Testing Drugs for

Physical

Dependence

Potential and

Abuse Liability

DEPARTMENT OF HEALTH AND HUMAN SERVICES

Public Health Service

Alcohol. Drug Abuse, and Mental Health Administration
Testing Drugs for Physical

Dependence Potential and

Abuse Liability

The Committee on Problems of

Drug Dependence, Inc.

Editors:

Joseph V. Brady, Ph.D.

The Johns Hopkins University School of Medicine

Scott E. Lukas, Ph.D.

Addiction Research Center

National Institute on Drug Abuse

NIDA Research Monograph 52

1984

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, Maryland 20857

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

Washington, D.C. 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 Institute

Troy. New York

RICHARD E. BELLEVILLE, Ph.D.

NB Associates, Health Sciences

Rockville. Maryland

KARST J. BESTEMAN

Alcohol and Drug Problems Association

of North America

Washington, D.C

GILBERT J. BOTVIN, Ph.D.

Cornell University 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

SIDNEY COHEN, M.D.

Los Angeles, California

MARY L. JACOBSON

National Federation of Parents for

Drug-Free Youth

Omaha, Nebraska

REESE T. JONES, M.D.

Langley Porter Neuropsychratric Institute

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

JEAN PAUL SMITH, Ph.D.

Acting Associate Director for Science, NIDA

Acting Editor

Parklawn Building. 5600 Fishers Lane, Rockville, Maryland 20857
ACKNOWLEDGMENTS

The material in this monograph was developed and reviewed by

members of The Committee on Problems of Drug Dependence, Inc. The

senior editor, Dr. Joseph V. Brady, is Professor of Behavioral

Biology and Professor of Neuroscience at the Johns Hopkins

University School of Medicine, Baltimore. Dr. Scott E. Lukas

organized the document, reviewed the relevant research literature,

and wrote substantial portions of the monograph. Until March 1984

Dr. Lukas was a staff fellow with the National Institute on Drug

Abuse, Addiction Research Center. He is currently Assistant

Professor of Psychiatry (Pharmacology) at the Harvard Medical

School and the McLean Hospital Alcohol and Drug Abuse Research

Center.

PUBLIC DOMAIN NOTICE

All material appearing in this volume is in the public domain and

may be reproduced or copied without permission from the Institute

or the authors. Citation of the source is appreciated.

The U.S. Government does not endorse or favor any specific

commercial product or commodity. Trade or proprietary names

appearing in this publication are used only because they are

considered essential in the context of the studies reported herein.

Library of Congress catalog card number 84-601076

DHHS Publication No. (ADM)90-1332

Printed 1984, Reprinted 1987, 1990

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.
MEMBERS

Committee on Problems of Drug Dependence

Joseph V. Brady, Chair

Martin Adler

Sidney Archer

William Beaver

Richard Bonnie

Theodore Cicero

Marian Fischman

Charles Gorodetzky

Roland Griffiths

Leo Hollister

Jerome Jaffe

Donald Jasinski

Harold Kalant

William Martin

Charles O'Brien

C. R. Schuster, Jr.

A. E. Takemori

Members

Board of Directors

E. L. Way, Chairman

Raymound W. Houde

Keith F. Killam

Everette May

Jack Mendelson

Beny J. Primm

Lee N. Robins

Edward C. Senay

James Woods

Executive Secretary

Joseph Cochin

Liaison Participants

Louis S. Harris

Arthur E. Jacobson

Howard McClain

Heinz Sorer

Edward C. Tocus
FOREWORD

The National Institute on Drug Abuse is pleased to publish as a

NIDA Research Monograph this updated state-of-the-art review of the

procedures utilized for testing drugs for their physical dependence

potential and abuse liability. NIDA has a long history of working

with the Committee on Problems of Drug Dependence (CPDD) to attain

our mutual objective in broadening the knowledge base in this area.

It has been 11 years since the previous review by the Committee on

Problems of Drug Dependence, and 9 years since the publication by

WHO of Evaluation of Dependence Liability and Dependence Potential

of Drugs. During this period, the field has advanced

significantly, particularly as new classes of compounds have been

included within the range of those known to result in human

dependence. In addition, this review represents a major conceptual

clarification in distinguishing between testing of physical

dependence potential and abuse liability. The rationale this

distinction is well described in Chapter II.

The ultimate objective is to develop and refine methods that will

allow for the prediction of the human dependence or addiction

potential of a compound. Too often, the problems of human drug

dependence have been attributed to the physical dependence

potential of a drug. The use of the word dependence in two very

different senses has contributed its share of confusion.

effort to maintain clarity, the authors of the manuscript have

consistently referred to measurement of physical dependence

liability so as not to confuse that concept with the clinical

syndrome of drug dependence. This is particularly important since

increasingly the evidence supports the view that the problems of

human drug dependence are more closely related to the abuse

liability of a drug than to its physical dependence potential.

In the Prefatory note that follows, the importance of these 

procedures in making national and international regulatory

decisions has been emphasized. Not to be over looked is the fact 

that the laboratory work necessary for the development of these

testing procedures has also contributed to the identification and

characterization of novel compounds and the elucidation of 

fundamental biobehaviorial mechanisms of drug action. The recent Text

v
growth of knowledge of the neural mechanisms involved in drug abuse

has been explosive. Along with this has come a rapidly developing

ability to synthesize new drugs with markedly greater, specificity,

increasing the requirement for refined methods of assessing abuse

liability. Future development in the area may be expected to focus

on the role of drug interactions, the role of environmental and

social factors and, perhaps most importantly, the role of

individual genetic differences in the liability to abuse of drugs.

The CPDD can be expected to continue to play an important

leadership role in providing a forum for the integration of the

many scientific, social and legal issues involved in coping with

the problems of human drug dependence.

Jean Paul Smith, Ph.D.

Acting Associate Director for Science

National Institute on Drug Abuse

vi
PREFACE

For many years, the Department of Health and Human Services (DHHS)

has had a special interest in developing methodologies for

assessing abuse liability of drugs, irrespective of their known or

potential therapeutic usefulness. More recently, additional

interest in this area has grown as a result of the authorities

granted to the Secretary of DHHS in the Comprehensive Drug Abuse

Prevention and Control Act (P.L. 91-513) and the Psychotropic

Substances Act of 1978 (P.L. 95-633). These statutes provide the

Secretary with the authority to determine the abuse liability of

drugs and make scheduling recommendations with regard to the need

for domestic and international controls in an effort to prevent or

reduce the abuse. Thus, efforts to expand our knowledge to make

informed scheduling decisions has received support as part of the

overall drug abuse prevention effort.

Within the DHHS, the National Institute on Drug Abuse (NIDA) is

authorized by the Secretary to conduct the necessary studies for

the proper evaluation of the abuse liability of drugs and other

substances and to evaluate the nature and extent of associated

health risks. Pursuant to these responsibilities, NIDA supports

various intramural and extramural research efforts both to

determine the abuse liability of new drugs and to develop methods

for making more specific determinations about new classes of

compounds. These efforts have been significantly enhanced by close

collaboration with the Committee on Problems of Drug Dependence

and the National Institute of Arthritis, Diabetes, and Digestive

and Kidney Diseases.

It is hoped that this scholarly and comprehensive review of the

existing methodologies will be of benefit to both the public and

private sector domestically, as well as internationally. In

addition to being a unique reference, it can serve as a guide for

future drug abuse liability screening initiatives and a foundation

on which newer and more specific methodologies can be developed.

The monograph may be of assistance to other countries interested in

incorporating some abuse liability testing as a requisite for drug

registration and assist the World Health Organization (WHO) in its

efforts to identify the range of existing parameters available for

assessing the abuse liability of drugs required of them by

international treaties.

James R. Cooper, M.D.

Associate Director for Medical

and International Affairs

National Institute on Drug Abuse

vii

CONTENTS

Foreword

Jean Paul Smith

Preface

James R. Cooper

I. Introduction

References

II. Concepts and Definitions

References

III. General Methodological Considerations

IV. Specific Drugs and Drug Classes

A. General Considerations

B. Opioids

C. CNS Depressants: Sedative/Hypnotics,

Anesthetics, Anxiolytics, and Antihistamines

D. CNS Stimulants: Anorectics, Local

Anesthetics, and Antidepressants

E. Hallucinogens and Anticholinergics

F.

G .

