cocaine (“crack”). Am J Dis

Child 141:937-938, 1987.

Levitt, P., and Moore, R.Y. Development of the noradrenergic

innervation of neocortex. Brain Res 162:243-259, 1979.

Loizou, A.L. The postnatal ontogeny of monoamine-containing

neurons in the CNS of albino rats. Brain Res 40:395-418, 1972.

London, E.D.; Wilkerson, G.; Goldberg, S.R.; and Risner, M.E. Effects

of L-cocaine on local cerebral glucose utilization in the rat.

Neurosci Lett 68:73-78, 1986.

MacGregor, S.N.; Keith, L.G.; Chasnoff, I.J.; Rosner, M.A.; Chisum,

G.M.; Shaw, P.; and Minogue, J.P. Cocaine use during pregnancy:

Adverse perinatal outcome. Am J Obstet Gynecol 157:686-690, 1987.

Mahalik, M.P.; Gautieri, R.F.; and Mann, D.E. Teratogenic potential

of cocaine hydrochloride in CF-1 mice. J Pharm Sci 69:703-709,

1980.

Moon-Edley, S., and Herkenham, M. Comparative development of

striatal opiate receptors and dopamine revealed by autoradiography

and histofluorescence. Brain Res 305:27-42, 1984.

Murrin, L.C.; Gibbens, D.L.; and Ferrer, J.R. Ontogeny of dopamine,

serotonin and spirodecanone receptors in rat forebrain-an

autoradiographic study. Dev Brain Res 23:91-109, 1985.

Nehlig, A.; Porrino, L.; Crane, A.; and Sokoloff, L. Local cerebral

glucose utilization in normal female rats: Variations during the

estrus cycle and comparison with males. J Cereb Blood Flow Metab

5:393-400, 1985.

Nomura, Y.; Naitoh, F.; and Segawa, T. Regional changes in

monoamine content and uptake of the rat brain during postnatal

development. Brain Res 101:305-315, 1976.

302
Porrino, L.J.; Domer, F.R.; Crane, A.M.; and Sokoloff, L. Selective

alterations in cerebral metabolism within the mesocortical dopaminergic system produced by acute cocaine administration in rats.

Neuropsychopharm 1:109-118, 1988.

Post, R.M. Central stimulants: Clinical and experimental evidence on

tolerance and sensitization. In: Israel, Y.; Glaser, F.; Kalant, H.;

Popham, R.E.; Schmidt, W.; and Smart, R., eds. Research Advances

in Alcohol and Drug Problems. New York: Plenum Press, 1981.

pp. 1-65.

Rosengarten, H., and Friedhoff, A.J. Enduring changes in dopamine

receptor cells of pups from drug administration to pregnant and

nursing rats. Science 203:1133-1135, 1979.

Silbergeld, E.K., and Goldberg, A.M. Lead-induced behavioral

dysfunction: An animal model of hyperactivity. Exp Neural 42:146-

157, 1974.

Sokoloff, L.; Reivich, M.; Kennedy, D.; Des Rosiers, M.H.; Patlak, C.S.;

Pettigrew, KD.; Sakurada, O.; and Shinohara, M. The [

14C]

deoxyglucose method for measurement of local cerebral glucose

utilization: Theory, procedure and normal values in the conscious

and anesthetized albino rat. J Neurochem 28:897-916, 1977.

Spear, LP.; Kirstein, C.; Bell, J.; Greenbaum, R.; O’Shea, J.;

Yoottanasumpun, V.; Hoffman, H.; and Spear, N.E. Effects of

prenatal cocaine on behavior during the early postnatal period in

rats. Teratology 35:BTS12, 1987.

Vingan, R.D.; Dow-Edwards, D.L; and Riley, E.P. Effects of prenatal

exposure to ethanol on local cerebral glucose utilization.

Alcoholism: Clin Exp Res 10:22-26, 1986.

ACKNOWLEDGMENTS

These studies were funded in part by National Institute on Drug

Abuse grant DA 04118, National Institute of Neurological and

Communicative Disorders and Stroke grant NS22766, and the State

University of New York. The financial contribution of T.H. Milhorat

is acknowledged. L.A. Freed and R.J. Wolintz are acknowledged for

their assistance in data analysis and in other technical aspects of

these studies.

AUTHOR

Diana L. Dow-Edwards, Ph.D.

State University of New York

Health Science Center at Brooklyn

Brooklyn, NY 11203

303
Cardiovascular Toxicity of Cocaine

R. Douglas Wilkerson

Although cocaine has been utilized for many years as a local anesthetic in the practice of medicine (Ritchie and Greene 1985) and in

the research laboratory as a tool to block the neuronal uptake of

catecholamines (Trendelenburg 1959), the cardiovascular actions of

cocaine itself have not been widely studied. Deaths associated with

intravenous cocaine abuse, however, have been reported over the last

10 years, and the circumstances surrounding these deaths were clearly

consistent with sudden cardiac death (Lundberg et al. 1977; Di Maio

and Garriott 1978; Kossowsky and Lyon 1984; Nanji and Filipenko

1984). Recently, Mittleman and Wetli (1987) restudied autopsy

material from 24 patients who had apparently suffered sudden

“natural” deaths and found that cocaine was present in the blood of

11 of the 24 patients. Cerebrovascular complications have also been

reported after cocaine use, and these reports have recently been the

subject of a review by Levine and Welch (1987).

The vast majority of recent reports of cocaine-associated cardiovascular toxicity have implicated cocaine in episodes of myocardial

ischemia, which frequently resulted in myocardial infarction. These

reports have been the subject of recent reviews by Mathias (1988)

and Duke (1988). In addition, Smith and coworkers (1987) recently

reviewed the pertinent data from all 38 patients reported to have

suffered myocardial ischemic episodes temporally associated with

cocaine use. A noteworthy finding was that a significant number of

patients with documented evidence of myocardial infarction were

subsequently shown to have angiographically normal coronary arteries.

This finding of normal coronary arteries subsequent to a documented

ischemic episode, temporally related to cocaine use, has led a number

of investigators to suggest that cocaine may induce coronary vasospasm (Schachne et al. 1984; lsner et al. 1988; Zimmerman et al. 1987)

304
and/or promote coronary thrombosis (Zimmerman et al. 1987; Gardezi

1987; Kirlin et al. 1987). Although there is only circumstantial

evidence of cocaine-induced coronary vasospasm, cocaine has been

shown to cause vasospasm in other vascular beds (Altura et al. 1985).

The cardiovascular actions of cocaine are extremely complex. As a

result of its local anesthetic actions, one might expect vasodilation in

most vascular beds and a reduced tendency for cardiac arrhythmias.

On the other hand, the excitatory effects of cocaine, like other local

anesthetics in the central nervous system, might be expected to

increase peripheral sympathetic tone (Ritchie and Greene 1985). This

central stimulation of sympathetic outflow, combined with cocaine’s

well-established action to inhibit the neuronal uptake of norepinephrine, would be expected to elicit intense vasoconstriction in most

vascular beds, an increase in heart rate and myocardial contractility,

and an increased tendency for cardiac arrhythmias. These factors

would also lead to an increase in myocardial oxygen consumption.

Thus, the two best-understood actions of cocaine (i.e., local anesthesia and inhibition of neuronal uptake of catecholamines) would be

expected to produce opposing effects on the heart and circulation.

The local anesthetic effect on the heart and blood vessels would

result in antiarrhythmic and vasodilatory actions, while local anesthetic actions and effects at adrenergic, dopaminergic, and serotonergic synapses within the central nervous system would result in

excitation leading to seizure activity and increased peripheral

sympathetic tone. The effects of the norepinephrine released by this

increased sympathetic tone to the heart and blood vessels should be

potentiated by the action of cocaine to inhibit the major mechanism

for terminating the action of norepinephrine, that is, neuronal uptake.

Because of this complexity of actions, it is essential that studies

designed to clarify the cardiovascular actions of cocaine be performed

utilizing a model that preserves all aspects of cocaine action. This

can only be accomplished in an intact, conscious animal model, which

allows the integrated central nervous system and cardiovascular

actions of cocaine to be fully expressed without interference from

general anesthesia.

To better understand the contribution of the central nervous system

to the cardiovascular actions of cocaine, we compared the effects of

intravenous cocaine, 1 mg/kg, on blood pressure in conscious dogs

with the effects of the same dose in dogs anesthetized with 32 mg/kg

pentobarbital sodium (figure 1). In anesthetized dogs, this dose of

cocaine produced a 9.5±3.1 mmHg increase in blood pressure, while in

conscious dogs, the same dose of cocaine produced a 45.8±5.2 mmHg

305
elevation in blood pressure (Wilkerson 1988). Thus, it is clear that

depression of the central nervous system significantly reduced the

peripheral vascular actions of cocaine which, presumably, were

mediated, at least in part, by increased peripheral sympathetic tone

resulting from its central stimulant actions.

FIGURE 1. Influence of pentobarbital anesthesia on the blood

pressure response elicited by 1 mg/kg of cocaine

administered intravenously in dogs

NOTE: The bar on the left illustrates the blood pressure response to 1 mg/kg of

cocaine in a group of 10 dogs anesthetized with pentobarbital sodium

(32 mg/kg). The bar on the tight illustrates the blood pressure response to

the same dose of cocaine in a group of 10 trained conscious dogs. Each bar

represents the mean ± standard error for the number of animals indicated.

To confirm the importance of a central nervous system component to

the cardiovascular effects of cocaine, we compared the effects of

cocaine on blood pressure in conscious dogs with its effects in the

same animals, on a different day, after pretreatment with 10 mg/kg

of hexamethonium, a ganglionic blocking drug. This dose of hexamethonium did not significantly alter blood pressure, but did significantly increase heart rate. In addition, hexamethonium abolished

306
the reflex increase in heart rate elicited by intravenous administration of nitroglycerin. It can be seen in figure 2 that this dose of

hexamethonium significantly reduced the pressor response elicited by

the intravenous administration of 2 mg/kg of cocaine from 70±4.2 to

25.3±3.8 mmHg. The fact that hexamethonlum itself at this dose had

very little effect on blood pressure suggests that sympathetic

FIGURE 2. The effect of ganglionic blockade with hexamethonium on

the blood pressure response to cocaine (2 mg/kg, IV) in

trained, conscious dogs

NOTE: The left bar depicts the response of six dogs to this dose of cocaine without

any pretreatment. The right bar depicts the response to this same dose of

cocaine in the same six dogs on a different day, but after pretreatment with

10 mg/kg of hexamethonium. Each bar represents the mean ± standard error

of six dogs.

tone to blood vessels in these conscious dogs was quite low

(Wilkerson 1988). This observation argues against the possibility that

the major action of cocaine to increase blood pressure was merely by

blocking neuronal norepinephrine uptake and, thus, potentiating the

effect of norepinephrine being released as a result of normal sympathetic tone. These findings of a pivotal role of the central

307
nervous system in the cardiovascular actions of cocaine are in accord

with the findings of Matsuzaki and coworkers (1978), who have

demonstrated that EEG changes and behavioral hyperexcitation

accompanied the cardiovascular effects of cocaine in subhuman

primates.

It would be expected that the action of cocaine to inhibit the

neuronal uptake of norepinephrine at peripheral adrenergic synapses

should play a major role in the cardiovascular effects of this drug.

Indeed, cocaine potentiates and prolongs the effects of exogenously

administered norepinephrine (Trendelenburg 1959; Whitby et al. 1960).

Despite this well-documented action to potentiate the effects of

exogenously administered norepinephrine, however, the effects of

cocaine on the response to sympathetic nerve stimulation are still

debated. lt might seem straightforward that, if cocaine inhibits the

neuronal uptake of norepinephrine, then the norepinephrine released

from sympathetic nerves should accumulate in adrenergic synapses and

thus the effects of sympathetic nerve stimulation should be potentiated. Indeed, some studies have reported that cocaine does increase

the effects of sympathetic nerve stimulation (Moore 1966; Langer and

Enero 1974). Yet, other investigators have been unable to observe

any potentiation of the effects of sympathetic nerve stimulation by

cocaine (Koerker and Moran 1971; Matsuda et al. 1979; Levy and

Blattberg 1978). Moreover, Koerker and Moran (1971) demonstrated

that cocaine pretreatment did not increase the amount of norepinephrine in the venous outflow from the heart during sympathetic nerve

stimulation as would be expected if the synaptic concentration of

norepinephrine was greatly increased.

This apparent difference in the effects of cocaine on the actions of

exogenously administered norepinephrine and on the actions of

sympathetic nerve stimulation (i.e., endogenous norepinephrine) is

undoubtedly the result of synaptic control mechanisms, by which

synaptic norepinephrine effectively controls the rate of its own

release by activating presynaptic -adrenergic receptors (Yamaguchi

et al. 1977). Inhibition of neuronal catecholamine uptake may

momentarily result in an increase in synaptic norepinephrine, but its

action on presynaptic -adrenergic receptors would decrease further

release of norepinephrine. This control mechanism is “bypassed” when

norepinephrine reaches the synapse via a process that does not

require its neuronal release (e.g., intravenous injection).

Recently, it has been demonstrated that, even after inhibition of the

neuronal uptake of norepinephrine, the amount of norepinephrine in

308
the synapse is not elevated when sympathetic tone is relatively low

(Cousineau et al. 1986). These same workers also demonstrated,

however, that although this negative feedback control of norepinephrine release was very effective at low to moderate rates of sympathetic nerve stimulation, at high rates of sympathetic nerve

stimulation, this control mechanism was not as effective, and synaptic

norepinephrine levels were increased after neuronal uptake inhibition.

This suggests that the efficacy of the negative feedback control of

norepinephrine release is not sufficient to effectively control synaptic

norepinephrine concentration when sympathetic tone is elevated. lt

is only in this setting that inhibition of the neuronal uptake of

norepinephrine leads to its accumulation in the synapse. Thus, when

inhibition of norepinephrine neuronal uptake can be matched effectively by inhibition of its neuronal release, synaptic norepinephrine

concentration will not increase, but when this is not the case (e.g.,

when sympathetic tone is elevated), synaptic norepinephrine concentration will be increased by inhibition of neuronal uptake.

In addition to the control mechanism described above, Cohen et al.

(1984) have described a second presynaptic mechanism by which the

rate of norepinephrine release may be controlled. These workers

have shown that acetylcholine inhibits the release of norepinephrine

from adrenergic neurons and that this action is blocked by muscarinic

antagonists such as atropine. The involvement of these and perhaps

other synaptic processes acting to control the rate of norepinephrine

release may explain the divergent results of some workers who have

attempted to study the effects of cocaine on the cardiovascular

actions of sympathetic nerve stimulation. Undoubtedly, some of the

confusion also results from the fact that these studies employed

relatively large doses of cocaine (5 mg/kg) and that they were

performed in anesthetized animals.

As shown in figures 1 and 2, the cardiovascular actions of cocaine

clearly have an important component that involves the central

nervous system, which results in an increase in sympathetic outflow

to the periphery. Thus, the cardiovascular actions of cocaine are not

solely dependent upon inhibition of neuronal uptake of norepinephrine

at peripheral adrenergic synapses to provide increased synaptic

concentrations of norepinephrine. In addition to blocking uptake,

cocaine apparently increases sympathetic tone, as evidenced by the

action of hexamethonium to inhibit its pressor response. Chieuh and

Kopin (1978) have also demonstrated that cocaine exerts an action in

the central nervous system to cause the release of norepinephrine and

epinephrine from the sympathoadrenal axis in conscious rats.

309
Furthermore, cocaine appears to cause the release of norepinephrine

from isolated cardiac atrial tissue (Trendelenburg 1968). Thus, the

adrenergic actions of cocaine cannot be thought of as simply the

result of peripheral inhibition of neuronal catecholamine uptake. The

central stimulant component of its cardiovascular effects is probably

what separates cocaine from other drugs, such as the tricyclic

antidepressants, which also very effectively inhibit catecholamine

uptake, but whose cardiovascular actions are not as prominent as

those of cocaine.

We have recently demonstrated the importance of these synaptic

control mechanisms in modulating the cardiovascular actions of

cocaine (Wilkerson 1987). Yohimbine, a selective -adrenergic

antagonist (Goldberg and Robertson 1983), which is widely abused for

its alleged aphrodisiac properties (Linden et al. 1987), was utilized to

assess the influence of presynaptic adrenergic control of norepinephrine release on the actions of cocaine. This drug was employed alone

and in combination with atropine which, as described above, inhibits

cholinergic mechanisms that have also been shown to modulate

norepinephrine release.

Yohimbine, 0.25 mg/kg, increased the effects of cardiac sympathetic

nerve stimulation on heart rate at low stimulation frequencies, but

failed to alter the frequency-response curve at higher frequencies

(figure 3). This finding is in accord with Cousineau and coworkers

(1986), who demonstrated that the presynaptic -adrenergic mechanism for controlling norepinephrine release is most effective at low

stimulation frequencies. In contrast, cocaine, 1 mg/kg, increased the

effects of sympathetic nerve stimulation on heart rate at higher

frequencies, but not at low frequencies (figure 4). This is not

surprising, since, at higher stimulation frequencies, where feedback

control of norepinephrine release is less effective, inhibition of

neuronal uptake of norepinephrine may not be countered by a

corresponding decrease in its neuronal release. This would allow the

synaptic concentration of norepinephrine to rise and the effects of

sympathetic nerve stimulation to be potentiated by cocaine.

Figure 5 illustrates the effects of inhibition of either or both of the

previously described mechanisms controlling neuronal release of

norepinephrine on the blood pressure response to intravenous cocaine,

1 mg/kg, in anesthetized dogs. The effect of cocaine alone at this

310
FlGURE 3. lnflunce of pretreatment with the -adrenergic

antagonist yohimbine on the heart rate response to

cardiac sympathetic nerve stimulation in anesthetized

dogs

NOTE: The left ansa subclavia was electrically stimulated at the frequencies shown on

the abcissa prior to (open bars) and after (hatched bars) treatment with

yohimbine (0.25 mg/kg, IV). Heart rate was electronically derived from the

electrocardiogram. Each bar represents the mean ± standard error of nine

dogs.

dose in an anesthetized animal was a 9.6±2.2 mmHg increase in blood

pressure. When the same dose of cocaine was administered after

pretreatment with yohimbine, 0.25 mg/kg, the blood pressure response

to cocaine was significantly increased to 20.6±2.8 mmHg. After

pretreatment with atropine, 1 mg/kg, the blood pressure response to

the 1 mg/kg dose of cocaine was increased to 17.6±3.5 mmHg.

Pretreatment with both yohimbine and atropine resulted in a 44.2±7.2

mmHg elevation in blood pressure after administration of this same

dose of cocaine.

311
FIGURE 4. Influence of pretreatment with cocaine (1 mg/kg, IV) on

the heart rate response to cardiac sympathetic nerve

stimulation in anesthetized dogs

NOTE: The left ansa subclavia was electrically stimulated at the frequencies shown on

the abdssa prior to (open bars) and after (hatched bars) cocaine administration.

Heart rate was electronically derived from the electrocardiogram. Each bar

represents the mean ± standard error of nine dogs.

In this same study, each animal was also instrumented for the

measurement of myocardial contractile force using a Walton-Brodie

strain gauge arch, which was sutured to the lateral wall of the left

ventricle. The segment of myocardium subtended by the strain gauge

arch was stretched incrementally to the peak of its length-tension

curve by adjusting the distance between the feet of the arch. Thus,

this segment of muscle contracted isometrically and, therefore, its

force of contraction was not influenced by peripheral vascular

alterations, which resulted in changes in ventricular preload or

afterload. In addition, a pulsed doppler blood flow probe was placed

around the left circumflex coronary artery. This probe allowed the

measurement of the velocity of blood flow through its lumen utilizing

the doppler shift principle. Each blood flow probe was precalibrated

312
FIGURE 5. Modification of the blood pressure response elicited by

cocaine (1 mg/kg, IV) in anesthetized dogs by pretreatment with yohimbine (0.25 mg), atropine (1 mg/kg), or

yohimbine plus atropine in these same doses

NOTE: The left bar depicts the blood pressure response to 1 mg/kg of cocaine when

given without any pretreatment, the left-center bar depicts the blood pressure

response to the same dose of cocaine when administered after yohlmbine

pretreatment, the right-center bar depicts the response to the same dose of

cocaine when administered after atropine pretreatment, and the right bar

depicts the blood pressure response to cocaine when administered after

pretreatent with both yohimbine and cocaine. Each bar represents the mean

± standard error of seven dogs.

to allow conversion of the measured doppler frequency shift to blood

flow units (i.e., ml/min).

Coronary blood flow is typically regulated very effectively to match

the heart’s demand for oxygen and/or nutrients. This regulation of

blood flow is the result of adjustments in the caliber of coronary

arterioles (i.e., changes in resistance). Typically, in response to

increased demand for oxygen and/or nutrients, coronary arteries

dilate to increase blood flow to meet that demand. An example of

313
FIGURE 6. Coronary vascular response to increased myocardial oxygen

consumption elicited by administration of 0.25μg/kg of

norepinephrine to anesthetized dogs

NOTE: The left bar depicts the effects of norepinephrine (norepi.) on left ventricular

contractile force (C.F.), the left-center bar depicts the effects of norepinephrine on the product of heart rate and blood pressure (RxP). The right-center

and right bars illustrate the coronary vascular response to this change in

myocardial oxygen demand. The right-center bar illustrates the decrease in

coronary vascular resistance (C.V.R.), which resulted in the increase in coronary blood flow (C.B.F.) shown in the right bar. Each bar represents the mean

± standard error of six dogs.

this phenomenon in the preparation described above is illustrated

in figure 6. When norepinephrine is administered at a dose of

0.25μg/kg, myocardial contractile force and the product of heart rate

and blood pressure are increased, thus increasing myocardial oxygen

consumption. This increase in the demand for oxygen is met by an

increase in coronary blood flow, which is the result of a reduction in

coronary vascular resistance (i.e., coronary vasodilation).

Figure 7 illustrates the effects intravenous cocaine, 1 mg/kg, on myocardial contractile force, the product of heart rate and blood

314
FIGURE 7. Effects of 1 mg/kg of cocaine on left ventricular contractile force (open bars), the product of heart rate and

blood pressure (right-slant hashed bars), and the

coronary vascular response (left-slant hashed bars) to

the change in myocardial oxygen consumption elicited by

these effects of cocaine in anesthetized dogs

NOTE: Cocaine was admlnistered alone and after pretreatment with yohimblne

(0.25 mg/kg), atropine (1 mg/kg), or a combination of yohimbiie and atropine

in these same doses. Each bar represents the mean ± standard error of seven

dogs.

pressure, and coronary vascular resistance in anesthetized dogs

instrumented as described above. When cocaine was administered

alone and after pretreatment with yohimbine or atropine, it decreased

myocardial contractile force but increased the rate-pressure product.

Thus, in each case, it elicited effects that increased myocardial

oxygen consumption (i.e., an elevation in the rate-pressure product)

and other effects that decreased myocardial oxygen consumption (i.e.,

a reduction in contractile force). The reduction in myocardial

contractile force was presumably the result of the local anesthetic

actions of cocaine. However, when cocaine was administered after

pretreatment with a combination of yohimbine and atropine, dramatic

315
increases in both myocardial contractile force and the rate-pressure

product were observed. Clearly, in this latter instance, myocardial

oxygen consumption was greatly increased, but this did not result in

the expected decrease in coronary vascular resistance, which would be

required to support an increase in coronary blood flow sufficient to

meet the requirements of the heart during this response to cocaine.

This apparently inappropriate response of the coronary vasculature to

elevated myocardial oxygen demand elicited by cocaine administration

suggests that cocaine may exert an action on the coronary circulation

to prevent its normal dilation in response to increased oxygen

demand. This finding, although not evidence of coronary vasospasm,

clearly suggests that cocaine modifies the ability of the coronary

circulation to dilate appropriately in response to normal physiologic

stimuli (i.e., increased myocardial oxygen demand). These data

further suggest that synaptic mechanisms that control the rate of

norepinephrine release modulate the cardiovascular effects of cocaine

and that any pharmacologic influence that interferes with these

control mechanisms might be expected to significantly enhance its

cardiovascular actions.

More comprehensive studies of the effects of cocaine on cardiovascular function in animals have also been performed, but these

studies are few in number and all were performed in anesthetized

animals or isolated hearts where the component of cocaine’s cardiovascular action that is dependent upon central nervous system

stimulation is absent or diminished. Pierre et al. (1985) have shown

that a single large intravenous dose (200 mg) of cocaine in dogs

results in a reduction in arterial blood pressure, cardiac output, and

coronary blood flow.

Catravas and coworkers have reported two studies of cocaine toxicity

in conscious dogs (Catravas et al. 1978; Catravas and Waters 1981).

All animals in both studies exhibited cardiovascular effects of cocaine

(e.g., increased blood pressure and heart rate). Although the precise

role of the cardiovascular effects of cocaine in the lethality of this

drug when infused at a rate of 0.5 mg/kg/min until death in dogs was

unclear, these workers observed that the only interventions capable

of protecting all animals from a 39.5 mg/kg dose of cocaine (three

standard deviations above their previously determined mean lethal

dose) were chlorpromazine pretreatment and reduction of ambient

temperature to -5 °C.

The conscious dog is quite responsive to the cardiovascular effects of

cocaine. In this model, mean blood pressure was elevated 95.8±11.0

316
mmHg after a single 8 mg/kg dose, and smaller cocaine doses also

elicited dose-dependent increases in blood pressure (figure 8). Dogs

with chronically implanted arterial and venous catheters were given

cocaine intravenously in a range of doses (0.063, 0.125, 0.25, 0.5, 1.0,

2.0, 4.0, and 8.0 mg/kg) to establish the dose-response relationship

for this drug on blood pressure in this model. The data for this

dose-response curve were obtained over a period of 8 days with only

one cocaine dose being administered each day. Convulsions were

observed in all animals after administration of the 8 mg/kg dose of

cocaine, but no convulsions were noted at doses of 4.0 mg/kg or less.

FlGURE 8. Dose-response relationship for the effects of cocaine on

blood pressure in trained, conscious dogs

NOTE: The responses depicted were obtained when each dose of cocaine was administered as a single intravenous bolus with a minimum interval of 24 hours

between doses. Each point represents the mean ± standard error of seven

dogs.

Figure 9 illustrates the dose-dependent increase in the rate-pressure

product observed after administration of increasing doses of cocaine

on different days in conscious dogs instrumented for the measurement

317
of blood pressure, heart rate, and coronary blood flow as described

above. At all doses studied, cocaine administration resulted in a

significant increase in the rate-pressure product, and, at doses above

2 mg/kg, this index of myocardial oxygen consumption was more than

doubled. As described above, typically, increases in myocardial

oxygen demand are appropriately matched by coronary vascular

dilation in order to meet this increased need for oxygen and nutrients. At all doses of cocaine studied, coronary vascular resistance

increased despite apparent increases in myocardial oxygen demand.