Cannabinoids

Dissociative Anesthetics

V. Animal Testing Procedures

A. General Methods

1. Procedures for Characterization

of Drug Effects

a. Acute Drug Effects

i. Spontaneous Locomotor Activity

ii. Forced Motor Activity

iii. Electroencephalographic

(EEG) Activity

iv. Behavioral Performance

v. Binding Assays

2. Procedures for Assessing Physical

Dependence Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Administration

3. Procedures of Assessing Abuse Liability

a. Drug Self-Administration

i. Progressive Ratio Procedures

ii. Response Rate Analysis

iii. Choice Procedures

iv. Concurrent Schedules

b. Drug Discrimination Procedures

c. Behavioral Toxicity

References

v

vii

1

2

3

6

8

10

10

10

1 1

12

13

13

13

15

15

15

15

15

16

16

18

19

19

19

20

20

21

21

22

22

22

23

24

ix
B. Procedures for Assessing Opioids

1. Characterization of Opioid Effects

a. Acute Drug Effects

i. Analgesia

ii. Spontaneous Locomotor Activity

iii. Electroencephalographic

(EEG) Activity

iv. Behavioral Performance

v. Seizure Threshold

vi. Spinal Dog Preparation

vii. Gastrointestinal Motility

viii. Body Temperature

ix. Pupillary Effects

x. In Vitro Procedures

xi. Binding Assays

2. Opioid Physical Dependence Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Administration

i. Primary Physical Dependence

ii. Single Dose Substitution

3. Opioid Abuse Liability

a. Drug Self-Administration

i. Continuous Self-Administration

ii. Substitution

iii. Reinforcing Efficacy Analysis

iv. Tolerance and Physical

Dependence

b. Drug Discrimination

c. Behavioral Toxicity

References

C. Procedures for Assessing CNS Depressants:

Sedative/Hypnotics, Anesthetics, Anxiolytics,

and Antihistamines

29

29

2 9

2 9

30

30

3 1

3 1

3 2

3 2

33

34

34

34

3 5

35

36

3 7

3 8

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38

39

39

39

40

40

41

41

4 8

4 8

4 8

48

4 8

4 8

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49

50

50

5 1

52

53

53

53

53

54

54

1.

2.

3.

Characterization of CNS Depressant Effects

a. Acute Drug Effects

i. Sleep Induction

ii. Spontaneous Locomotor Activity

iii. Electroencephalographic

(EEG) Activity

iv. Behavioral Performance

v. Binding Assays

CNS Depressant Physical Dependence

Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Administration

i. Primary Physical Dependence

ii. Single Dose Substitution

CNS Depressant Abuse Liability

a. Drug Self-Administration

i. Continuous Self-Administration

ii. Substitution

iii. Intragastric, oral, and

Inhalation Self-Administration

x
iv. Reinforcing Efficacy Analysis

v. Tolerance and Physical

Dependence

b. Drug Discrimination

c .

References

Behavioral Toxicity

D. Procedures for Assessing CNS Stimulants:

Anorectics. Local Anesthetics, and Antidepressants

1. Characterization of CNS Stimulant Effects

a. Acute Drug Effects

i. Spontaneous Locomotor Activity

56

56

56

57

58

64

ii. Electroencephalographic

(EEG) Activity

iii. Behavioral Performance

iv. Anorexia

v. Intracranial Electrical

Self-Stimulation

2. CNS Stimulant Physical Dependence

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Potential

Administration

3. CNS Stimulant Abuse Liability

a . Drug Self-Administration

i. Continuous Self-Administration

ii. Substitution

iii. Reinforcing Efficacy Analysis

b. Drug Discrimination

c.

References

Behavioral Toxicity

E. Procedures of Assessing Cannabinoids

1. Characterization of Cannabinoid Effects

a. Acute Drug Effects

i. Spontaneous Locomotor Activity

ii. Analgesic Activity

iii. "Hallucinatory" Behavior

iv. Dog Ataxia Test

v. Schedule-Controlled Behavior

vi. Electroencephalographic

(EEG) Activity

2. Cannabinoid Physical Dependence Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Administration

3. Cannabinoid Abuse Liability

a. Drug Self-Administration

i. Continuous Self-Administration

ii. Substitution

b. Drug Discrimination

c .

References

Behavioral Toxicity

F. Procedures for Assessing Hallucinogens and

Anticholinergics

1. Characterization of Hallucinogen and

Anticholinergic Effects

a. Acute Drug Effects

64

64

64

64

64

65

65

65

66

66

66

66

66

67

66

67

68

71

71

71

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73

74

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75

75

76

81

81

81

xi
i. Abnormal Behaviors

ii. Spinal Dog Preparation

iii. Schedule-Controlled Behavior

iv. Electroencephalographic

(EEG) Activity

v. Binding Assays

2. Hallucinogen and Anticholinergic

Physical Dependence Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug Effects

3. Hallucinogen and Anticholinergic

Abuse Liability

a. Drug Self-Administration

i. Continuous Self-Administration

ii. Substitution

b. Drug Discrimination

c. Behavioral Toxicity

References

G. Procedures for Assessing Dissociative

Anesthetics

1. Characterization of Dissociative

Anesthetic Effects

a. Acute Drug Effects

i. Spontaneous Locomotor Activity

ii. Forced Motor Activity

iii. Electroencephalographic

(EEG) Activity

iv. Behavioral Performance

v. Binding Assays

2. Dissociative Anesthetic Physical

Dependence Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Administration

3. Dissociative Anesthetic Abuse Liability

a. Drug Self-Administration

b. Drug Discrimination

c. Behavioral Toxicity

References

VI. Human Testing Procedures

A. General Methods

81

81

82

82

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83

83

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88

88

8 8

88

8 8

8 8

8 9

8 9

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89

9 0

90

90

9 1

9 1

9 4

9 4

9 4

9 4

9 4

9 8

99

9 9

100

100

100

101

1.

2.

3.

Procedures for Characterization of

Drug Effects

a. Acute Drug Effects

i. Analgesia

ii. Electroencephalographic

(EEG) Activity and Sleep

Procedures for Assessing Physical

Dependence Potential

a. Tolerance to Drug Effects

b. Withdrawal from Chronic Drug

Administration

Procedures for Assessing Abuse Liability

a. Drug Self-Administration

b. Self-Report of Drug Effects

xii
c. Behavioral Toxicity 102

References 103

B. Procedure for Assessing Opioids 110

1. Characterization of Opioid Drug Effects 110

a. Acute Drug Effects 110

i. Analgesia 110

ii. Electroencephalographic 112

(EEG) Activity and Sleep

iii. Gastrointestinal Motility 113

Cough 114

2. Opioid Physical Dependence Potential 114

a. Tolerance to Drug Effects 114

b. Withdrawal from Chronic Drug 115

Administration

3. Opioid Abuse Liability 116

a. Drug Self-Administration 116

b. Self-Report of Drug Effects 117

c. Behavioral Toxicity 118

References 119

C. Procedures for Assessing CNS Depressants: 123

Sedative/Hypnotics, Anesthetics, Anxiolytics,

and Antihistamines

1. Characterization of CNS Depressant Effects 123

a. Acute Drug Effects 123

i. Anxiolytic and Sedation 123

ii. Electroencephalographic 123

(EEG) Activity and Sleep

2. CNS Depressant Physical Dependence 124

Potential

a. Tolerance to Drug Effects 124

b. Withdrawal from Chronic Drug 125

Administration

3. CNS Depressant Abuse Liability 126

a. Drug Self-Administration 126

b. Self-Report of Drug Effects 126

c. Behavioral Toxicity 127

References 127

D. Procedures for Assessing CNS Stimulants: 130

Anorectics, Local Anesthetics, and

Antidepressants

1. Characterization of CNS Stimulant Effects 130

a. Acute Drug Effects 130

i. Electroencephalographic 130

(EEG) Activity and Sleep

ii. Appetite 130

2. CNS stimulant Physical Dependence 130

Potential

a. Tolerance to Drug Effects 130

b. Withdrawal from Chronic Drug 131

Administration

3. CNS Stimulant Abuse Liability 131

a. Drug Self-Administration 131

b. Self-Report of Drug Effects 132

c. Behavioral Toxicity 132

References 133

xiii
E. Procedures for Assessing Cannabinoids 135

1. Characterization of Cannabinoid Effects 135

a. Acute Drug Effects 135

i. Analgesia 135

ii. Electroencephalographic 135

(EEG) Activity and Sleep

2. Cannabinoid Physical Dependence Potential 136

a. Tolerance to Drug Effects 136

b. Withdrawal from Chronic Drug 136

Administration

3. Cannabinoid Abuse Liability 136

a. Drug Self-Administration 136

b. Self-Report of Drug Effects 137

c. Behavioral Toxicity 137

References 138

F. Procedures for Assesing Hallucinogens 140

1. Characterization of Hallucinogen Effects 140

a. Acute Drug Effects 140

i. Electroencephalographic 140

(EEG) Activity

2. Hallucinogen Physical Dependence Potential 140

a. Tolerance to Drug Effects 140

b. Withdrawal from Chronic Drug 141

Administration

3. Hallucinogen Abuse Liability 141

a. Drug Self-Administration 141

b. Self-Report of Drug Effects 141

c. Behavioral Toxicity 142

References 142

G. Procedures for Assessing Dissociative 144

Anesthetics

1. Characterization of Dissociative 144

Anesthetic Effects

a. Acute Drug Effects 144

i. Anesthesia 144

ii. Analgesia 144

iii. Electroencephalographic 145

(EEG) Activity

2. Dissociative Anesthetic Physical 145

Dependence Potential

a. Tolerance to Drug Effects 145

b. Withdrawal from Chronic Drug 145

Administration

3. Dissociative Anesthetic Abuse Liability 146

a. Drug Self-Administration 146

b. Self-Report of Drug Effects 146

c. Behavioral Toxicity 146

References 146

List of Monographs 148

xiv
I. Introduction

The Committee on Problems of Drug Dependence. (CPDD) has been in

existence since 1929 and is the longest standing group in the

United States concerned with drug dependence and abuse. Its history

from 1929 until the early 1970s was summarized by the late Nathan

B. Eddy (1973). From 1929 until 1976, the CPDD was associated with

the National Academy of Sciences - National Research Council. Since

1976, it has existed as an independent body presently affiliated

with ten national societies representing various disciplines

concerned with problems of drug dependence and abuse.