Thus, in the conscious dog model, there appears to be evidence

FlGURE 9. The effects of cocaine on myocardial oxygen consumption

and coronery artery function in trained, conscious dogs

NOTE: Cocaine was administered In the doses shown with a minimum of 24 hours

between doses. The product of heart rate and blood pressure (open bars) was

utilized as an index of myocardial oxygen consumption, and coronary vascular

resistance (hatched bars) was calculated from arterial blood pressure and left

circumflex coronary artery blood flow. Each bar represents the mean ±

standard error of seven dogs.

for an action of cocaine to modify normal coronary function.

Whether or not the observed inappropriate increase in coronary

vascular resistance, in the face of increased myocardial oxygen

318
demand, is sufficient to produce myocardial infarction is an open

question. lt does seem quite clear, however, that cocaine exerts an

action to interfere with the normal control of coronary Mood flow.

Such an action could have disastrous consequences in individuals

whose ability to increase myocardial blood flow in response to an

increase in myocardial oxygen demand is already compromised by

atherosclerotic changes in their coronary vessels. Even in individuals

with normally functioning coronary arteries, the actions of cocaine to

increase myocardial oxygen demand, while interfering with the normal

blood flow regulation in the heart, could conceivably result in

myocardial ischemia.

There have been several studies of the cardiovascular effects of

cocaine in human subjects who were cocaine users. These subjects

received intravenous doses of cocaine ranging from 4 to 48 mg and

typically exhibited dose-related increases in heart rate and blood

pressure (Fischman et al. 1976; Resnick et al. 1977; Javaid et al. 1978;

Fischman et al. 1983). To put these doses in perspective, Fischman

and Schuster (1982) asked their subjects to compare the cocaine doses

received to their typical “street” dose. A dose of 16 mg was generally rated as similar to their average “street” dose, while doses of 24

and 32 mg were rated as among the highest those subjects had ever

experienced.

Recently, Fischman et al. (1985) reported that acute tolerance

develops to the techycardia associated with cocaine administration in

humans. When subjects were allowed to self-administer 96 mg of

cocaine intranasally 1 hour prior to an intravenous dose of cocaine,

the heart rate response to the intravenous dose was significantly

reduced in comparison to the response to the same intravenous

cocaine dose in the same subjects 1 hour after intranasal administration of a placebo. Although the increments in plasma cocaine

concentration produced by similar intravenous doses were not

significantly different in the two groups, the residual plasma cocaine

concentration remaining from the intranasal cocaine dose was

approximately 160 ng/ml at the time of intravenous cocaine administration in the one patient whose plasma cocaine concentrations

were presented. The authors interpreted these findings to be

indicative of acute tolerance to the actions of cocaine. Clearly,

these results demonstrate that, when cocaine is present in the body

at the time of administration of an acute intravenous cocaine dose,

the heart rate response to that dose is less than when the same

intravenous dose is administered in the absence of residual plasma

cocaine. Whether this is the result of functional tolerance, or merely

319
due to the fact that the dose (concentration)-response relationship

for cocaine, like most drugs, is not linear but rather forms a

rectangular hyperbola is unclear. That is, the “dose-response” curve

is linear only when response is plotted as a function of log dose.

Thus, a 450 ng/ml increment in plasma cocaine concentration would

be expected to produce a greater response, for example, when it

occurs from 0 to 450 ng/ml than when it occurs from 160 to

610 ng/ml. In this regard, it would be interesting to know whether

or not the reduced responsiveness to the intravenous cocaine dose

would occur if it were administered a sufficient time after the

intranasal cocaine dose for the residual plasma cocaine concentration

to fall to a negligible level. Similarly, it is important to determine

whether this reduced responsiveness is peculiar to heart rate or

whether it also occurs with other cardiovascular parameters. The

very efficient control of heart rate by baroreceptor reflexes makes

heart rate a particularly complex parameter to use to evaluate

tolerance, since a major component of the cardiovascular response to

cocaine is an increase in arterial blood pressure. Finally, as pointed

out recently by Porcet and his colleagues (1987), when the concentration of a drug that distributes rapidly to its site of action

cannot be measured at that active site, it may not be possible to

assess acute tolerance by bolus-dose experiments alone. Steady-state

experiments may also be required to distinguish acute functional

tolerance from apparent tolerance resulting from distributional

differences with drugs such as cocaine or nicotine, which distribute

very rapidly into the brain. Matsuzaki and coworkers (1978) also

repotted the development of tolerance to the cardiovascular actions

of cocaine after chronic administration, but they also reported that

chronic cocaine treatment decreased the elimination half-life of

cocaine by almost 50 percent. This suggests that the reduced

response to cocaine that they observed may have been the result of

lower cocaine plasma concentrations resulting from its faster metabolism rather than from a true functional tolerance. Unfortunately,

they did not measure cocaine plasma concentrations in conjunction

with their physiological measurements. Whether or not acute

functional tolerance actually occurs to the cardiovascular effects of

cocaine is uncertain at this time, but because of this uncertainty we

have modified our acute study protocols so that no more than one

cocaine dose is administered during any 24-hour period.

In conclusion, despite a lack of concrete evidence demonstrating

cardiovascular toxicity of cocaine in man, there is a growing volume

of circumstantial evidence that suggests that cardiac and vascular

toxicity associated with cocaine use may lead to myocardial infarction

320
and stroke. These data are complemented by observations in

conscious animal preparations that cocaine increases myocardial

oxygen consumption, as measured by the product of heart rate and

blood pressure, while simultaneously interfering with metabolic

vasodilation in the coronary circulation. Indeed, there is an inappropriate increase in coronary vascular resistance as a result of

cocaine treatment. Clearly, more studies are needed to better define

the effects of cocaine on the cardiovascular system, but because of

the complex effects of cocaine on the central nervous system and on

the heart and blood vessels, the integrated effects of cocaine on the

cardiovascular system can only be studied appropriately in the intact,

fully conscious animal.

REFERENCES

Altura, B.M.; Altura, B.T.; and Gebrewold, A. Cocaine induces spasms

of cerebral blood vessels: Relation to cerebral vascular accidents,

strokes and hypertension. Fed Proc 44:1637, 1985.

Catravas, J.D., and Waters, I.W. Acute cocaine intoxication in the

conscious dog: Studies on the mechanism of lethality. J Pharmacol

Exp Ther 217:350-356, 1981.

Catravas, J.D.; Waters, I.W.; Walz, M.A.; and Davis, W.M. Acute

cocaine intoxication in the conscious dog: Pathophysiologic profile

of acute lethality. Arch Int Phermecodyn Ther 235:328-340, 1978.

Chiueh, C.C., and Kopin, I.J. Centrally mediated release by cocaine

of endogenous epinephrine and norepinephrine from the sympathoadrenal medullary system of unanesthetized rats. J Pharmacol Exp

Ther 205:148-154, 1978.

Cohen, R.A.; Shepperd, J.T.; and Vanhoutte, P.M. Neurological

cholinergic prejunctional inhibition of sympathetic beta-adrenergic

relaxation in the canine coronary artery. J Pharmacol Exp Ther

229:417-421, 1984.

Cousineau, D.; Goresky, C.A.; and Rose, C.P. Decreased basal cardiac

interstitial norepinephrine release after neuronal uptake inhibition

in dogs. Circ Res 58:859-866, 1986.

Di Maio, V.J.M., and Garriott, J.C. Four deaths due to intravenous

injection of cocaine. Forensic Science Int 12:119-125, 1978.

Duke, M. Cocaine, myocardial infarction and arrhythmias--a review.

Conn Med 50:440-442, 1986.

Fischman, M.W., and Schuster, C.R. Cocaine self-administration in

humans. Fed Proc 41:241-246, 1982.

Fischman, M.W.; Schuster, C.R.; and Hatano, Y. A comparison of the

subjective and cardiovascular effects of cocaine and lidocaine in

humans. Pharmacol Biochem Behev 18:123-127, 1983.

321
Fischman, M.W.; Schuster, C.R.; Javaid, J.; Hatano, Y.; and Davis, J.

Acute tolerance development to the cardiovascular and subjective

effects of cocaine. J Pharmacol Exp Ther 235:677-682, 1985.

Fischman, M.W.; Schuster, C.R.; Resnekov, L.; Shick, J.F.E.;

Krasnegor, N.A.; Fennell, W.; and Freedmen, D.X. Cardiovascular

and subjective effects of intravenous cocaine administration in

humans. Arch Gen Psychiatry 33:983-989, 1976.

Gardezi, N. Cardiovascular effects of cocaine. JAMA 257:979-980,

1987.

Goldberg, M.R., and Robertson, D. Yohimbine: A pharmacological

probe for study of the alpha-2 adrenoreceptor. Pharmacol Rev

35:143-180, 1983.

Isner, J.M.; Estes, N.A.M.; Thompson, P.D.; Costanzo-Nordin, M.R.;

Subramanian, R.; Miller, G.; Katsas, G.; Sweeney, K.; and

Sturner, W.Q. Acute cardiac events temporally related to cocaine

abuse. N Engl J Med 315:1438-1443, 1986.

Javaid, J.I.; Fischman, M.W.; Schuster, C.R.; Dekirmenjian, H.; and

Davis, J.M. Cocaine plasma concentration: Relation to physiological and subjective effects in humans. Science 202:227-228, 1978.

Kirlin, P.C.; Kittleson, M.D.; and Johnson, L.E. Neurohumoral and

cardiopulmonary response to sustained submaximal exercise in the

dog. J Appl Physiol 62:1040-1045, 1987.

Koerker, R.L., and Moran, N.C. An evaluation of the inability of

cocaine to potentiate the responses to cardiac sympathetic nerve

stimulation in the dog. J Pharmacol Exp Ther 178:482-496, 1971.

Kossowsky, W.A., and Lyon, A.F. Cocaine and acute myocardial

infarction. Chest 86:729-731, 1984.

Langer, S.Z., and Enero, M.A. The potentiation of responses to

adrenergic nerve stimulation in the presence of cocaine: its relationship to the metabolic fate of released norepinephrine.

J Pharmacol Exp Ther 191:431-443, 1974.

Levine, S.R., and Welch, K.M.A. Cocaine and stroke: Current

concepts of cerebrovascular disease. Stroke 22:25-30, 1987.

Levy, M.N., and Blattberg, B. The influence of cocaine and desipramine on the cardiac responses to exogenous and endogenous

norepinephrine. Eur J Pharmacol 48:37-49, 1978.

Linden, C.H.; Vellman, W.P.; and Rumack, B. Yohimbine: A new

street drug. Ann Emerg Med 14:1002-1004, 1987.

Lundberg, G.D.; Garriott, J.C.; Reynolds, P.C.; Cravey, R.H.; and

Shaw, R.F. Cocaine-related death. J Forensic Sci 22:402-408, 1977.

Mathias, D.W. Cocaine-associated myocardial ischemia: Review of

clinical and angiographic findings. Am J Med 81:675-678, 1986.

322
Matsuda, Y.; Masuda, Y.; and Levy, M.N. The effects of cocaine and

metanephrine on the cardiac responses to sympathetic nerve

stimulation in dogs. Circ Res 45:180-187, 1979.

Matsuzaki, M.; Spingler, P.J.; Whitlock, E.G.; Misra, A.L.; and Mule,

S.J. Comparative effects of cocaine and pseudococaine on EEG

activities, cardiorespiratory functions, and self-administration

behavior in the rhesus monkey. Psychopharmacology (Berlin)

57:13-20, 1978.

Mittleman, R.E., and Wetli, C.V. Cocaine and sudden “natural” death.

J Forensic Sci 32:11-1 9, 1987.

Moore, J.I. Potentiation of the cardiac and pressor responses to

electrical stimulation of the cardiac sympathetic nerves by cocaine

in open-chest dogs. J Pharmacol Exp Ther 153:218-224, 1966.

Nanji, A.A., and Filipenko, J.D. Asystole and ventricular fibrillation

associated with cocaine intoxication. Chest 85:132-133, 1984.

Pierre, A.: Kossowksy, W.; Chou, S.-T.; and Abadir, A.R. Coronary

and systemic hemodynamics after intravenous injection of cocaine.

Anesthesiology 63:A28, 1985.

Porcet, H.C.; Benowitz, N.L.; Sheiner, L.B.; and Copeland, J.R.

Apparent tolerance to the acute effect of nicotine results in part

from distribution kinetics. J Clin Invest 80:1466-1471, 1987.

Resnick, R.B.; Kestenbaum, R.S.; and Schwartz, L.K. Acute systemic

effects of cocaine in man: A controlled study by intranasal and

intravenous routes. Science 195:696-698, 1977.

Ritchie, J.M., and Greene, N.M. Local anesthetics. In: Gilman, A.G.;

Goodman, L.S.; Rall, T.W.; and Murad, F., eds. Goodman and

Gilman’s The Pharmacologic Basis of Therapeutics. New York:

Macmillan, 1985. pp. 302-321.

Schachne, J.S.; Roberts, B.H.; and Thompson, P.D. Coronary-artery

spasm and myocardial infarction associated with cocaine use.

N Engl J Med 310:1665-1666, 1984.

Smith, H.W.B.; Liberman, H.A.; Brody, S.L.; Battey, L.L.;

Donohue, B.C.; and Morris, D.C. Acute myocardial infarction

temporally related to cocaine use. Ann Intern Med 107:13-18, 1987.

Trendelenburg, U. The supersensitivity caused by cocaine.

J Pharmacol Exp Ther 125:55-65, 1959.

Trendelenburg, U. The effect of cocaine on the pacemaker of

isolated guinea-pig atria. J Pharmacol Exp Ther 161:222-231, 1968.

Whitby, L.G.; Hertting, G.; and Axelrod, J. Effect of cocaine on the

disposition of noradrenaline labelled with tritium. Nature

187:604-605, 1960.

Wilkerson, R.D. Yohimbine pretreatment enhances the cardiovascular

actions of cocaine in anesthetized dogs. Fed Proc 46:1143, 1987.