A major thrust of the CPDD has been to facilitate the development of

safer and more effective therapeutic agents while reducing the risks

of dependence and/or abuse. Most of the drugs evaluated by the CPDD

are drugs with well-defined current or potential clinical use.

Knowledge of the dependence potential and/or abuse liability of a

drug proposed for marketing is of obvious importance with respect to

its scheduling both within the United States and internationally.

By appropriate screening prior to marketing, it should be possible

to avoid cases of iatrogenic dependence and/or self-generated abuse

that would have occurred and formed the basis for later scheduling

of the drug after post-marketing experience.

In some cases, discovery of dependence potential and/or abuse

liability might be grounds for a decision not to manufacture a drug.

Dependence and abuse are among the costs, along with other adverse

side effects, that must be weighed against a drug's potential

therapeutic efficacy. Ordinarily, a drug will not be tested for

dependence potential and/or abuse liability until other aspects of

efficacy and safety have been demonstrated in a scientifically sound

and systematic manner employing appropriate standards of measurement

and controls to avoid bias. At this point, a serious level of

dependence potential and/or abuse liability can counterbalance the

prospect for a modest therapeutic advance. Thus screening for

dependence potential and/or abuse liability may prevent the

marketing of an undesirable new product.

The drug assessment activities of the CPDD. then, have come to serve

three general purposes. In the first instance, they have provided

1
information to the pharmaceutical industry and the public to assist

in evaluating the dependence potential and/or abuse liability of new

drugs. Secondly, they have advanced the basic scientific purpose of

identifying and characterizing truly novel compounds, and

stimulating their synthesis. In this regard, the CPDD has

collaborated closely with the National Institutes of Health. And

thirdly, the CPDD has implemented and supported the assessment of

dependence potential and abuse liability in both animal laboratory

and human clinical settings. In 1947, the CPDD initiated and

supported an animal testing facility at the University of Michigan.

In the early 1970s. a similar unit was formed at the Medical College

of Virginia. The focus of activity at both facilities has been upon

testing opioids, although in recent years other types of drugs have

been studied more often. In 1980, the CPDD awarded, on a

competitive basis, starter grants to laboratories for the

development of clinical procedures for evaluating drug dependence

potential and abuse liability in man. And, in 1982, initiatives

were undertaken to broaden the scope of CPDD's testing activities to

encompass CNS stimulants and depressants.

The CPDD has twice before, in 1966 and 1973, published position

papers on the testing of drugs for "dependence liability" in animals

and humans (CPDD 1966; 1973). The present paper represents a

continuation of these state-of-the-art reviews. A revision is

timely now because there have been several new and important

developments over the past decade. First, a growing number of

opioids of the mixed agonist-antagonist type are in various stages

of development as anesthetics and analgesics. Reports indicate that

the analgesic effects of these new compounds are not accompanied by

the subjective mood changes associated with abuse liability, but the

evidence in this regard is inconclusive. Second, considerable

public concern has been generated by reports of physical dependence

involving commonly used sedatives and anxiolytics even at

therapeutic doses. Third, a number of cannabinoids and their

homologs are under evaluation as potential therapeutic agents.

Fourth, expanded development of anorectic agents and antidepressants

has emphasized the importance of abuse liability assessment of the

centrally active stimulants. And fifth, the verification of the

street-use of dissociative anesthetics (e.g., phencyclidine and its

congeners) has added another group of substances with high abuse

liability. These substances produce a constellation of effects

including sedation, stimulation, and hallucinosis.

References

Committee on Problems of Drug Dependence, Addendum I: Testing for

dependence liability in animals and man. Bulletin, Problems of

Drug Dependence. Washington, D.C.: National Academy of

Sciences -- National Research Council, 1966. pp. 1-10.

Committee on Problems of Drug Dependence: Testing for dependence

liability in animals and man (Revised 1972). Bull Narc

25:25-39, 1973.

Eddy, N.B. The National Research Council Involvement in the Opiate

Problem: 1928-1971. Washington, D.C.: National Academy of

Sciences, 1973.

2
II. Concepts and Definitions

Traditional concepts and definitions of drug dependence and drug

abuse are in need of re-evaluation and revision. Advances in

knowledge about drug actions, and particularly in research

technology, have made possible an operational approach to

pharmacological assessment of the risks of dependence on, and abuse

of, psychoactive drugs of various categories, yet non-operational

terminology still persists. The distinction between "physical" and

"psychic" or "psychological" dependence, for example, has long since

outlived its theoretical basis. Even the dichotomy between

"physical" and "behavioral" factors has not provided a particularly

useful framework for analyzing the essential dimensions of

drug-related problems. And while the terms "dependence" and "abuse"

are generally considered preferable to the word "addiction" as a

basis for the development of an operational language in this field,

persistent terminological ambiguities must be acknowledged.

One such problem derives from the continued use of the word

"dependence" in at least two quite different ways. In the first

instance, "physical dependence" is used with reference to the

biochemical, physiological, and behavioral consequences of repeated

exposure to a drug resulting in tolerance, and an abstinence

syndrome when the drug is withdrawn. A second, less technical, use

of the term "dependence" (or, more commonly, the phrase "drug

dependence") refers to a complex of behavioral phenomena often

described by such terms as "loss of voluntary control over drug

taking," "compulsive drug use," 'and "reduced range of behavioral

options." In this second case, the obvious overlap with the term

"abuse" (or more commonly, the phrase "drug abuse") further

complicates both technical and non-technical language usage.

In response to these problems, a recently published World Health

Organization Memorandum on Nomenclature and Classification (Bull.

WHO 1981) recommended substitution of the word "neuroadaptation" for

physical dependence, and deletion of the term "abuse" from the drug

lexicon on the grounds that it is essentially a value-judgmental

term rather than an operational one. In addition, the WHO

Memorandum further proposes use of the term "drug dependence

syndrome" to encompass virtually all the phenomena currently

3
described by the two terms "dependence" (i.e., "neuroadaptation"

plus "drug dependence") and "abuse." The main advantages of the

terminology suggested in the WHO Memorandum lay in its clear

differentiation between the primary process (drug selfadministration) and secondary consequences (neuroadaptation), and

its emphasis on experimentally or clinically operational terms

devoid of value judgments.

From a scientific and technical perspective, the phrase "drug

dependence syndrome," as defined in the WHO Memorandum, is

conceptually indistinguishable from the older term "addiction" when

the latter is used to designate the complex of persistent behavioral

and physiological changes associated with chronic selfadministration of drugs. However, the WHO Memorandum was not

designed as an operational guide to the testing of those properties

of drugs that might be related to the risk of dependence and/or

abuse. In contrast, the present document is intended specifically

for that purpose. Since the testing procedures to be described are

largely based on concepts and terminology that had evolved before

the publication of the WHO document, and that still enjoy wide

acceptance, we will not at this time attempt to revise the

operational terms that are in general use.

The final resolution of these differences in terminology has not

been achieved. Many workers in the field as well as the lay public

are still unaware of the new terminology proposed by WHO.

Accordingly, for purposes of this discussion we shall continue to

use the terms "physical dependence" and "abuse" as conventionally

defined. These familiar terms are encoded in the laws and

regulations which control the scheduling of drugs and the

determination of manufacturing quotas. Should the proposed WHO

terminology come into general use, future revisions of the present

document will provide for the necessary changes.

"Physical dependence," as used in the following sections of this

review. refers to physiological and behavioral alterations that

become- increasingly manifest when drug administration is stopped

after repeated exposure to a pharmacologic agent. Changes

associated with (but not limited to) the development of tolerance

and the physiological effects of drug withdrawal are expressed as an

abstinence syndrome (Cochin 1970; Kalant et al. 1971; Eddy 1973;

Clouet and Iwatsubo 1975). "Abuse" is used with reference to events

that precede or accompany strong drug-seeking, drug discrimination,

and drug-taking behaviors in association with self-administration of

a pharmacological agent, often in a social context (Brady and

Griffiths 1977; Griffiths et al. 1980; Brady 1981; Woods et al.