323
Wilkerson, R.D. Cardiovascular effects of cocaine in conscious dogs.

FASEB J 2:A1802, 1988.

Yamaguchi, N.; DeChamplain, J.; and Nadeau, R.A. Regulation of

norepinephrine release from cardiac sympathetic fibers in the dog

by presynaptic alpha and beta receptors. Circ Res 41:108-117, 1977.

Zimmerman, F.H.; Gustafson, G.M.; and Kemp, H.G. Recurrent

myocardial infarction associated with cocaine abuse in a young man

with normal coronary arteries: Evidence for coronary artery spasm

culminating in thrombosis. J Am Coll Cardiol 9:964-968, 1987.

ACKNOWLEDGMENTS

This research was supported by grant DA-04038 and contract 271-87-

8130 from the National Institute on Drug Abuse.

AUTHOR

R. Douglas Wilkerson, Ph.D.

Department of Pharmacology

Medical College of Ohio

C.S. #10008

Toledo, OH 46699

324
Arteriosclerotic Toxicity of Cocaine

Ronald O. Langner, Collette L. Bement, and

Lynda E. Perry

INTRODUCTlON

The purpose of our research is to test the hypothesis that cocaine is

a potential risk factor for the premature development of arteriosclerosis. The abuse of cocaine has been associated with a number of

cardiovascular toxicities including tachycardia, hypertension, ventricular arrhythmias, and death. In small doses, cocaine can depress

the heart rate as a result of central vagal stimulation. When cocaine

is used intravenously in high concentrations, it produces a doserelated increase in heart rate and blood pressure (Fischman et al.

1976). These effects are due both to central sympathetic stimulation

and the peripheral effects of cocaine in inhibiting the reuptake of

catecholamines at adrenergic nerve endings. The resulting increased

sympathetic tone leads to tachycardia and raises the blood pressure,

which in turn increases the oxygen demand of the heart. This

sequence of events has been suggested as a possible cause of myocardial infarctions observed in some individuals following the abuse of

cocaine (Ring and Butman 1986).

Cardiovascular disease remains the leading cause of death in the

United States and Western Europe. Arteriosclerosis, which accounts

for the majority of these deaths, is a group of diseases characterized

by increased collagen deposition and loss of elasticity in the arterial

wall. Atherosclerosis, which is a form of arteriosclerosis, is characterized by the presence of yellowish plaques (atheromas) containing

cholesterol, as well as by the loss of elastin and deposition of collagen. These lesions are normally found in the intima and media of

large- and medium-sized arteries. The earliest arteriosclerotic lesions

are thought to begin during the first decade of life (McGill 1984) and

may or may not contain increased amounts of cholesterol. During the

325
third and fourth decades of life, fibrous plaques with necrotic, lipidrich cores surrounded by smooth muscle and connective tissue appear.

These lesions undergo vascularization, hemorrhage, and ulceration, and

some eventually are covered by thrombi, which occlude the artery and

cause ischemic necrosis in the affected tissue. Even though the clinical manifestations of arteriosclerosis vary with the location of the

lesions, all the underlying arterial lesions are essentially the same.

Many risk factors have been associated with the development of

arteriosclerosis. Hopkins and Williams (1981) listed 246 different

factors that have been implicated in the development of the disease.

These were divided into primary and secondary risk factors with

elevated serum cholesterol, hypertension, cigarette smoking, and

obesity being considered as primary risk factors. In a recent review

of the mechanism of action in atherogenesis, Ross (1986) suggests

that all atherosclerotic risk factors cause an injury to the blood

vessel wall and represent some of the earliest events in lesion forma

tion. When the injury is recurrent, the resulting damage to the

arterial lining stimulates migration and proliferation of smooth muscle

cells to the damage site. In the presence of moderate to elevated

serum cholesterol, smooth muscle cells are thought to accumulate

large amounts of cholesterol, resulting in cellular necrosis. This is

further complicated by a buildup of fibrous tissue (mostly collagen)

and calcium leading to a clinically significant narrowing of the

arteries.

Several investigators have demonstrated that intravenous injection of

epinephrine results in the development of arteriosclerotic lesions,

which are similar to those found in the early stages of human arteriosclerosis (Oester 1959; Lorenzen 1961). The lesions are primarily

localized in the thoracic aorta and the ascending and descending portions of the aortic arch. The histological changes induced in the

aorta were primarily in the medial layers, with some changes in the

intima, and were characterized by damage or necrosis of the inner

arterial walls, breakdown, splitting, and eventual disappearance of the

elastic lamella and an infiltration of the injured area with water,

proteins, and glycosaminoglycans. Surrounding this necrotic area are

layers of smooth muscle cells, which appear to be active both as

fibroblasts and phagocytes. Calcification of the injured area seems to

be a secondary event, which follows the deposition of glycosaminoglycans. These epinephrine-induced lesions do not exhibit increased

lipid accumulation. However, experimental studies by Constantinides

et al. (1958) have shown that arteries injured by epinephrine will

326
incorporate lipids when rabbits are fed a high cholesterol diet either

during or after epinephrine treatment.

Fuller and Langner (1970), studying the biochemical changes induced

by epinephrine treatment, found that one of the earliest changes was

an increase in the rate of aortic collagen synthesis. This change was

observed after only 4 days of treatment and before any gross alterations in aortic morphology were noted. The authors suggested that

changes in aortic collagen synthetic rates represented a primary biochemical defect. The mechanism whereby epinephrine causes these

changes is unknown. Waters and de Suto-Nagy (1950) suggested that

the hypertensive effect of epinephrine caused an injury to the endothelial lining, which was responsible for the observed changes. Since

cocaine exerts its cardiovascular effects by potentiating the effects

of epinephrine and norepinephrine, these studies were initiated to

establish the potential of intravenous cocaine to cause an arteriosclerotic injury to the vessel wall.

METHODS

Fifteen male New Zealand rabbits weighing 2.8 to 3.2 kg were used in

these studies. Five rabbits were placed in a saline control group and

the remaining 10 rabbits were placed in a drug treatment group. All

animals were given standard rabbit chow and water ad libitum. Each

rabbit was weighed daily and injected via the marginal ear vein for

14 days using an infusion pump set to deliver fluid at a rate of 1 ml

per minute. The saline control group received injections of sterile

saline for 1 minute. The cocaine-treated rabbits were injected with

the following doses of drug: days 1 and 2, 5.5 mg/kg; days 3 through.

10, 4.5 mg/kg; and days 11 through 14, 5.0 mg/kg.

The initial dose of 5.5 mg/kg was selected following preliminary

studies that indicated that this dose of cocaine would result in an

elevation in the heart rate. After 2 days, the dose of cocaine was

decreased to 4.5 mg/kg, because the initial dose of cocaine resulted

in the death of two rabbits and visibly stressed the remaining

animals. After 10 days, the dose was increased to 5.0 mg/kg, because

the rabbits were exhibiting little visible response to the cocaine,

suggesting that some degree of tolerance was developing.

Twenty-four hours after their last injection, the rabbits were lightly

anesthetized with ether and killed by cervical dislocation. The thoracic aorta and the liver were quickly removed and placed in cold

Kreb’s bicarbonate buffer (pH 7.4). The thoracic aorta was cleaned

327
of loosely adhering fat and tissue, opened longitudinally, and inspected for gross aortic lesions. Aortic samples from the descending

aortic arch were removed for histological evaluation. The liver was

handled in a similar manner. The aorta and liver were then placed in

fresh buffer containing 3 uCi of 14C-proline and 0.2 mM L-proline

for either 90 minutes (aorta) or 60 minutes (liver), following

procedures previously reported (Langner and Bement 1985).

Following the incubation period, the tissues were rinsed in 5 percent

ice-cold trichloroacetic acid (TCA) to prevent further incorporation of

the labeled proline. The tissues were then homogenized in 9 volumes

of normal saline using a glass coaxial homogenizer. Appropriate aliquots were taken for analysis of collagen hydroxyproline content

(Kivirrikko et al. 1967), protein content (Lowry et al. 1951), cholesterol content (Franey and Amador 1968). and protein synthetic rate

(Langner and Bement 1985). For histological evaluation, all samples

were fixed in 10 percent formalin and were stained with hematoxyline

and eosin, and Verhoeff’s. The data were evaluated for statistically

significant differences using the Student’s t-test (Snedecor and

Cochran 1976).

RESULTS

As shown in table 1, the cocaine treatment did not cause any

alteration in serum cholesterol levels. The final body weights of the

TABLE 1. Effect of cocaine on final body weight and serum

cholesterol levels

Group N

Final Serum Final Body

Cholesterol Weight

(mg/100 ml) (kg)

Saline

Cocaine

5 44.12±7.74 3.46±0.2

8 49.14±5.89 2.90±0.1*

*p<.05 compared to saline-treated controls.

NOTE: Data expressed as mean ± SE.

328
FIGURE 1. Aorta from a rabbit treated for 14 days with cocaine,

showing general disruption of elastic fibers

NOTE: The spaces between the elastic fibers appear widened with increased deposition

of granular material. (Verhoeff’s, 250X.)

329
FIGURE 2. Aorta from a rabbit treated for 14 days with cocaine,

showing a compression of the medial layer

NOTE: The intimal layer is thickened and appears to contain increased numbers of

foam cells. (Verhoeff's, 250X.)

cocaine-treated rabbits were significantly lower than the salinetreated controls. Visual inspection of the aorta revealed that five of

the eight cocaine-treated animals had some focal lesions, primarily in

the area of the upper thoracic and aortic arch. These lesions were

small and raised and had a pearly white appearance. There were no

observable lesions in the saline-injected controls.

Histological evaluation of the aortic lesions (figures 1 and 2)

demonstrated that the cocaine-induced lesions could be quite

different. In figure 1, the lesion appears to be primarily located in

the media with little evidence of intimal involvement. The lesioned

area is characterized by edema and widening of the spaces between

the elastic lamella. In addition, the elastic fibers show fragmentation

and general disruption. Within the widened spaces there appears to

be increased deposition of a granular exudate. In figure 2, the lesion

is much different and shows a compression of the media with little

330
rearrangement or damage. In this lesion, however, there is intimal

proliferation over the area of compressed media, which appears to

have a foam-cell-like appearance. Histological evaluation of the

livers from cocaine-treated rabbits did not exhibit any pathological

alterations.

The data in table 2 show the tissue body weight ratios of both the

liver and thoracic aorta. As can be seen, the administration of

cocaine for 14 days did not cause any change from control values

even though the total body weights of the cocaine-treated rabbits

were less than the saline animals (table 1). There also was no

change in tissue cholesterol content in either the aorta or the liver.

TABLE 2. Tissue to body weight ratios and total cholesterol content

of thomcic aortas and livers of rabbits treated with

cocaine

Thoracic Aorta Liver

Tissue/Body Cholesterol Tissue/Body Cholesterol

Group (g/kg) (mg/g tissue) (g/kg) (mg/g tissue)

Saline 0.25±.02 3.94±.52 19.40±1.37 4.48±16

Cocaine 0.28±02 3.83±.39 18.37±0.59 4.38±16

NOTE: Data expressed as mean ± SE.

The total collagen content in both the aorta and liver were estimated

by measuring tissue hydroxyproline content. Hydroxyproline is primarily found in collagen. Smaller amounts of hydroxyproline are also

found in elastin and the C1q component of complement. As seen in

table 3, the administration of cocaine did not cause any significant

alteration in either aortic or liver hydroxyproline content. The

protein content of both the aorta and liver also were not altered by

the administration of cocaine for 14 days.

The data in table 4 show that there was a significant increase in the

rate of protein synthesis in the thoracic aortas of rabbits treated

with cocaine, as indicated by a threefold increase in the rate of 14Cproline incorporation into aortic proteins. The livers of the cocainetreated rabbits did not demonstrate any change in protein synthetic

331
activity in response to the cocaine administration. These data, therefore, suggest a relatively specific response of the blood vessel wall to

the administration of cocaine.

TABLE 3. Hydroxyproline and protein content of thoracic aortas and

livers of rabbits treated with cocaine

Thoracic Aorta Liver

Hydroxyproline Protein Hydroxyproline Protein

Group (μg/kg) (mg/g) (μg/g) (mg/g)

Saline 7696±394 104±11 737±81 169±4

Cocaine 7520±460 98±9 810±89 174±2

NOTE: Data expressed as mean ± SE.

TABLE 4. Protein synthetic activity of thomcic aortas and livers of

rabbits treated with cocaine

Group

Thoracic Aorta Liver

dpm/mg Protein dpm/mg Protein

90 min 60 min

Saline

Cocaine

226.6±60 199.5±35

688.7±165* 282.7±29

*p<.05 compared to saline controls.

NOTE: dpm of 14C-proline incorporated into either aortic or liver proteins per

milligrarn of tissue protein. All data expressed as mean ± SE.

DISCUSSION

The purpose of our studies was to determine the potential of chronic

cocaine administration to induce early arteriosclerotic changes in the

blood vessels of rabbits. In humans, arteriosclerosis frequently begins

in the form of scattered foci in which the innermost layers of the

vascular wall show signs of damage, accompanied by the growth of

332
repair tissue (Constantinides 1965; Daoud et al. 1964). Injury is

characterized by the disruption and disintegration of the innermost

elastic lamella and by an increase in amorphous materials, particularly

proteins and glycosaminoglycans. At this early lesion stage there is

not an abnormal amount of cholesterol. Lipids appear to accumulate

slowly until the lesions become grossly visible and are observed as

fatty streaks (Constantinides 1965). The histological changes we

observed in the cocaine-treated rabbits appear to be similar to early

human lesions described by Constantinides (1965). The lesions are

primarily located in the medial layers of the aortic wall and may or

may not be grossly visible. These early cocaine-induced lesions do

not contain elevated levels of cholesterol (table 2) and do not appear

to be the result of any alteration in serum cholesterol levels

(table 1). The observed increase in aortic protein synthetic rates

(table 4) suggests that the tissues are in a proliferative stage of

development. The absence of any change in total collagen or total

proteins may be a result of the lesions being in an early stage of

development. At this stage of development, sufficient time may not

have elapsed to allow for a detectable accumulation of these proteins.