1982). The abuse concept clearly encompasses toxicity' as well,

1 . e., adverse physiological and/or behavioral consequences (Schuster

and Fischman 1975).

The relevance and importance of this distinction between "physical

dependence" and "abuse," for drug evaluation purposes, resides in

the fact that while both these processes are of obvious public

health concern, their defining properties are not coextensive and

they do not invariably occur together. The assessment of "physical

4
dependence potential" in terms of tolerance and withdrawal, while

obviously an important aspect of drug evaluation, does not by itself

predict the drug-seeking and drug self-administration which are

essential features of abuse, and which one attempts to predict by

assessment of "abuse liability." Drug-seeking and drug-taking

behaviors can be maintained in strength by doses of cocaine or

morphine, for example, which produce no significant degree of

tolerance or withdrawal (Schuster and Woods 1967; Johanson et al.

1976; Jones and Prada 1977). Conversely, some compounds such as

propranolol have clear dependence potential (i.e., development of

tolerance after repeated dosing, and an abstinence syndrome after

drug withdrawal) at doses which have little or no attendant abuse

liability, i.e., do not give rise to drug-seeking or drug-taking

(Myers and Austin 1929; Crandall et al. 1931; Ambrus et al. 1951;

Rector et al. 1955; Jaffe 1980).

In summary, from the perspective of drug testing, to which this

review is addressed, pharmacologic assessment can most conveniently

be related to the changes or events antecedent to repeated drugtaking, on the one hand, or consequent to it, on the other. It is

primarily the reactive biochemical, physiological, and behavioral

consequences, of drug administration, both acute and chronic (in

terms of tolerance and withdrawal), which define a pharmacologic

agent's physical dependence potential. The proactive drug-seeking,

and drug discrimination which occur as antecedents to habitual drug

use, on the other hand, together with the adverse effects of such

use (i.e., a combination of the drug's reinforcing properties and

its toxicity), define a drug's abuse liability.

Physical dependence and abuse, as defined, do of course frequently

occur together. Changes in drug-seeking and drug-taking often occur

as sequelae to both the acute effects of a drug and to tolerance and

withdrawal after chronic drug exposure (Musto 1973). Conversely,

the biochemical, physiological, and/or behavioral changes which

define physical dependence can be sequelae of abuse-related drug

self-administration (Jaffe 1980). But the relative contributions of

these distinguishable processes to drug-related problems can vary

widely with different pharmacologic agents as a function of such

factors as dose level, environmental interactions, and previous drug

history (Mendelson and Mello 1982). Moreover, the methods used to

assess physical dependence potential and abuse liability, both in

humans and in laboratory animals, are quite distinct. Therefore

these terms are retained throughout the following description of the

methods by which they are assessed.

Beyond the problem of terminology, of course, fundamental issues

concerning the validity of biochemical, physiological, and

behavioral methods for assessing dependence potential and abuse

liability continue to be debated (Martin 1966; Jasinski 1977). The

ultimate test animal, and the most precious, is of course, the

human. Situations have arisen in which drugs were passed through

the animal screens only to show abuse liability in humans. And

street users have been able to discover abusable drugs or their

combinations which were never tested in animals and were never

suspected of abuse liability even after extensive clinical use.

5
Other drugs, such as the benzodiazepines, may be abused more often

by humans than might have been expected on the basis of animal

screening studies.

Such considerations notwithstanding, the focus developed in this

document on formal changes and their temporal ordering in

relationship to the drug intake event does provide an operational

basis for characterizing the full range of a drug's functional

properties and for evaluating all available measures of its

structure (i.e., physico-chemical) and activity (i.e.,

physiological-behavioral) in comparison to known standards. In the

final analysis, such "pharmacological equivalence" remains our most

reliable approach to the assessment of physical dependence potential

and abuse liability.

References

Ambrus, J.L.; Ambrus, C.M.; and Harrisson, J.W.E. Effect of

histamine desensitization on histamine induced gastric

secretion of guinea pigs. Gastroentero, 18:249, 1951. 

Brady, J.V. Common mechanisms in substance abuse. In: Thompson.

T., and Johanson, C., eds. Behavioral Pharmacology of Human

Drug Dependence. National Institute on Drug Abuse Research

Monograph 37. DHHS Pub. No. (ADM) 81-1137. Washington, D.C.:

U.S. Govt. Print. Off., 1981. pp. 11-20.

Brady, J.V., and Griffiths, R.R. Drug-maintained performance

procedures and the assessment of drug-abuse liability. In:

Thompson, T., and Unna, K., eds. Predicting Dependence

Liability of Stimulant and Depressant Drugs. Baltimore:

University Park Press, 1977 pp. 165-184.

Clouet. D.H., and Iwatsubo. K. Mechanisms of tolerance to and

dependence on narcotic analgesia drugs. Ann Rev Pharmacol

Toxicol, 15:49-71, 1975.

Cochin, J. Possible mechanisms in development of tolerance. Fed

Proc, 29:19-27, 1970.

Crandall, L.A.; Leake, C.D.; Loevenhart, A.S.; and Muehlberger, C.W.

Acquired tolerance to and cross tolerance between the nitrous

and nitric acid esters and sodium nitrite in man. J Pharmacol

Exp Ther, 41:103, 1931.

Eddy, N.B. The National Research Council Involvement in the Opiate

Problem: 1928-1971. Washington, D.C.: National Academy of

Sciences, 1973

Griffiths, R.R.; Bigelow, G.E.; and Henningfield, J.H. Similarities

in animal and human drug-taking behavior. In: Mello, N.K.,

eds. Advances in Substance Abuse. Vol. I. Greenwich, CT:

JAI Press, 1980. pp. 1-90.

Jaffe, J.H. Drug addiction and drug abuse. In: Gilman, A.G.,

Goodman, L.S., and Gilman. A., eds. The Pharmacological Basis

of Therapeutics. 6th Edition. New York: MacMillan, 1980.

pp. 535-584

Jasinski, D.R. Assessment of the abuse potentiality of morphinelike

drugs (methods used in man). In: Martin, W.R., ed. Handbook

of Experimental Pharmacology Vol. 45/I. Drug Addiction I.

Berlin: Springer-Verlag, 1977. pp. 197-258.

6
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.

Jones, B.E., and Prada, J.A. Drug-seeking behavior in the dog:

Lack of effect of prior passive dependence on morphine. Drug

Alchol Depend 2:287-294, 1977 

Kalant, H.; leBlanc, A.E.; and Gibbins, R.J. Tolerance to and

dependence on some non-opiate psychomotor drugs. Pharmacol

Rev, 23:135-191, 1971.

Martin, R. Assessment of dependence-producing potentiality of

narcotic analgesics. In: Radouco-Thomas, C., and Lasagna, L.,

eds. International Encyclopedia of Pharmacology and

Therapeutics. Glasgow: Pergamon Press, 1966 

Mendelson, J.H., and Mello, N.K. Commonly Abused drugs. In:

Harrison's "Principles of Internal Medicine. Chapter 241.

New York: McGraw Hill Book Co., 1982. pp. 1541-1546.

Musto, D.F. The American Disease - Origins of Narcotic Control.

New Haven, CT: Yale University Press, 1973.

Myers, H.B., and Austin, V.T. Nitrite toleration. J Pharmacol Exp

Ther, 36:227, 1929.

Rector, F.C.; Seldin, D.W.; and Copenhaver, J.H. The mechanism of

ammonia excretion during ammonium chloride acidosis. J Clin

Invest, 34:20, 1955.

Schuster, C.R., and Fischman, M.W. Amphetamine toxicity:

Behavioral and neuropathological indices. , Fed Proc

34:1845-1851, 1975.

Schuster, C.R., and Woods, J.H. Morphine as a reinforcer for

operant behavior: The effects of dosage per injection. Report

to Committee on Problems of Drug Dependence. Washington, D.C.:

National Academy of Science, 1967. pp. 5067-5072.

Woods, J.H.; Young, A.M.; and Herling, S. Classification of

narcotics on the basis of their reinforcing, discriminative,

and antagonist effects in rhesus monkeys. Fed Proc,

41:221-227, 1982.

World Health Organization: Nomenclature and classification of drugand alcohol-related problems: A WHO Memorandum. Bull WHO,

59(2):225-242, 1981.