Another possibility is that cocaine treatment has induced both

synthesis and degradation of proteins, resulting in no net accumulation of either collagen or total proteins.

The changes induced by cocaine administration were found only in

the aorta and not in the liver. Histological and biochemical examination of liver sections revealed no significant alterations. These data

would suggest that the induction of aortic protein synthetic rates is

the result of a direct change in the aortic wall and not the result of

some generalized increase in body protein synthetic activity. When

cholesterol is used to induce atherosclerotic lesions in rabbits, both

the aorta and the liver have markedly elevated tissue cholesterol

levels and increased rates of protein synthetic activity (Langner and

Modrak 1981). The ability of cocaine to induce arteriosclerotic

changes appears to be more specific in that only the aortic wall is

affected. The mechanism by which cocaine produces its effect is not

known. The biochemical and histological changes produced by cocaine

administration, however, are very similar to the changes observed

following epinephrine administration (Langner and Fuller 1973). lt

has been suggested that epinephrine administration causes an increase

in blood pressure, which in some manner may cause an injury to the

intimal lining of the blood vessel (Waters and de Suto-Nagy 1950).

Since cocaine has the ability to potentiate endogenous catecholamine

activity, the arteriogenic effect of cocaine may be the result of

increased catecholamine activity resulting in an increase in heart rate

333
and blood pressure. The exact mechanism by which increases in

blood pressure and heart rate damage the arterial wall is unknown

and needs to be investigated.

The significance of our observations in rabbits to the development of

arteriosclerotic disease in man remains to be elucidated. Further

experiments using different doses of cocaine for varying times must

be investigated. In addition, the potential for cocaine to act

synergistically with other known cardiovascular risk factors to cause

premature atherosclerosis should also be studied. Our data do,

however, clearly demonstrate that cocaine has the potential to cause

an injury to the blood vessel wall that may result in arterioscleroticlike changes.

SUMMARY

The repeated injection of cocaine results in the formation of

arteriosclerotic lesions in rabbits, which appear to be similar to early

arteriosclerotic lesions seen in man. Biochemically, the aortas

exhibited increased rates of protein synthesis, without any change in

tissue cholesterol content. The cocaine-treated rabbits exhibited no

change in serum cholesterol levels. These data suggest that the

abuse of cocaine may result in damage to the aorta, which could

result in the premature onset of cardiovascular disease and its

complications.

REFERENCES

Constantinides, P. Experimental Atherosclerosis. New York:

Elsevier, 1965. 91 pp.

Constantinides, P.; Gutmann-Augersperg, N.; and Hospes, D.

Acceleration of intimal atherogenesis through prior medial injury.

AMA Arch Path 66:247-254, 1958.

Daoud, A.; Jarmolych, J.; Zumbo, A.; and Fani, K. “Pre-Atheroma”

phase of coronary atherosclerosis in man. Exp Mol Pathol 3:475

484, 1964.

Fischman, M.W.; Schuster, C.R.; Resnekov, L.; Shick, J.F.E.;

Krasnergor, N.A.; Fennell, W.; and Freedman, D.X. Cardiovascular

and subjective effects of intravenous cocaine administration in

humans. Arch Gen Psychiatry 33:983-989, 1978.

Franey, R.J., and Amador, E. Serum cholesterol measurement based

on ethanol extraction and ferric chloride-sulfuric acid. Clin Chim

Acta 21:255-263, 1968.

334
Fuller, G.C., and Langner, R.O. Elevation of aortic proline hydroxylase: A biochemical defect in experimental arteriosclerosis.

Science 168:987-989, 1970.

Hopkins, P.N., and Williams, R.R. A survey of 246 suggested coronary

risk factors. Atherosclerosis 40:1-52, 1981.

Kivirrikko, K.I.; Laitinen, O.; and Prockop, D.J. Modification of a

specific assay for hydroxyproline in urine. Anal Biochem 19:249

255, 1967.

Langner, R.O., and Bement, C.L. Lesion regression and protein

synthesis in rabbits after removal of dietary cholesterol.

Arteriosclerosis 5:74-79, 1985.

Langner, R.O., and Fuller, G.C. Collagen synthesis in thoracic aortas

of rabbits with epinephrine-thyroxine induced arteriosclerosis.

Atherosclerosis 17:463-469, 1973.

Langner, R.O., and Modrak, J.B. Alteration of collagen synthesis in

different tissues of the atherosclerotic rabbit. Artey 9:253-261,

1981.

Lorenzen, I. Vascular connective tissue under the influence of

thyroxin. I. Epinephrine-thyroxine induced arteriosclerotic lesions.

Acta Endocrinol 36:197-211, 1961.

Lowry, O.H.; Rosebrough, J.J.; Farr, A.L.; and Randall, R.J. Protein

measurement with the folin-phenol reagent. J Biol Chem 193:265-

275, 1951.

McGill, H.C., Jr. Persistent problems in the pathogenesis of

atherosclerosis. Arteriosclerosis 4:443-451, 1984.

Oester, Y.T. Adrenal medullary hormones and arteriosclerosis. Ann

NY Acad Sci 72:885-895, 1959.

Resnick, R.B.; Kestenbaum, R.S.; and Schwartz, L.K. Acute systemic

effects of cocaine in man: A controlled study by intranasal and

intravenous routes. Science 195:696-698, 1977.

Ring, M.E., and Butman, S.M. Cocaine and premature myocardial

infarction. Drug Ther 16:117-125, 1986.

Ross, R. The pathogenesis of atherosclerosis--an update. N Engl J

Med 314:488-500, 1986.

Snedecor, G.W., and Cochran, W.G. Statistical Methods. 6th Edition.

Ames, Iowa: Iowa State University Press, 1976. 534 pp.

Waters, L.L., and de Suto-Nagy, G.I. Lesions of the coronary arteries

and great vessels of the dog following injection of adrenaline.

Science 111:634-635, 1950.

ACKNOWLEDGMENTS

These studies were supported in part by U.S. Public Health Service

grant DA 03846 from the National Institute on Drug Abuse.

335
AUTHORS

Ronald O. Langner, Ph.D.

Collette L. Bement, M.S.

Lynda E. Perry, B.S.

Department of Pharmacology and Toxicology

University of Connecticut

Storrs, CT 06268

336
Mechanisms of Cocaine Abuse

and Toxicity: An Overview

Thomas V. Dunwiddie

lNTRODUCTlON

lt is characteristic of pharmacology that drugs are often initially

perceived in terms of their beneficial or therapeutic uses, and only

after they have come into common use by a large number of individuals do the drawbacks and difficulties attendant upon their use

become known. Certainly this has been the case with cocaine.

Initially introduced and popularized for various purposes such as

increasing alertness, elevation of mood, overcoming opiate addiction,

etc., it became clear even during the last century that cocaine

overuse had the potential for severely disrupting both physical and

psychological well-being. The neurobiological basis for the abuse

potential of cocaine was identified in a general sense by Sigmund

Freud, who did much to initially popularize the drug. Freud

suggested that the feelings of pleasure and well-being that accompanied the use of cocaine were quite similar to those engendered by

many other types of stimuli in daily life that give pleasure. Now,

with over a century of neurobiological research to lend further

insight, it is apparent that the ability of cocaine to so potently

disrupt normal activities stems from the fact that it can interact

directly with the same brain systems (sometimes called “reward”

pathways) that are normally activated indirectly by pleasurable

stimuli. In a sense, cocaine can “short-circuit” the means by which

gratification is usually achieved, and the typical pleasurable activities

(e.g., eating, drinking, sexual behaviors, etc.) become less satisfying.

For both the drug abuser of the 1980s and for laboratory animals in

drug self-administration protocols, cocaine acts as a potent reinforcer

that supersedes nearly all other kinds of reinforcement as a

determinant of behavior.

337
Unfortunately, it is clear that simply understanding the biological

bases for specific maladaptive behaviors does not immediately dictate

their solution. In fact, given our current understanding of the

biological underpinnings of drug self-administration behavior, one

might well adopt a more pessimistic view of the likelihood of

developing pharmacological treatments for substance abuse. lf drugs

such as cocaine directly activate neural substrates of reward, is it

even possible to develop pharmacological treatments that could block

pharmacological reward without severely disrupting normal appetitive

behaviors? At this point, it is perhaps premature to try to answer

this question. Nevertheless, as we begin to understand more about

the basic biology of drug abuse, it is clear that our attempts to

control such behaviors can shift from the purely empirical, to

strategies that are directed by an understanding of the neural

systems involved. Many of the papers in this monograph contribute

substantially to this understanding, and it seems probable that the

development of more successful treatments for drug abuse may

eventually develop out of the basic science presented at this meeting.

When considering the recent research advances in this field, it is

difficult to point to any specific observations as being “key” advances

in our understanding of drug abuse. What is apparent, however, is

that, at many different levels of inquiry, a simplistic model concerning the actions of cocaine is being replaced by a more detailed

and necessarily more complicated picture of action. The original

pharmacological picture of cocaine was that of a local anesthetic,

which upon further study was also found to be an inhibitor of the

uptake of the neurotransmitter norepinephrine. Now, however, we

know that cocaine inhibits not only the uptake of norepinephrine but

also that of dopamine and serotonin. Furthermore, the work of

several participants in this review (Reith, this volume; Hanbauer, this

volume) suggests that high-affinity dopamine uptake, which until now

has been regarded as a unitary process, may represent multiple

processes with independent pharmacological properties.

In a similar vein, some of the earlier pharmacological studies of drug

self-administration suggested that dopamine was centrally involved in

such behaviors as “the” reward transmitter. However, it is difficult

now to regard pharmacological reinforcement as a monolithic

“system,” in which the mere occupation of a dopamine receptor

somehow can be equated with positive reinforcement. Instead, it is

clear that a drug such as cocaine, which causes an inhibition of

dopamine uptake, elicits a complex range of effects on dopaminergic

systems. Moreover, it is likely that a variety of transmitter systems

338
(opioid, cholinergic, and most likely peptides such as neurotensin and

cholecystokinin as well) may also be involved in ways that remain

undetermined. As was suggested by several participants at this

conference, reward systems may have distinct initiation, mediation,

and presumably termination components, and pharmacological agents

might interact with any one of these components to modify drugseeking and self-administration behaviors. Similarly, when dealing

with drugs that are self-administered, as well as with drugs that

disrupt such behaviors, one cannot focus simply upon the rewarding

aspects of these drugs, but must also take into account their effects

upon motor systems and their abilities to act as discriminative cues.

It seems likely that only by subdividing every aspect of cocaine’s

actions (drug self-administration into its component behavioral

processes; the biochemical actions of cocaine into effects upon different transmitters, ion channels, receptors, and uptake sites) will we

be able to fully understand the way in which substance abuse evolves.

RECENT PROGRESS IN UNDERSTANDING THE ACTlONS OF

COCAINE

Biochemical Aspects of Cocaine Action

The last decade has seen a remarkable change in the level of sophistication at which we understand the interactions of transmitters with

receptors and the secondary processes by which receptor activation

by a transmitter is translated into the physiological responses of

neurons to such transmitters. However, our understanding of drugs

that interact with more peripheral aspects of neurotransmission, such

as allosteric regulation of transmitter binding, neurotransmitter

uptake, etc., has perhaps lagged. In part, this is due to the fact

that it is more difficult to measure physiological responses to these

types of drugs; if a drug such as cocaine inhibits uptake, this does

not usually lead directly to any well-defined physiological response,

but only to indirect effects that represent the interaction of

increased extracellular concentrations of transmitters with their

receptors. Correspondingly, studies of cocaine binding in the brain

have been hampered somewhat by the difficulties in linking specific

binding sites to functional effects; although binding can be observed,

it is often complex, and the physiological consequences of binding to

these sites are often subtle.

One approach to these difficulties has been to correlate the relative

potencies of drugs in terms of well-defined biochemical properties

with their ability to elicit relatively complex behaviors such as drug

339
self-administration, without attempting to define the intervening

physiological events. Kuhar adopted the approach of labeling specific

neurotransmitter uptake sites with selective ligands, and then characterizing their displacement by a variety of abused and nonabused

drugs (Kuhar et al., this volume; Ritz et al. 1987). Binding to the

dopamine uptake site generally correlated well with the extent to

which these drugs are self-administered in animals, whereas their

ability to inhibit the uptake of serotonin and norepinephtine did not

display such a relationship. These data provide further support for

the hypothesis that potentiation of the effects of dopamine is intimately involved in pharmacological reward. In addition, these studies

help to resolve the continuing dilemma of how to reconcile the

“dopamine hypothesis” with the experimental observation that a

number of local anesthetics are self-administered. The present

findings imply that the reinforcing actions of some local anesthetics

are a function of their actions upon the dopamine uptake site and

that local anesthetic actions may be only peripherally involved. The

use of newer pharmacological agents, such as the more selective

uptake inhibitors, possibly irreversible inhibitors of uptake, and local

anesthetics that have little or no effect upon uptake processes, may

lead to an even more finely drawn portrait of the mechanisms by

which monoamine uptake occurs, the ways in which cocaine can

interact with such processes, and how this can lead to pharmacological reinforcement.

The experimental results discussed by a number of the participants

also suggest that our current models of high-affinity uptake processes

for monoamine transmitters need revision. The observations of Reith

(this volume) made it clear that, although cocaine affects the uptake

of norepinephrine, dopamine, and serotonin, it is not a particularly

suitable ligand for characterizing the corresponding uptake sites. In

particular, the binding of labeled cocaine appears to correspond in

many regions with high-affinity uptake of dopamine and serotonin,

but not norepinephrine, even though cocaine is able to inhibit uptake

of norepinephrine quite well. Disparities also arose between some of

the effects of metaphit (an irreversible acylator of PCP-binding sites)

upon binding and behavior; for example, metaphit treatment in vivo

blocked the locomotor stimulant effects of cocaine (but not indirect

dopamine-releasing agents such as amphetamine), but did not affect

either dopamine uptake or cocaine binding. The work of Hanbauer

(this volume), who demonstrated that an unidentified endogenous

peptide as well as GABA modulin are allosteric regulators of 3Hcocaine binding, also suggests that the uptake processes may not be

simple transporters, but may have a complex regulation by endogenous

340
agents that might be perturbed by cocaine. Clearly, a better understanding of the transport process per se will lead to a more detailed

picture as to how cocaine acts as well.

Physiological Aspects of Cocaine Action

In terms of the electrophysiological actions of cocaine, there has

been relatively little work done in comparison with other drugs of

abuse such as amphetamine. Relatively recent reports have demonstrated that cocaine can inhibit the spontaneous firing of neurons in

the locus coeruleus, the ventral tegmental area, and the dorsal raphe

(White 1985; Pitts and Marwah 1986; Brodie and Dunwiddie 1986;

Lakoski and Cunningham, this volume). In each case where this has

been examined, the mechanism underlying this inhibition appears to

be a facilitation of the actions of the endogenous monoamine transmitter (norepinephrine, dopamine, and serotonin, respectively) upon

autoinhibitory receptors.

Two aspects of these responses will require further study. First, the

general hypothesis of cocaine action postulates a facilitatory action

of cocaine at monoaminergic synapses; however, if the neurons containing these transmitters are themselves inhibited by cocaine, then

there is no release of transmitter to provide a substrate upon which

cocaine can act. It is possible that cocaine in behaviorally active

doses does not totally suppress presynaptic activity, but merely

reduces it, and in this situation the net effect of cocaine administration could still be an enhanced postsynaptic effect. Alternatively,

as White (1985) has demonstrated, the mesocortical dopamine neurons

of the ventral tegmental area may be relatively insensitive to the

impulse-inhibiting effects of cocaine, since they appear to lack the

autoinhibitory receptors found on other ventral tegmental area

neurons. Thus, cocaine might selectively enhance activity in mesocortical pathways while having a less pronounced effect upon the

mesoaccumbens and other projections that have autoreceptors. The

other aspect of the physiological effects of cocaine that needs

further investigation concerns the relative sensitivity of different

monoamine systems to cocaine and the ways in which these systems

interact. Lakoski and Cunningham (this volume) presented evidence

that the raphe nucleus appears to be the most sensitive of the

monoamine-containing nuclei to the inhibitory effects of cocaine.

This effect was shared by other serotonin uptake inhibitors such as

fluoxetine, but not by selective dopamine and norepinephrine uptake

inhibitors. Since there is behavioral evidence to suggest that

serotonergic systems are not involved directly in the rewarding

341
effects of drugs such as cocaine (Koob and Hubner, this volume), it

is possible that inhibition of serotonergic systems might well relate to

some of the other, nonreinforcement properties of cocaine. Behavioral and electrophysiological studies involving selective serotonin

uptake inhibitors such as fluoxetine and paroxetine may help to

clarify these issues.

RELATlNG BIOCHEMICAL AND PHYSIOLOGICAL EFFECTS OF

COCAINE TO "REWARD"

Although the effects of cocaine can be studied independently from a

consideration of pharmacological reward, at some point it becomes

useful to try to relate specific changes in neuronal function to the

psychological concept of reinforcement. A problem that arose

repeatedly during this conference was the difficulty in quantifying

“reward” per se. As was pointed out by several speakers (Balster,

Winger, Dworkin, and Johanson), the rate of responding on a lever

for drug administration is a clearly inadequate measure of the

“potency” of a rewarding stimulus. Typically, animals will respond to

increasing doses of drug with increasing response rates up to a point,

but beyond that point responding drops off, so that at the highest

doses response rates may be quite low. Nevertheless, when allowed

to choose between a lower or higher dose of drug, animals typically

select the higher dose, even though it is associated with lower rates

of administration (Johanson, this volume). Several types of protocols

have potential for determining “reward potency,” such as choice

studies, and so-called break-point studies, in which the measure of

reward is determined by increasing the number of responses required

of the animal for each successive injection. The point at which the

animal stops working for a given dose of drug thus becomes the

“break-point” for that dose.

Presumably this technique will lead to the characterization of drugs

not only in terms of relative potency (i.e., the amount of drug

required to maintain equivalent break-points) but also in terms of

efficacy (i.e., the maximum break-point that can be established by

any dose of the drug). The development of more unequivocal techniques such as these for characterizing the rewarding properties of

drugs may lead to a better understanding of the neurobiological basis

for reward as well. Dworkin and Smith (this volume) also discussed

ways in which to dissociate the various pharmacological properties of

cocaine, viz, the reinforcing effects, the effects upon response rate,

and the stimulus properties. Different types of behavioral protocols

are sensitive to each of these variables, and by suitable experimental

342
design, it is possible to fractionate the changes in behavior attributable to each of these actions of cocaine.

The approach used by Kuhar et al. (this volume) to identify binding

sites in the brain that are related to the reinforcing properties of

drugs is an important first step in determining how drugs such as

cocaine act. With improving behavioral techniques for quantifying

reinforcement, these correlations between biochemistry and behavior

may become even tighter. However, it is also clear that defining

reinforcement by means of a single biochemical parameter may be too

simplistic. As was pointed out by Balster (this volume), in terms of

reward, cocaine and amphetamine appear quite similar, although

biochemically they probably act through quite different mechanisms.

On the other hand, other drugs that share some of cocaine’s actions

appear quite different in behavioral studies. There are a number of

possible explanations for this anomaly. First, it is clear that the

pharmacokinetics of the drug response is an important variable:

Johanson (this volume) demonstrated that the rate of drug intrusion

could be as important a determinant as dose in affecting reward

potency. When the rewarding dose of drug is injected over a

prolonged period of time, the rewarding effects are markedly

diminished. Thus, a drug that enters the brain relatively slowly may

have diminished rewarding potency as a result. Second, a

combination of actions may be required in order to elicit reward.

Effects upon nondopamine synapses may either contribute to or

diminish reinforcement; since many of the regions thought to be

important in the effects of cocaine also receive serotonergic and

noradrenergic inputs, it may be necessary to take into account the

net effect of any given drug and dose on activity in those regions.

For example, although imipramine, like cocaine, is a good inhibitor of

serotonin and norepinephrine uptake, it is a relatively poor dopamine

uptake inhibitor. This may explain its very weak efficacy as a

positive reinforcer.

Although biochemical experiments have given numerous insights into

the mechanisms of reinforcement, physiological studies may be

required in order to determine the nature of the changes in cellular

activity that accompany reward. As was mentioned above, the rate

of drug administration is one critical variable in reward; route of

administration may be a second, and possibly related, variable.

Porrino and Kornetsky (this volume) described 2-deoxyglucose studies

demonstrating that the alterations in patterns of cerebral metabolism

induced by cocaine are partially dependent upon the route of

administration; they are not simply linear functions of dose, but are

343
affected in complex ways by the amount of drug administered. Given

that the patterns of activity revealed by 2-deoxyglucose represent

averages of activity over relatively long periods of time, it is clear

that the short-term alterations in cerebral function may be even more

complex. In this regard, it seems particularly important that more

attention be focused on physiological studies of function in defined

cell populations. None of the biochemical techniques currently

available has a time resolution that even begins to approach that of

physiological studies; given the rapid and rate-dependent temporal

dynamics of reward, physiological techniques may ultimately be

required to characterize the precise nature of the physiological events

that accompany reward.

As mentioned previously, a variety of evidence supports the hypothesis that blockade of dopamine uptake is an important aspect of the

reinforcement component of the actions of cocaine. However, this

implies as a necessary consequence that dopamine receptors are

involved in the reinforcing effects of cocaine. The modulation of

cocaine reinforcement by dopamine antagonists, as was discussed by a

number of speakers, supports this hypothesis. Goeders (this volume)

presented evidence that the self-administration of cocaine directly

into the medial prefrontal cortex could be blocked by sulpiride, the

selective D2 antagonist, but not by a D1 antagonist. Similar issues

were addressed by Woolverton and Kleven (this volume), and, like

many other aspects of cocaine action, the answers are not simple. In

general, dopamine D1 agonists are not self-administered, whereas D2

agonists such as piribedil are. D2 antagonists are effective antagonists of the rewarding effects of cocaine, but there is also some

evidence that a D1 antagonist (SCH 23390) can also reduce both the

rewarding and discriminative cue properties of cocaine (Woolverton

and Kleven, this volume; Koob and Hubner, this volume). The basis

for these interactions is obscure but may relate to multiple effects in

different brain regions. Alternatively, given the electrophysiological

evidence to support the hypothesis that D1 receptors may play a

permissive role in the elaboration of dopamine D2 responses (Carlson

et al. 1987; White 1987), a similar role for D1 receptors in behavior

might hold.

In relation to the role of dopamine in the reinforcing effects of

cocaine, Winger (this volume) discussed the possible use of dopamine

agonists and antagonists in the treatment of cocaine abuse. Unlike

the situation with opiate abusers, where both agonists (e.g., methadone) and antagonists (e.g., naltrexone) can potentially be used as

adjuncts to treatment, it seems less likely that either dopamine

344
agonists such as bromocriptine, or dopamlne antagonists such as the

neuroleptic drugs, are likely to be as effective in cocaine abusers.

Both agonists and antagonists lead to substantial toxicity with chronic

use, the agonists producing stereotypic behavior, and the antagonists

producing the adverse effects already well estabished with moat

neuroleptic drugs, such as tardive dyskinesias, etc. In addition,

dopamine antagonists typically increase rather than decrease

responding for cocaine injections and decrease responding for other

reinforcers such as food; both of these types of effects would clearly

be undesirable in a treatment for cocaine abuse. The basis for the

rate-increasing effects of dopamine antagonists is still unclear; this

has often been interpreted as being a compensatory response to the

diminished reinforcing effects of cocaine, but it could also represent

an antagonism of the rate-decreasing effects associated with higher

doses of cocaine. The relative difficulties that have been encountered in developing practical pharmacological strategies for reducing

cocaine self-administration may reflect a more central role for dopamine in reinforcement than for opiates. However, the relative

contributions of opiate and dopamine systems in reward, and the relationships between the two systems, are still topics of considerable

dispute (Wise and Bozarth 1984; Koob 1987).

Although interactions with dopaminergic systems may be the most

important component of the reinforcing effects of cocaine, the relative significance of local anesthetic effects remains unclear. Blood

concentrations of cocaine following recreational use reach levels at

which local anesthetic actions can be observed, but whether these

actions contribute to the rewarding effects of cocaine or represent

essentially a “side effect” of this drug is unclear. Local anesthetics

commonly induce unique patterns of spindling in olfactory and limbic

brain regions, and Post and Weiss (this volume) reported that such

effects can be observed in human studies with procaine as well, following intranasal use of cocaine. The precise significance of these

effects is unclear, but it was proposed that excessive activity in limbic regions may contribute initially to the euphoric effects of cocaine,

but, following pharmacologically induced kindling, may lead to adverse

effects such as decreased seizure threshold, panic attacks, etc.

Determining which pharmacological actions of cocaine contribute to

specific behavioral effects is clearly a difficult task. A potentially

useful tool for approaching these issues was suggested by George and

Goldberg (this volume). Studies of cocaine action in different mouse

strains have demonstrated markedly different behavioral responses,

345
particularly in terms of locomotor stimulant effects. By characterizing the physiological and biochemical differences between these

strains, it may be possible to dissociate various components of the

behavioral response to cocaine and to link them to their appropriate

neurobiological substrates. In a similar vein, interspecies comparisons

may also be of value in some cases, since some cocaine like drugs

(e.g., mazindol) are self-administered in some species but avoided by

others.

IDENTIFICATION OF BRAIN REGIONS RELATED TO COCAINE

ABUSE

The identification of substrates of drug action within the central

nervous system has been a recurring topic of interest in the drug

abuse field. Considerable evidence supports a role for the ventral

tegmental region and its projection areas in the rewarding effects of

not only opiate drugs and psychomotor stimulants, but in the effects

of other drugs such as nicotine and alcohol as well. Porrino and

Kometsky (this volume) demonstrated that low doses of cocaine (0.5

mg/kg) could significantly affect glucose utilization in the nucleus

accumbens and in the medial prefrontal cortex, both regions that

receive important dopamine projections from the ventral tegmental

area. At higher doses (e.g., 5 mg/kg), many brain regions showed

increases in activity. Whether or not sensitivity of the metabolism of

a given brain region to cocaine is necessarily related to reward

remains to be determined, but other drugs of abuse that act on dopamine systems, such as amphetamine and methylphenidate, show somewhat similar patterns of activity.

Another approach to identifying important brain sites of cocaine

action linked to reward is to combine techniques such as lesions or

local administration of drug to defined sites in the brain with

behavioral measures of pharmacological reward. In summarizing these

types of studies, Koob and Hubner (this volume) presented considerable evidence in support of a central role for the dopamine projections to the nucleus accumbens. Work presented by Koob and Hubner

(this volume) and Goeders (this volume) suggested that 6-

hydroxydopamine (6-OHDA) lesions of the nucleus accumbens, but not

the frontal cortex, significantly reduced or abolished the reinforcing

properties of cocaine. However, in view of the previous observations

by Goeders and Smith (1993) that animals will self-administer cocaine

into the medial prefrontal cortex, but not the nucleus accumbens,

these results are difficult to interpret. Perhaps some of these results

could be explained if the mesocortical system were involved in the

346
initiation aspects of reinforcement, whereas the accumbens is

involved in the actual reward process. Thus, lesions of the

mesocortical system would only disrupt one of perhaps several

initiation systems, whereas accumbens lesions would disrupt the

reward system directly. Nevertheless, there remain a number of

unanswered questions that can be addressed only through further

experiments.