7
III. General Methodological Considerations

The profile of a specific drug's physical dependence potential and

abuse liability is defined by a number of its pharmacologic effects

observed over a wide range of doses. Assessment of these effects

involves a range of procedures that quantify the responses after

acute administration (e.g., analgesia, behavioral performances, EEG

patterns) and after chronic administration (e.g., tolerance

spontaneous and/or precipitated withdrawal). Furthermore

behavioral paradigms for assessing drug-seeking and drug-taking

(e-g., drug self-administration procedures), for determining the

stimulus properties of drugs (e.g., drug discrimination procedures),

and for assessing behavioral toxicity (e.g., sensorimotor

psychophysical procedures) have been employed with increasing

frequency in systematic drug evaluations. The standardization of

such assessment techniques thus makes it possible to provide a

comparative analysis of relative dependence potential and/or abuse

liability in both animals and humans across a number of drug doses

and drug classes with increasing confidence.

Evaluating the acute effects of a drug is a necessary antecedent to

the subsequent study of tolerance, and ultimately, physical

dependence potential. In this regard, numerous pharmacological

assays are employed that serve not only as the comparison response

for tolerance studies, but as the basis for screening unknown

compounds for physical dependence potential and/or abuse liability,

as well. These assessments are made by comparing the effects of the

unknown drug to a standard compound (e.g., morphine, pentobarbital,

amphetamine, etc.). Once the acute effects of a drug are well

characterized, the development of tolerance to these effects can be

adequately quantified. While the fact that tolerance develops after

chronic administration of a drug does not necessarily mean that the

drug possesses significant dependence potential and/or abuse

liability, it may serve as a partial explanation for gradual changes

in intake over time as observed in drug self-administration studies.

Under any circumstances, judicious interpretation of the results of

tolerance studies serves an important function in the ultimate

assessment of a drug's dependence and/or abuse profile.

8
Procedures for assessing the relative dependence potential of a drug

have historically relied upon either substitution or primary

dependence methods. The discovery of specific antagonists has added

an important dimension to the study of physical dependence potential

by providing a means of precipitating withdrawal signs. Another

critical consideration in comparing the physical dependence

potential of two or more drugs is the degree to which the effects of

the drugs during chronic dosing are equivalent. Procedures for

assessing the abuse liability of a drug have focused predominantly

upon drug self-administration and drug discrimination paradigms. In

the former case, methods have been developed for determining not

only whether a drug will be self-injected (e.g., substitution

procedures), but for determining the relative reinforcing efficacy

of a given drug as compared to standard drugs of abuse (e.g.,

progressive ratio and choice procedures). Drug discrimination in

animals, which approximates measures of subjective reports in

humans, has been utilized to measure and compare the discriminative

stimulus properties of drugs both within and between .pharmacologic

classes.

Most importantly, laboratory animal methodologies have provided an

applied technology for quantifying the acute and chronic effects of

a drug in relationship to known standards of dependence potential

and abuse liability. By and large, these methods complement, and in

some cases extend, those employed in human research. Testing in

human beings is complicated by the fact that there is great interindividual variability in reactivity to and tolerance for various

substances and that this initial tolerance level can be drastically

modified by prior drug experience. Therefore, it is necessary not

only to require of volunteers a drug-free period of several weeks to

be certain that any drugs taken either therapeutically or

recreationally have been eliminated, but control and experimental

groups should be matched with respect to drug-taking histories,

baseline reactivity, and tolerance. Simple random assignment to

drug or control group is usually not sufficient to assure equivalent

groups because most drug trials are based on small samples.

In addition to matching experimental and control groups, attention

should be paid to obtaining subjects who will represent the

population likely to be administered the drug clinically. This

means that tests should not be restricted to persons with extensive

prior drug experience, and that subjects should come from the age

and sex groups likely to be given the drug. And since attrition

rates may be high for human drug studies that require weeks or

months of administration and follow-up, rewards for participation

should be increased in proportion to the length of the study to

reduce dropouts. Additionally, attention to methods for endpoint

analysis to avoid loss of data for those who do not complete the

test period is essential, since those compliant with long and

difficult studies are unlikely to be representative of the general

population of potential users.

9
IV. Specific Drugs and Drug Classes

A. General Considerations

Many, though not all, of the substances under consideration are

drugs with well-defined clinical uses, and for some, there are

currently no good therapeutic alternatives. These facts must be

given due consideration in the overall evaluation of drugs with

physical dependence potential and/or abuse liability. New compounds

in these classes should, however, be expected to have either unique

therapeutic properties or fewer adverse side effects than currently

available drugs.

B. Opioids

The opioids (opiates) are a heterogeneous set of compounds widely

utilized in suppressing the symptoms of pain, in the treatment of

cough, dyspnea, and diarrhea, and, in certain circumstances, in the

production of anesthesia, either as a primary agent or in

combination with non-opiate anesthetics. Certain opioids are also

utilized in maintenance programs for the management of opioid

dependence and abuse. Morphine remains the major reference

compound, though many other agents in this class produce comparable

acute and chronic effects. In general, there are three types of

opioids: pure agonists, pure antagonists, and those with mixed

agonist/antagonist actions. Distinctions between pure agonists and

mixed agonist/antagonists can be made on the basis of their effects

on any one or all four distinct opioid receptor populations: mu,

kappa, sigma, and delta, of which the prototypes are morphine,

ethylketazocine, N-allylnormetazocine (SKF 10,047), and enkephalin,

respectively. There is now considerable debate over whether the

sigma opioid receptors are not, in fact, the same as the

phencyclidine receptors or binding sites. This area remains to be

clarified. Most of the compounds are structurally related to the

naturally occurring 4,5-epoxymorphinans (morphine, codeine, and

thebaine). While morphine was formerly difficult to make in the

laboratory, many semisynthetic analogues (e.g., diacetylmorphine or

heroin, hydromorphone, oxymorphone, hydrocodone, and oxycodone)

could be relatively easily synthesized from morphine, codeine, or

thebaine. These latter three key compounds can now be synthesized

10
from readily available simple compounds in good overall yield by a

process which can be adapted to large scale synthesis.

Phenylpiperidines and related compounds such as meperidine,

alphaprodine, loperamide, diphenoxylate, and fentanyl are purely

synthetic drugs. Diphenoxylate and loperamide are particularly

useful in treating diarrhea. Meperidine and alphaprodine are useful

analgesics with a relatively short duration of action. Fentanyl is

used as an anesthetic and in combination with droperidol used to

produce neuroleptic analgesia. Propoxyphene, methadone and its

congeners, and levo-alpha-acetylmethadol (LAAM) are also

efficacious analgesics, but both methadone and LAAM are used

primarily in the management of opioid dependence and abuse.

Relatively pure antagonists such as naloxone, naltrexone, and

nalmefene are essentially free of morphine-like effects, and can

block the effects of morphine and other opioids. The efficacy of

these antagonists varies with the type of opioid receptor acted upon

by the agonist. The opioid agonist/antagonist analgesics represent

a unique class of compounds with very different pharmacological

profiles. Nalorphine

dysphoric effects.

and cyclazocine both possess pronounced

Pentazocine, butorphanol, and nalbuphine are all

effective analgesics, produce physical dependence, and

precipitate

may

withdrawal signs in opioid-dependent subjects.

Buprenorphine, a semisynthetic oripavine derivative, is a highly

lipophilic compound that is a potent analgesic but can precipitate

opioid withdrawal. Because it exhibits a "ceiling" effect to both

its analgesic and side effects,

agonist of the morphine type.

it has been classified as a partial

A recent new use of opiates and opioids has been as the sole agents

in anesthesia. Morphine,. as well as fentanyl, and mixed agonist/

antagonist analgesics have been used in very large doses. Since

respiration is maintained artificially during surgery, the

depressant effect on respiration is obviated. The opioid effects

are then terminated by administering large doses of naloxone.

With the discovery of endogenous ligands (e.g., enkephalins and

endorphins) for the opioid receptor, attempts have been made to

synthesize longer acting peptides (e.g., D-ala2-D-leu5-enkephalin)

with the hope of finding a compound that is selective for

subpopulations of opioid receptors.

C. CNS Depressants Sedative/Hypnotics, Anesthetics,

Anxiolytics, and Antihistamines

CNS depressants that are employed as sedative/hypnotics or as

anxiolytics comprise a wide range of chemically different compounds.

While few qualitative differences exist between these compounds,

they possess marked quantitative differences with respect to the

degree of CNS depression. The older hypnotics include the inorganic

bromides, chloral hydrate, and paraldehyde, while the barbiturates

(e.g., pentobarbital) are currently more widely used. More

recently, the use of nonbarbiturate sedative/hypnotics belonging to

various distinct chemical classes has increased steadily. These

include the acylureas (e.g., carbromal and bromvaletone), carbamates

11
(e-g, ethinamate and meprobamate), piperidinediones (e.g.,

gluthethimide and methyprylon), acetylenic carbinols (e.g.,

methaqualone), benzodiazepines (e.g., diazepam and nitrazepam), and

the aliphatic alcohols (e.g., ethanol).

Another class of compounds that exerts generalized CNS depression is

the volatile anesthetics. Abuse of such licit compounds as various

halogenated anesthetic gases including halothane, methoxyflurane,

and enflurane, is relatively rare. The anesthetic nitrous oxide,

however, is currently enjoying a revival of illicit use. Obsolete

compounds include the ethers, cyclopropane, fluroxene, and ethylene.

Illicit use of compounds typically employed as propellants in

commercial spray cans (e.g., fluorocarbons), and various solvents

and glues such as aliphatic hydrocarbons (e.g., hexane and naphtha),

aromatic hydrocarbons (e.g., toluene, xylene, and benzene), ketones

(e.g., acetone and cyclohexanone), esters (e.g., amyl acetate),

alcohols (e.g., butyl, ethyl, methyl, and isopropyl alcohols),

glycols (e.g., ethylene glycol), and gasoline is more common.

The older antihistaminics (H1 blocking agents) are classified into

three chemical. groups:

tripelennamine),

ethylenediamines (e.g., pyrilamine and

doxylamine), and

aminoalkylethers (e.g., diphenhydramine

alkylamines e. g., chlorpheniramine and

pheniramine). Most of the H1 blockers produce some degree of

sedation and drowsiness at therapeutic doses. There are some

exceptions (phenendamine). Further, at higher doses the H1 blockers

have been observed to produce hallucinosis. The primary actions of

the H1 blockers are to block the histamine-induced contractions of

the bronchiolar .and gastrointestinal smooth muscle with partial

effects on the cardiovascular system and no effect on histamineinduced gastric secretions. The latter action is postulated to be

another receptor - H2. The prototypic H2 blocking agent,

cimetidine, has been reported to have CNS activity especially in the

elderly.

D. CNS Stimulants: Anorectics, Local Anesthetics, and

Antidepressants

Compounds that stimulate the CNS and/or elevate mood are represented

by numerous pharmacological classes. The xanthines (e.g., caffeine,

theobromine), phenylethylamines (e.g., amphetamine, pemoline), and

diphenylmethanes (e.g., methylphenidate, deanol) all increase;

wakefulness, talkativeness, and random motor activity.

often elevated, sometimes with euphoria, and respiration is

occasionally stimulated. Anorexia is a common effect as well. In

addition to CNS stimulation, compounds like cocaine (which is a

benzoic acid ester extracted from the leaves of Erythroxylon coca),

as well as its analogues (e.g., procaine, benzocaine) possess marked

local anesthetic action. While the tricyclic antidepressants such

as imipramine and amitriptyline elevate mood in depressed patients,

they are generally without effect in normal individuals. Newer

antidepressants of different chemical classes, however, may have

such potential.

12
E. Hallucinogens and Anticholinergics

Many compounds, when taken in sufficient doses, can cause

hallucinations that may be secondary to delirium, sedation, or

stimulation. The compounds appropriately classified as

hallucinogens produce. visual and/or auditory hallucinations not

necessarily associated with delirium, sedation, or stimulation.

Many compounds are derivatives or analogues of endogenous amino

acids. Compounds in this class include the substituted

indolealkylamines (e.g., lysergic acid diethylamide or LSD,

dimethyltryptamine or DMT, psilocybin. harmine, and ibogaine). and

the substituted phenylalkylamines (e.g., mescaline, 2,5-dimethoxy4-methylamphetamine or DOM). The 3-N-substituted piperidyl

benzilates (e.g. ditran) and the classical anticholinergics atropine

and scopalamine, when taken in sufficient doses, also produce

hallucinosis.

F. Cannabinoids

Residues from Cannabis sativa include a number of chemically

distinct compounds with the cannabinoids the

pharmacologically active group.

being major

At least 20 different naturally

occurring cannabinoids have been identified and include delta-gtetrahydrocannabinol (THC), delta-8-THC, cannabinol, and

cannabidiol. The most potent compound, delta-9-THC, has been shown

to be a potentially useful agent in the management of emesis,

glaucoma, and muscle spasm. Many congeners of delta-9-THC have been

synthesized, of which synhexyl (a delta-6a

-lOa, n-hexyl derivative

of THC) and DMHP (a dimethylheptyl side chain congener of synhexyl)

were the first to be studied. Significant undesired side effects

precluded further studies with these compounds, but other synthetic

analogues such as nabilone, SP 106, and CP 44,001 may be potentially

useful therapeutic agents in man.

The cannabinoids, unlike other compounds reviewed, present a 'range

of problems which have hindered the understanding of their

pharmacology. The identification and standardization of the active

constituents in these preparations has been difficult. The

compounds are relatively unstable, insoluble in aqueous solutions,

and have many metabolites, including some that are pharmacologically

active. Clearly, the development and refinement of methods for the

isolation and purification of the natural alkaloids of the plant

Cannabis sativa have been instrumental in the progress that is

reviewed here. In addition, a number of synthetic cannabinoids have

demonstrated some specificity in their actions, and, as such, may

prove to be more useful as potential therapeutic agents. The

assessment of the cannabinoids as a class, however, must necessarily

involve procedures sensitive to all known actions of these

compounds; the standard for comparison is the major active agent,

delta-9-tetrahydrocannabinol.

G. Dissociative Anesthetics

The prototypic compound, 1-(1-phenylcyclohexyl) piperidine or PCP

produces a wide spectrum of effects, from stimulation and

13
hallucinations to depression and analgesia. One analogue, ketamine,

enjoys a widespread use in veterinary medicine and some special uses

as a short-acting anesthetic in man. Other analogues have been

synthesized which are essentially based on substitutions of one of

three functional groups of the PCP nucleus: phenyl ring, cyclohexyl

ring, or the amino group. The latter analogues are not legitimately

manufactured or marketed, but widespread illicit synthesis has been

documented.

While most studies on the structure-activity relationships of PCP

analogues have used compounds with substitutions on the phenyl or

cyclohexyl ring, the majority of illicitly synthesized compounds are

N-substituted.

been the

Small chain alkyl groups (e.g., ethyl, propyl) have

most common substitutions which N-ethyl-lphenylcyclohexylamine (PCE) and 1-(n-propyl)-1-phenylcyclohexylamine

(NPPCA), respectively. The former compound, PCE,is currently under

Schedule I control as is 1-(1-phenylcyclohexyl) pyrrolidine or PCPY.

Toxicological manifestations of PCP analogue abuse have also been a

major cause for concern with respect particularly to the compound

l-piperidinocyclohexanecarbonitrile or PCC which, upon entering a

biological system, releases hydrogen cyanide. While PCC itself is

currently under Schedule II control (like PCP), it is a precursor to

PCP synthesis and thus tends to be a common contaminant in illicitly

synthesized PCP and other analogues. PCP analogue abuse is a

complex problem which essentially stems from the fact that the

identity of the compound is rarely confirmed and very little

information is available about the many easily synthesized

analogues.

14
V. Animal Testing Procedures

A. General Methods

1. Procedures for Characterization of Drug Effects

a. Acute Drug Effects

i) Spontaneous locomotor activity

The jiggle cage method of Harned et al. (1952) determines the

effects of test compounds on spontaneous motor activity.

rats are placed in a cage suspended in air by a spring.

Usually,

Movements

of the rat are recorded kymographically on chart paper and can be

quantified with respect to control animals given vehicle. An

additional measure of locomotor activity is obtained using an openfield grid (Brimblecombe 1963). An observer (blind to the

treatment) counts the number of squares entered over a set period of

time. The photo-cell annular cage is also employed to determine

motor activity. Typically, mice or rats are treated with either

vehicle or the test compound and the number of times an animal

interrupts the light beam over a given interval is recorded.

Time-course data are easily obtained with this method using

cumulative recorders.

ii) Forced motor activity

A measure of motor impairment (ability of rats or mice to remain on

a rotating rod) can be obtained using a variety of procedures, one

example being the rotorod assay (Dunham and Miya 1957; Kinnard and

Carr 1957). The device consists of a motor-driven, aluminum rod

(6-8 cm diameter) which is divided into several segments by circular

aluminum plates which serve to limit lateral movements of the

animals on the rod. Once the rodent is placed on the rod, it is set

in motion and the time until the animal falls off is recorded.

Typically, a maximum cut-off of 1-2 minutes is used. Animals can be

tested numerous times after a single injection to generate

time-course data.

15
iii) Electroencephalographic (EEG) activity

This procedure employs the brain's electrical activity as a measure

of a drug's effect (Klemm 1969). Recently developed computerassisted analysis (e.g., power spectrum) has permitted not only

qualitative but quantitative measures of drug-induced changes in EEG

activity. The EEG has been used to quantify acute and chronic drug

effects in various species including the mouse (Morley and Bradley

1977), rat (Lukas et al. 1980). cat (Schallek et al. 1967), dog

(Wikler and Altschul 1950), and monkey (Gehrmann and Killam 1976).