ACUTE TOXICITY

A number of speakers addressed the issue of the toxic effects of

cocaine and the hazards associated with its use. Desplte the recent

occurrence of highly publicized deaths attributed to cocaine, it is

clear that, in general, the acute toxicity cannot be particularly high,

given the relatively large number of individuals who abuse cocaine.

Nevertheless, in specific cases in which individuals have conditions

that may predispose them to the toxic effects of cocaine, such as a

pseudocholinesterase deficiency or compromised cardiovascular function, acute toxic effects may present a serious risk. Wilkerson (this

volume) addressed some of the cardiovascular problems associated

with cocaine use, such as myocardial ischemia, ventricular tachycardias disruptions in local flow, etc., and presented work on a dog

model concerning the cardiovascular effects of cocaine. Although the

local anesthetic properties of cocaine would be expected to make it

an antiarrhythmic drug, both the effects upon the central nervous

system (increased sympathetic tone) and the ability of cocaine to

potentiate the effects of norepinephrine released at synapses are

arrhythmogenic. Determining which of these opposing effects predominate under various conditions and how they relate to the other

cardiovascular effects of cocaine will remain an important area of

work. Studies on dogs, reported by Wilkerson, clearly stressed the

importance of using unanesthetized subjects, since the use of anesthesia blunts the component of the response that is mediated via the

central nervous system.

Finally, Dow-Edwards (this volume) described some of the hazards

associated with use during pregnancy. Some of the adverse neurological consequences that have been reported in humans may be

related to pressor effects of cocaine (e.g., intracephalic hemorrhages),

but, in addition, there appear to be more subtle disruptions of development that are manifested only in rather general abnormalities (e.g.,

“jittery” behavior, decreased interaction, etc.) and remain only poorly

understood at this point. Rat studies (Dow-Edwards, this volume)

347
also have demonstrated lasting changes in both behavior and brain

neurochemistry following neonatal cocaine exposure.

EFFECTS OF CHRONIC DRUG USE

The chronic use of cocaine presents two very different and largely

unrelated issues. The first concerns changes in sensitivity to the

acute effects of cocaine with repeated administration (sensitization),

and the second concerns cumulative toxic effects associated with

repeated drug administration. While most drugs typically induce a

condition of reduced sensitivity upon chronic use (drug tolerance),

under certain conditions the repeated use of cocaine can lead to

enhanced sensitivity, which was originally described in terms of

behavioral sensitization. Repeated use of cocaine can lead to marked

potentiation of some of its behavioral effects, such as the increases

in motor activity and stereotypy that can be observed following the

appropriate dosing regime. Reith (this volume) addressed some of the

critical variables in terms of the protocols required to observe behavioral sensitization. In general, these effects are only seen with

repetitive administration of lower doses of cocaine, and only with

phasic administration of drug. Thus, when the drug was administered

chronically with an osmotic minipump, or when high (40 mg/kg) doses

were used, locomotor activity following acute cocaine administration

seemed to be reduced. However, it remains to be determined whether

this reduction in activity constitutes real tolerance or might be the

result of the development of “endstage” behaviors (e.g., extreme

stereotypy) that might present as reductions in motor activity.

Although the neurochemical bases for sensitization are unclear, some

of the biochemical data that were presented by Zahniser et al. (this

volume) provided interesting potential directions for research. In

these studies, single injections of cocaine were observed to facilitate

subsequent amphetamine-induced dopamine release from striatal brain

slices for at least 2 weeks. The functional relevance of amphetaminestimulated release is unclear, but the existence of such long-term

changes following treatment with a single dose of cocaine might well

provide a mechanism by which sensitization could arise. On the

other hand, data presented concerning neurochemical changes following 8 to 14 days of cocaine administration, under conditions that

caused behavioral sensitization, did not demonstrate any alterations in

dopamine receptors that would suggest potential mechanisms for this

process.

348
With regard to the toxic effects of chronic cocaine administration,

several types of studies were presented. Langner et al. (this volume)

addressed concerns regarding the potential for cocaine to induce the

formation of arteriosclerotic lesions, particularly with repetitive use.

Shuster et al. (this volume) described a type of hepatotoxicity that

can be observed in mice, particularly following high doses of cocaine.

This type of toxicity is considerably more difficult to elicit in rats

and may occur relatively rarely in humans, although this is difficult

to establish because in many cases the relationship of hepatotoxicity

to cocaine abuse may not have been suspected. In mice, a unique but

still only partially defined combination of conditions was required to

see more severe forms of toxicity (e.g., it did not occur in adrenalectomized mice). Further work may help to identify the conditions

under which such toxicity can occur and may provide insights as to

which drug abusers are particularly at risk.

Seiden and Kleven (this volume) discussed the neurotoxicity observed

with cocaine, amphetamine, and related compounds. Unlike amphetamine, which can release dopamine from the cytosolic pool, drugs that

induce release from the granular pool (cocaine, methylphenidate) do

not appear to induce neurotoxic changes. A proposed mechanism of

action involving the in vivo formation of toxic metabolites (e.g., 6-

OHDA) was presented, but it will take further work before this can

be established.

FUTURE DIRECTIONS

One of the major observations that evolved from this conference was

that rapid progress is now being made in characterizing the actions

of cocaine on the nervous system, and the behavioral manifestations

of such actions. Nonetheless, it is equally clear that the systems

upon which cocaine is thought to act are perhaps more complex than

would have been suspected based upon the knowledge of these

systems that we had even 10 years ago. Despite this complexity, it

seems likely that our level of understanding not only of drug abuse

per se, but also of the neuronal systems that underlie drug selfadministration behavior, should be markedly enhanced by studies now

in progress. What is perhaps not as readily apparent is that research

on drug abuse is likely to lead to insights not only into drug-seeking

and self-administration behaviors, but into a variety of aspects of

neurobiology that are only remotely related to drug abuse. In this

concluding section, some of these other aspects of this work will be

addressed.

349
There are several levels at which the study of cocaine action upon

the central nervous system can prove informative. First, cocaine

provides a way in which to perturb neurotransmission at serotonergic,

noradrenergic and dopaminergic synapses. The responses to such

changes in terms of receptor binding, transmitter uptake, etc., can

tell us much about the complex manner in which such systems are

regulated, and about ways that different transmitter systems interact.

Many useful antidepressant drugs share pharmacological properties

with cocaine, which is not a therapeutically useful antidepressant. By

comparing the actions of cocaine to such antidepressant drugs, it may

be possible to gain further insights into the specific actions that

ameliorate depression, and which changes are unrelated to this therapeutic action. In addition, characterizing the changes that occur

following long-term, high-dose use of cocaine, which results in

schizophreniclike behaviors in man, may provide a way of understanding some of the biological bases for the schizophrenic process

that it emulates.

To a psychobiologist interested in appetitive systems in the brain,

cocaine can serve as a remarkably powerful tool that can be

employed to define the neuronal circuits involved in pharmacological

reward. The observation has frequently been made that associating a

specific behavior with cocaine administration is perhaps one of the

easiest tasks that a rat can learn. Along these lines, cocaine may

ultimately prove to be of interest not only from the standpoint of

understanding the neuropharmacological basis for reward but also for

providing a key to understanding the learning process itself. Cocaine

administration clearly alters the central nervous system in such a

manner that events associated with cocaine administration quickly

become potent stimuli for eliciting the behaviors that result in drug

administration.

In this context, cocaine also provides a ready means of generating

positive reinforcement that can be studied with a variety of neurobiological techniques. As one example, cocaine has potential use in

defining the physiology of reward, i.e., in determining which patterns

of neuronal activity are most closely associated with the behavioral

state of positive reinforcement. This in turn may provide a means of

understanding some of the more unusual behavioral aspects of reward,

such as why a rapid onset of drug action is preferable to a slow

onset. In this context, it will also be important to differentiate

between various kinds of cocaine administration, such as passive vs.

self-administration, and intraperitoneal vs. intravenous administration.

By determining how cocaine changes neuronal activity under a variety

350
of conditions, we may gain insight into how other drugs of abuse

act and may ultimately develop more rational methods of treating

substance abuse.

From a basic science standpoint, cocaine also provides an invaluable

tool for characterizing the high-affinity uptake process for monoamine transmitters. Studies utilizing the binding of radiolabeled

cocaine and other inhibitors of uptake have led to the hypothesis

that they inhibit uptake by binding to an allosteric regulator of

uptake, since dopamine is not an effective competitor for such

binding. Further work in this area is likely to lead to a better understanding of the uptake process and its possible regulation by endogenous modulators as well. Studies of other novel aspects of cellular

function, such as the calcium-independent release of transmitter from

somatodendritic sites in the ventral tegmental area and substantia

nigra, have to some extent evolved out of the study of the actions of

drugs such as cocaine.

Finally, the study of cocaine might lead to a better understanding of

local anesthetic actions in the central nervous system and in the

heart. Many of the actions of cocaine remain obscure. Why should a

local anesthetic drug cause seizures? Which brain regions are

involved? What cellular alterations lead to such excitability, and how

can this lead to kindling following repetitive drug administration? In

addition, it is unclear how a drug such as cocaine (or other local

anesthetics) not only can have antiarrhythmic effects but also can

increase the probability of arrhythmias, particularly in lower dose

ranges. What we learn from cocaine in this regard will considerably

enhance our understanding of the basic biology of these systems.

There clearly remain a number of highly important yet unresolved

issues concerning the pharmacological effects of cocaine. In

answering earlier questions concerning the actions of cocaine, different and often more complex questions are raised in their stead.

However, the results presented in this monograph provide manifest

evidence of the rapid evolution of our understanding of the actions

of cocaine. The primary intent of this introduction has been to

focus attention upon specific areas in which considerable progress has

been made in the last few years and, in addition, to suggest areas in

which directed efforts might be particularly successful in making

further advances. The hope is that a better understanding of the

mechanism by which cocaine acts may suggest better ways of treating

cocaine abuse. Nevertheless, it is also clear that if a better understanding of the mechanisms underlying drug self-administration can be

351
achieved, this will necessarily involve significant advances in our

understanding of the psychobiological bases for the regulation of

appetitive behaviors.

REFERENCES

Brodie, M.S., and Dunwiddie, T.V. The effects of cocaine on ventral

tegmental area spontaneous activity in vitro: Interactions with

dopamine, sulpiride and choiecystokinin. Paper presented at the

16th Annual Meeting of the Society for Neuroscience, Washington,

DC, November 1986.

Carlson, J.H.; Bergstrom, D.A.; and Walters, J.R. Stimulation of both

D1 and D2 dopamine receptors appears necessary for full expression

of postsynaptic effects of dopamine agonists: A neurophysiological

study. Brain Res 400:205-218, 1987.

Goeders, N.E., and Smith, J.E. Cortical dopaminergic involvement in

cocaine reinforcement. Science 221:773-775, 1983.

Koob, G.F.; Le, H.T.; and Creese, I. D-1 receptor antagonist

SCH 23390 increases cocaine self-administration in the rat.

Neurosci Lett 79:315-321, 1987.

Pitts, D.K., and Marwah, J. Electrophysiologicai effects of cocaine

on central monoaminergic neurons. Eur J Pharmacol 131:95-98,

1986.

Ritz, M.C.; Lamb, R.J.; Goldberg, S.R.; and Kuhar, M.J. Cocaine

receptors on dopamine transporters are related to self-administration of cocaine. Science 237:1219-1223, 1987.

White, F.J. D-1 dopamine receptor stimulation enables the

inhibition of nucleus accumbens neurons by a D-2 receptor agonist.

Eur J Pharmacol 135:101-105, 1987.

White, F.J. Electrophysioiogicai effects of cocaine in the

mesoaccumbens and mesocortical dopamine systems. Abstr Soc

Neurosci 11:828, 1985.

Wise, R.A., and Bozarth, M.A. Brain reward circuitry: Four circuit

elements “wired” in apparent series. Brain Res Bull 12:203-208,

1984.

ACKNOWLEDGMENTS

This work was supported by U.S. Public Health Service grant DA

02702 from the National Institute on Drug Abuse and by the Veterans

Administration Medical Research Service.

352
AUTHOR

Thomas V. Dunwiddie, Ph.D.

Department of Pharmacology

University of Colorado Health Sciences Center

Denver, CO 80262

and

Veterans Administration Medical Center

Denver, CO 80220

353
monograph series

While limited supplies last, single copies of the monographs may be

obtained free of charge from the National Clearinghouse for Alcohol

and Drug Information (NCADI). Please contact NCADI also for information about availability of coming issues and other publications of

the National Institute on Drug Abuse relevant to drug abuse research.

Additional copies may be purchased from the U.S. Government Printing Office (GPO) and/or the National Technical Information Service

(NTIS) as indicated. NTIS prices are for paper copy. Microfiche

copies, at $6.95, are also available from NTIS. Prices from either

source are subject to change.

Addresses are:

NCADI

National Clearinghouse for Alcohol and Drug Information

P.O Box 2345

Rockville, MD 20852

GPO

Superintendent of Documents

U.S. Government Printing Office

Washington, DC 20402

NTIS

National Technical Information

Service

U.S. Department of Commerce

Springfield, VA 22161

For information on availability of NIDA Research Monographs 1-24

(1975-1979) and others not listed, write to NIDA Office for Research

Communications, Room 10A-54, 5600 Fishers Lane, Rockville,

MD 20857.