This technique is most often employed in chronic animals which have

been implanted with cerebrocortical EEG electrodes, although subcortical electrodes have also been employed. The animals are

typically allowed to acclimate to the surroundings and control EEG

activity is recorded. Once the EEG recording is stable, a drug is

administered and' the resultant EEG activity is monitored on a

polygraph and stored on magnetic tape for subsequent computerized

power spectral analysis. This analysis consists of measuring the

amount of energy (or power) as a function of frequency.

Another component of the analysis of the EEG effects of drugs of

abuse relates to the quantification of various stages in the

sleep-wakefulness continuum. In all mammals, at least three stages

are readily distinguished - a waking stage, a slow-wave stage of

sleep, and a rapid-eye-movement (REM) stage of sleep. The EEG from

mammals can be scored for the onset, duration, and cyclicity of the

stages. Transitions between stages are also noted. These form the

basis for the evaluation of drug-induced alterations (van Twyver

1969).

In recent years, there has been increasing use of evoked potentials

in the EEG to define and quantify the actions of drugs. A constantintensity stimulus, such as a light flash, a signal tone, or a mild

electric shock, is administered repeatedly to the subject, and the

EEG activity is recorded during a fixed sweep of the last 500 msec

of each stimulus. Summation of these recordings in a computer

results in averaging-out of the random background activity, but

progressive augmentation of the specific wave pattern of response to

the stimuli. Drug effects on the latency and amplitude of each

component of the response can then be expressed in precise

quantitative terms (Kalant 1978a).

iv) Behavioral performance

Numerous drugs have been reported to modify a wide range of

performances including food and water consumption, exploratory and

aggressive behaviors, punishment, escape, avoidance, and conditioned

suppression responses. With the exception of the conflict procedure

(Geller and Seifter 1960), which is relatively selective for

anxiolytics, few behavioral paradigms are employed routinely in

animal models of abuse liability and/or dependence potential.

Consummatory behavior (e.g., food or water intake) has been shown to

increase after acute administration of opioid antagonists, but

benzodiazepines induce hyperphagia as well. In addition, procedures

which employ response rates as a measure of performance appear to be

16
affected by drugs belonging to a wide range of pharmacological

classes. This relationship is also dependent upon the response rate

prevailing at the time of drug administration (e.g., law of initial

effect); thus, procedures that maintain low response rates are

typically increased by many compounds while high response rate

performance is typically suppressed. This phenomenon has been

elegantly demonstrated using a multiple schedule technique (e.g.,

mult. FR/FI). Drug-induced changes of response rates in the same

animal can be demonstrated to be directly related to the schedule of

reinforcement (Dews 1955; Kelleher and Morse 1968).

The contributions of behavioral pharmacology to the assessment of

drug abuse liability and physical dependence potential have had

their greatest impact in the areas of tolerance and withdrawal. At

least three different behavioral factors have been shown to have

important effects upon the rate of development of tolerance, and

upon the degree of tolerance developed. The first is the presence

or absence of a requirement for task performance while the subject

is under the influence of a drug. Animals receiving the drug just

prior to the start of each training session acquire tolerance more

rapidly than animals receiving the same dose of the same drug

immediately after each training session (Kalant et al. 1971; LeBlanc

et al. 1973). The second factor is the effect of the drug upon the

relationship between the performance and its consequences. For

example, rats were trained to bar-press for food rewards on a

schedule of alternating periods of FI and DRL. Under the influence

of the first dose of amphetamine, their response rates increased

during both components of the schedule, but this caused loss of

rewards only in the DRL component and not during the FI. With

repeated amphetamine tests on the same schedule, the rats rapidly

developed tolerance to the rate-increasing effect during the DRL

portions, while showing no tolerance during the FI portions of the

schedule (Schuster et al. 1966). The same phenomenon has been

observed subsequently with other drugs (Krasnegor 1978). The third

factor is the presence or absence of environmental stimuli linked by

Pavlovian conditioning to the administration' of a drug. If an

animal receives a dose of a given drug repeatedly in the same

environment, it shows more tolerance when tested in the same

environment than when tested with the same dose in a new environment

(Siegel 1976; Le et al. 1979).

All three sets of observations mentioned in the preceding paragraph

have given rise to the concept of behavioral tolerance as distinct

from classical or pharmacological tolerance (Krasnegor 1978).

However, it has been argued that these are not separate forms of

tolerance, and that the behavioral factors simply alter the rate or

extent of development of an intrinsic biological adaptation (Kalant

et al. 1971; Kalant 1978b). The extent to which behavioral

variables contribute to the tolerance will vary according to the

specific circumstances. It is important, however, in testing drugs

for physical dependence liability, to ensure that the same test

paradigm is employed throughout, so that the behavioral influences

are kept constant for all the drugs under comparison.

17
Behavioral procedures for determining the appetitive and/or aversive

properties of drugs have as well extended the application of

classical conditioning methods to the assessment of drug abuse

liability. Conditioned taste aversion paradigms have been employed,

for example, in which the consumption of a saccharin solution to

which an animal has been previously exposed in association with

psychoactive drug injection is subsequently compared with water.

Saccharin preference under such conditions indicates appetitive drug

effects whereas saccharin rejection suggests aversive properties

even though drug self-administration tests may reflect reinforcement

effect (Cappell and LeBlanc 1975). A similar "place preference"

procedure involves a comparison of the time spent in each of two

distinctive environments (i.e., differing in visual, auditory,

olfactory, and/or tactile cues) as a function of different drug,

drug dose, or vehicle injections under such clearly discriminable

conditions. The appetitive effects of cocaine or morphine, for

example, are reflected in the increased time spent in the

environment associated with such drug injection, while aversive

effects produce avoidance of the drug injection site (Mucha et al.

1982).

v) Binding assays

With the discovery of specific receptors to which opioids,

stimulants, neuroleptics, and benzodiazepines bind, quantitative

study of structure-activity relationships (QSAR) has emerged as a

major pharmacological method which has provided extensive

information regarding the basic mechanism of action of drugs of

abuse. Binding sites in brain tissue have as well been reported for

both PCP and its analogues, and the good correlation between binding

affinity and potency in clinical use has served not only to enhance

understanding of drug/receptor interactions, but to validate the use

of this technique as a predictor of drug effects as well. While

binding to CNS tissue is normally associated with a drug's

physiological/behavioral effects, binding assays involving other

tissues (e.g., smooth muscle) have proven equally useful. In fact,

guinea pig ileum and mouse vas deferens have been used extensively

to characterize the binding characteristics of the opioids and

endogenous, opiate-like peptides (Kosterlitz et al. 1973; Woods et

al. 1981).

The basic concept in binding studies involves the use of a

radiolabelled (e.g, 3H, or less frequently, l4C) ligand (e.g.,

dihydromorphine, flunitrazepam) which is first incubated with the

tissue to which nonlabelled test drug is then added. By measuring

the remaining radioactivity on the tissue, the degree of

displacement of the labelled ligand can be determined, and thus a

measure of relative affinity for the binding site is obtained for

the test compound. Additional information regarding the binding

characteristics of drugs of abuse is obtained by determining the

effects of adding receptor antagonists or electrolytes (e.g., Na+)

to the incubation (Pert and Snyder 1974). Differential and

incomplete displacement of binding has been interpreted to mean that

there are multiple receptor populations for a specific drug class.

In many instances, a high correlation between the results of binding

18
studies and the observed physiological and behavioral effects with

certain drugs has been found. This has led to the inclusion of

binding characteristics in the pharmacological profile of some

drugs.

2. Procedures for Assessing Physical Dependence Potential

a. Tolerance to Drug Effects

The development of tolerance to a drug is reflected by an

attenuation of the original response with subsequent exposures to

the drug. Thus, the dose must be increased in order to re-establish

the original response. By quantifying the acute effects of a drug,

it is possible to track the development of tolerance to any one

effect. This is particularly important because tolerance does not

necessarily develop equally, that is, to the same degree or at the

same rate, to all of a drug's effects. Tolerance may be

dispositional or functional; the former is primarily due to an

increased rate of biotransformation and/or drug elimination from the

body, while the latter refers to a diminished sensitivity of the CNS

(Kalant et al. 1971). An additional dimension to tolerance which

has become evident as a result of refinements in the development of

performance procedures is behavioral tolerance. This phenomenon may

be apparent only with respect to some specific aspect of the

behavioral repertoire which has previously been "learned" under the

influence of the drug (see above "Behavioral Performance" section).