354
25 BEHAVIORAL ANALYSIS AND TREATMENT OF SUBSTANCE

ABUSE. Norman A. Krasnegor, Ph.D., ed.

GPO out of stock NCADI out of stock

NTIS PB #86-112428 $24.95

26 THE BEHAVIORAL ASPECTS OF SMOKING. Norman A. Krasnegor,

Ph.D., ed. (Reprint from 1979 Surgeon General’s Report on Smoking

and Health.)

GPO out of stock NTIS PB #80-118755 $18.95

30 THEORIES ON DRUG ABUSE: SELECTED CONTEMPORARY

PERSPECTIVES. Dan J. Lettieri, Ph.D.; Mollie Sayers; and Helen W.

Pearson, eds.

GPO Stock #017-024-00997-1 $10 NCADI out of stock

Not available from NTIS

31 MARIJUANA RESEARCH FINDINGS: 1980. Robert C. Petersen,

Ph.D., ed.

GPO out of stock NTIS PB #80-215171 $24.95

32 GC/MS ASSAYS FOR ABUSED DRUGS IN BODY FLUIDS. Rodger

L. Foltz, Ph.D.; Allison F. Fentiman, Jr., Ph.D.; and Ruth B. Foltz.

GPO out of stock NTIS PB #81-133746 $24.95

36 NEW APPROACHES TO TREATMENT OF CHRONIC PAIN: A REVIEW OF MULTIDISCIPLINARY PAIN CLINICS AND PAIN CENTERS.

Lorenz KY. Ng, M.D., ed.

GPO out of stock NCADI out of stock

NTIS PB #81-240913 $24.95

37 BEHAVIORAL PHARMACOLOGY OF HUMAN DRUG DEPENDENCE.

Travis Thompson, Ph.D., and Chris E. Johanson, Ph.D., eds.

GPO out of stock NCADI out of stock

NTIS PB #82-136961 $30.95

38 DRUG ABUSE AND THE AMERICAN ADOLESCENT. Dan J.

Lettieri, Ph.D., and Jacqueline P. Ludford, M.S., eds. A RAUS Review

Report.

GPO out of stock NCADI out of stock

NTIS PB #82-148198 $18.95

355
40 ADOLESCENT MARIJUANA ABUSERS AND THEIR FAMILIES.

Herbert Hendin, M.D.; Ann Pollinger, Ph.D.; Richard Ulman, Ph.D.; and

Arthur Carr, Ph.D.

GPO out of stock NCADI out of stock

NTIS PB #82-133117 $18.95

42 THE ANALYSIS OF CANNABINOIDS IN BIOLOGICAL FLUIDS.

Richard L. Hawks, Ph.D., ed.

GPO out of stock NTIS PB #83-136044 $18.95

44 MARIJUANA EFFECTS ON THE ENDOCRINE AND REPRODUCTlVE

SYSTEMS. Monique C. Braude, Ph.D., and Jacqueline P. Ludford,

M.S., eds. A RAUS Review Report.

GPO out of stock NCADI out of stock

NTIS PB #85-150563/AS $18.95

45 CONTEMPORARY RESEARCH IN PAIN AND ANALGESIA, 1983.

Roger M. Brown, Ph.D.; Theodore M. Pinkert, M.D., J.D.; and

Jacqueline P. Ludford, M.S., eds. A RAUS Review Report.

GPO out of stock NCADI out of stock

NTIS PB #84-184670/AS $13.95

46 BEHAVIORAL INTERVENTION TECHNIQUES IN DRUG ABUSE

TREATMENT. John Grabowski, Ph.D.; Maxine L. Stitzer, Ph.D.; and

Jack E. Henningfield, Ph.D., eds.

GPO out of stock NCADI out of stock

NTIS PB #84-184688/AS $18.95

47 PREVENTING ADOLESCENT DRUG ABUSE: INTERVENTION

STRATEGIES. Thomas J. Glynn, Ph.D.; Carl G. Leukefeld, D.S.W.; and

Jacqueline P. Ludford, M.S., eds. A RAUS Review Report.

GPO Stock #017-024-01180-1 NTIS PB #85-159663/AS $24.95

$5.50

48 MEASUREMENT IN THE ANALYSIS AND TREATMENT OF SMOKING BEHAVIOR. John Grabowski, Ph.D., and Catherine S. Bell, M.S.,

eds.

GPO Stock #‘017-024-01181-9 NCADI out of stock

$4.50 NTIS PB #84-145184/AS $18.95

50 COCAINE: PHARMACOLOGY, EFFECTS, AND TREATMENT OF

ABUSE. John Grabowski, Ph.D., ed.

GPO Stock #017-024-01214-9 $4 NTIS PB #85-150381/AS $18.95

356
51 DRUG ABUSE TREATMENT EVALUATION: STRATEGIES, PROGRESS, AND PROSPECTS. Frank M. Tims, Ph.D., ed.

GPO Stock #017-024-01218-1 NTIS PB #85-150365/AS $18.95

$4.50

52 TESTING DRUGS FOR PHYSICAL DEPENDENCE POTENTIAL AND

ABUSE LIABILlTY. Joseph V. Brady, Ph.D., and Scott E. Lukas,

Ph.D., eds.

GPO Stock #017-024-01204-1

$4.25

NTIS PB #85-150373/AS $18.95

53 PHARMACOLOGICAL ADJUNCTS IN SMOKING CESSATION. John

Grabowski, Ph.D., and Sharon M. Hall, Ph.D., eds.

GPO Stock #017-024-01266-1 NCADI out of stock

$3.50

54 MECHANISMS OF TOLERANCE AND DEPENDENCE. Charles Wm.

Sharp, Ph.D., ed.

GPO out of stock

56 ETIOLOGY OF DRUG ABUSE: IMPLICATIONS FOR PREVENTION.

Coryl LaRue Jones, Ph.D., and Robert J. Battjes, D.S.W., eds.

GPO Stock #017-024-01250-5 $6.50

57 SELF-REPORT METHODS OF ESTIMATING DRUG USE: MEETING

CURRENT CHALLENGES TO VALIDITY. Beatrice A. Rouse, Ph.D.;

Nicholas J. Kozel, M.S.; and Louise G. Richards, Ph.D., eds.

GPO Stock #017-024-01246-7 $4.25

58 PROGRESS IN THE DEVELOPMENT OF COST-EFFECTIVE TREATMENT FOR DRUG ABUSERS. Rebecca S. Ashery, D.S.W., ed.

GPO Stock #017-024-01247-5 $4.25

59 CURRENT RESEARCH ON THE CONSEQUENCES OF MATERNAL

DRUG ABUSE. Theodore M. Pinkert M.D., J.D., ed.

GPO Stock #017-024-01249-1 $2.50

60 PRENATAL DRUG EXPOSURE: KINETICS AND DYNAMICS.

C. Nora Chiang, Ph.D., and Charles C. Lee, Ph.D., eds.

GPO Stock #017-024-01257-2 $3.50

357
61 COCAINE USE IN AMERICA: EPIDEMIOLOGIC AND CLINICAL

PERSPECTIVES. Nicholas J. Kozel, M.S., and Edgar H. Adams, M.S.,

eds.

GPO Stock #017-024-01256-1 $5

62 NEUROSCIENCE METHODS IN DRUG ABUSE RESEARCH. Roger

M. Brown, Ph.D., and David P. Friedman, Ph.D., eds.

GPO Stock #017-024-01266-2 $3.50

63 PREVENTION RESEARCH: DETERRING DRUG ABUSE AMONG

CHILDREN AND ADOLESCENTS. Catherine S. Bell, M.S., and Robert

J. Battjes, D.S.W., eds.

GPO Stock #017-024-01263-7 $5.50

64 PHENCYCLIDINE: AN UPDATE. Doris H. Clouet, Ph.D., ed.

GPO Stock #017-024-01281-5 $6.50

65 WOMEN AND DRUGS: A NEW ERA FOR RESEARCH. Barbara A.

Ray, Ph.D., and Monique C. Braude, Ph.D., eds.

GPO Stock #017-024-01283-1 $3.25

66 GENETIC AND BIOLOGICAL MARKERS IN DRUG ABUSE AND

ALCOHOLISM. Monique C. Braude, Ph.D., and Helen M. Chao, Ph.D.,

eds.

GPO Stock #017-024-01291-2

$3.50

NCADI out of stock

68 STRATEGIES FOR RESEARCH ON THE INTERACTIONS OF DRUGS

OF ABUSE. Monique C. Braude, Ph.D., and Harold M. Ginzburg, M.D.,

J.D., M.P.H., eds.

GPO Stock #017-024-01296-3

$6.50

NCADI out of stock

69 OPIOID PEPTIDES: MEDICINAL CHEMISTRY. Rao S. Rapaka,

Ph.D.; Gene Barnett, Ph.D.; and Richard L. Hawks, Ph.D., eds.

GPO Stock #017-024-01297-1 $11

70 OPIOID PEPTIDES: MOLECULAR PHARMACOLOGY, BIOSYNTHESIS, AND ANALYSIS. Rao S. Rapaka, Ph.D., and Richard L.

Hawks, Ph.D., eds.

GPO Stock #017-024-01298-0 $12

358
71 OPIATE RECEPTOR SUBTYPES AND BRAIN FUNCTION. Roger M.

Brown, Ph.D.: Doris H. Clouet, Ph.D.; and David P. Friedman, Ph.D.,

eds.

GPO Stock #017-024-01303-0 $6

72 RELAPSE AND RECOVERY IN DRUG ABUSE. Frank M. Tims,

Ph.D., and Carl G. Leukefeld, D.S.W., eds.

GPO Stock #017-024-01302-1 $6

73 URINE TESTING FOR DRUGS OF ABUSE. Richard L. Hawks,

Ph.D., and C. Nora Chiang, Ph.D., eds.

GPO Stock #017-024-01313-7 $3.75

74 NEUROBIOLOGY OF BEHAVIORAL CONTROL IN DRUG ABUSE.

Stephen I. Szara, M.D., D.Sc., ed.

GPO Stock #017-024-01314-5 $3.75

75 PROGRESS IN OPIOID RESEARCH. PROCEEDINGS OF THE 1986

INTERNATIONAL NARCOTICS RESEARCH CONFERENCE. John W.

Holaday, Ph.D.; Ping-Yee Law, Ph.D.; and Albert Herz, M.D., eds.

GPO Stock #017-024-0315-3 $21 NCADI out of stock

76 PROBLEMS OF DRUG DEPENDENCE, 1986. PROCEEDINGS OF

THE 48TH ANNUAL SCIENTIFIC MEETING, THE COMMlTTEE ON

PROBLEMS OF DRUG DEPENDENCE, INC. Louis S. Harris, Ph.D., ed.

GPO Stock #017-024-01316-1 $16 NCADI out of stock

NTIS PB #88-208111/AS $44.95

77 ADOLESCENT DRUG ABUSE: ANALYSES OF TREATMENT

RESEARCH. Elizabeth R. Rahdert, Ph.D., and John Grabowski, Ph.D.,

eds.

GPO Stock #017-024-01348-0 $4

78 THE ROLE OF NEUROPLASTICITY IN THE RESPONSE TO DRUGS.

David P. Friedman, Ph.D., and Doris H. Clouet, Ph.D., eds.

GPO Stock #017-024-01330-7 $6

79 STRUCTURE-ACTIVITY RELATIONSHIPS OF THE CANNABINOIDS.

Rao S. Rapaka, Ph.D., and Alexandros Makriyannis, Ph.D., eds.

GPO Stock #017-024-01331-5 $6

359
80 NEEDLE SHARING AMONG INTRAVENOUS DRUG ABUSERS:

NATIONAL AND INTERNATIONAL PERSPECTIVES. Robert J. Battles,

D.S.W., and Roy W. Pickens, Ph.D., eds.

GPO Stock #017-024-01345-5 NTIS PB #88-236138/AS $25.95

$5.50

81 PROBLEMS OF DRUG DEPENDENCE, 1987. PROCEEDINGS OF

THE 49TH ANNUAL SCIENTIFIC MEETING, THE COMMlTTEE ON

PROBLEMS OF DRUG DEPENDENCE, INC. Louis S. Harris, Ph.D., ed.

GPO Stock #017-024-01354-4 $17

82 OPIOIDS IN THE HIPPOCAMPUS. Jacqueline F. McGinty, Ph.D.,

and David P. Friedman, Ph.D., eds.

GPO Stock #017-024-01344-7 $4.25

63 HEALTH HAZARDS OF NlTRlTE INHALANTS. Harry W. Haverkos,

M.D., and John A. Dougherty, Ph.D., eds.

GPO Stock #017-024-01351-0 $3.25

64 LEARNING FACTORS IN SUBSTANCE ABUSE. Barbara A. Ray,

Ph.D., ed.

GPO Stock #017-024-01353-6 $6

86 COMPULSORY TREATMENT OF DRUG ABUSE: RESEARCH AND

CLINICAL PRACTICE. Carl G. Leukefeld, D.S.W., and Frank M. Tims,

Ph.D., eds.

IN PRESS

85 EPIDEMIOLOGY OF INHALANT ABUSE: AN UPDATE. Raquel A.

Crider, Ph.D., and Beatrice A. Rouse, Ph.D., eds.

89 BIOLOGICAL VULNERABILlTY TO DRUG ABUSE. Roy W.

Pickens, Ph.D., and Dace S. Svikis, B.A., eds.

360

U.S GOVERNMENT PRINTING OFFICE 1989-248-701/00726
DHHS Publication No. (ADM)59-1585

Alcohol, Drug Abuse, and Mental Health Administration

Printed 1988, Reprinted 1989