The development of tolerance is most apparent when exposure to the

drug is continual. Various techniques have been employed to

maximize drug exposure, including continual infusions, multiple

parenteral injections, addition of a compound to food or drinking

water, and implantation of subcutaneous pellets or miniature osmotic

pumps.

b. Withdrawal from Chronic Drug Administration

The abstinence syndrome is defined by the physiological and

behavioral consequences of terminating chronic administration of a

drug (also called abrupt withdrawal). Both the dose and duration of

exposure determine the dependence potential of a drug, and the

degree of dependence potential differs among drugs from different

classes. The basic procedures for assessing these effects, however,

are similar in format, and involve two basic techniques: direct or

primary (dependence), and single dose substitution (Fraser and

Jasinski 1977). In the former, the test compound is administered on

a chronic basis to drug-naive animals; the initial dose is usually

low, and as tolerance to toxic effects develops, the dose is

increased. At the end of the dosing schedule the drug is withheld,

and the animals are observed for physical signs and symptoms of

withdrawal. Single dose substitution procedures involve rendering

animals dependent on a standard drug (e.g., morphine, barbital) for

a specified period of time. The drug is then withheld, and the

animals are observed for withdrawal signs. Once the withdrawal

syndrome appears, a single dose of the test compound is administered

19
to determine whether the withdrawal signs can be reversed or

attenuated.

A recent development in the study of relative dependence potential

has been the use of a functionally equivalent dosing regimen for the

drugs being compared (Boisse and Okamoto 1978). This procedure

takes into account the physicochemical properties of a drug which

determine metabolic and elimination rates (and thus, the amount of

drug in the body) as a factor influencing dependence potential. The

most common procedures for accomplishing functional equivalence are

behavior rating scales (for degree of CNS depression) and devices

for measuring ataxia (e.g., rotating rod). Doses are adjusted in

order to maintain equal degrees of sedation.

Most dependence studies are conducted using spontaneous withdrawal

techniques (i.e., the drug is simply discontinued). Techniques for

assessing opioid dependence potential, however, have been extended

by the development of an opioid receptor antagonist, naloxone.

Thus, precipitated withdrawal is possible and the need to wait 12-24

hours for withdrawal signs is obviated. The recent discovery of a

benzodiazepine receptor antagonist (Ro 15-1788) has also provided

for a similar extension of dependence assessment procedures with

this increasingly more prominent class of clinically useful

compounds (Lukas and Griffiths 1982).

3. Procedures for Assessing Abuse Liability

a. Drug Self-Administration

Drug self-administration procedures represent the most widely used

method for assessing relative abuse liability with drugs belonging

to various classes. With the development of a chronic intravenous

catheter (Clark et al. 1961; Weitzman et al. 1961) and methods for

automated delivery of drugs, studies of drug self-administration

were first undertaken in rats over two decades ago (Weeks 1962;

Weeks and Davis 1964). Procedures for implanting i.v. catheters in

non-human primates have since been refined and detailed (Deneau et

al. 1969; Lukas et al. 1982). Chronic intragastric catheters have

also been developed for rats (Gotestam 1973; Lukas and Moreton 1979)

and primates (Altshuler et al. 1975; Lukas et al. 1982)) and

automated drinking systems have been described in reports of oral

drug self-administration studies (Meisch and Henningfield 1977; Ator

and Griffiths 1982).

Two basic types of drug self-administration procedure are commonly

used: 1) continuous or direct, and 2) substitution. In the former,

the animal is allowed access to the test compound without prior

experience. This procedure is useful in assessing whether an animal

will initiate self-injections of the test compound, and if so,

whether responding for the compound will change over time. In the

substitution procedure, drug self-administration is first

established with a standard compound (e.g., cocaine, morphine) known

to be reinforcing, that is, animals will work to obtain access to

the compound. Once performance and drug intake have stabilized, the

test compound is substituted for the standard. The abuse liability

20
pattern is defined by numerous characteristics including intake,

time course, and stability of responding. Once self-administration

behavior is established with a compound (e.g., dose-response

functions are generated, relative potencies are determined), the

next step in characterizing abuse liability is to determine relative

degree of reinforcing efficacy. Numerous procedures including

progressive ratio, response rate analysis, discrete-trial choice,

and concurrent schedule control, have been utilized to address this

question.

i) Progressive Ratio Procedures

Progressive ratio procedures involve a two-step operation in which

stable operant performance (typically maintained on a fixed-ratio

schedule) is first obtained with the reinforcer. Subsequently, the

number of responses required for delivery of reinforcement is

systematically increased until responding falls below some criterion

level. The highest ratio at which performance remained above

criterion is referred to as the "breaking point"; Hodos and Kalman

(1963) demonstrated that the higher the concentration or volume of

liquid reinforcer the greater the breaking point. Because of this

relationship the breaking point is taken to represent a measure of

relative reinforcer strength.

The extent to which a progressive ratio procedure can provide useful

data regarding the reinforcing efficacy of a drug is dependent upon

the fulfillment of certain criteria. First, since the actual

procedure followed varies substantially between laboratories, it is

important to recognize that the measures obtained are not absolute,

but rather are relative to some standard. Therefore, it is crucial

that all interpretations of reinforcing strength be made on the

basis of relation to a standard compound (e.g., morphine,

pentobarbital, cocaine, etc.). Second, since the reinforcing

properties of drugs are dose related, there is every reason to

suspect that the breaking point may also be dose related. Thus,

complete dose-effect curves must be evaluated in order to determine

not only whether different drugs maintain higher breaking points,

but also whether different doses maintain this performance as well.

ii) Response Rate Analysis

Interpretations of relative reinforcing strength can sometimes be

measured in terms of the rate of responding prior to the drug

injection. The degree to which this applies, however, is dependent'

upon the schedule of reinforcement employed. Historically, a

negative correlation has been shown to exist between drug dose and

response rate (Downs and Woods 1974; Goldberg et al. 1972; Pickens

and Thompson 1968). The response requirement for these studies was

typically a continuous reinforcement or a low fixed ratio, and thus,

the direct effect of the self-injected drug was most likely

responsible for the decreases' in response rate. Using schedules

that employ either a fixed-interval schedule (Balster and Schuster

1973) combined with a time-out (15 minute) period after each selfinjection or a fixed ratio schedule with a long time-out period

(Griffiths et al. 1981), however, it is possible to observe a

21
positive relationship between increasing drug dose and higher rates

of responding. This relationship does nonetheless have the

limitation that, as the direct effects of higher doses of the drug

persist for, longer periods of time, response rates will decrease.

iii) Choice Procedures

An obvious extension of procedures that assess the reinforcing

strength of a single drug is one which permits an animal to select a

preferred drug dose, or to choose between two different drugs.

Typically, different colored lights are used to signal the

availability of different reinforcers. After initial exposure with

various doses of a single drug (Brady and Griffiths 1977) or two

different drugs (Johanson and Schuster 1977), the animal is given

the opportunity to choose between the doses or drugs by operating a

third lever which switches the color of a viewing light indicating

which reinforcer is available. Only one reinforcer is available at

a time. Completion of the response requirement on the appropriate

lever results in delivery of drug previously associated with that

1 color. The percentage of choices of one drug or dose over another

and the response rates maintained by the injection are determined to

reflect the relative degree of reinforcing strength. Attempts to

eliminate possible confounding effects of drug interactions have

resulted in procedures which offer choices between drug and food

(Griffiths et al. 1975).

iv) Concurrent Schedules

The procedures for assessing concurrent drug availability differ

from choice procedures in that concurrent schedules make both drugs

(or different doses of the same drug) available simultaneously.

Using variable interval schedules, higher rates were generally

associated with the response alternative which delivered higher

cocaine doses (Iglauer et al. 1975). As with other procedures, this

relationship became asymptotic as higher doses were studied.

b. Drug Discrimination Procedures

Assessment of the discriminative stimulus properties of drugs has

been recently demonstrated to provide a useful adjunct to selfinjection techniques for the evaluation of a pharmacological agent's

abuse liability. The value of drug discrimination procedures in

this regard relates to the fact that these procedures are designed

to utilize the physiological or interoceptive alterations induced by

many psychoactive drugs as the "cue" for an appropriate response to

obtain food or avoid shock (Overton 1971). Thus, these procedures

can provide information analogous to a human testing situation in

which subjects categorize drugs with respect to their subjective

effects.

The basic procedure involves training animals to respond

differentially (depending upon the nature of drug pretreatment) in

either a T-maze (e.g., go left if drugged; go right if nondrugged)

or a two-lever choice situation (e.g., left lever produces food or

avoids shock if drugged; right lever produces food or avoids shock

22
if nondrugged). Training time can be decreased by introducing

differential drug conditions on the first exposure rather than

pretraining on both levers or arms of a T-maze (Overton 1979). Once

stable responding has been established and the percent of correct

responding is high (e.g., ›)85%) then test sessions are initiated in

which novel drug conditions are presented in order to assess

generalization. Within the context of abuse liability testing,

three basic procedures can then be followed. First, generalization

of the training dose can be demonstrated for other doses of the

training drug (Schuster and Balster 1977). This ensures that the

discriminative stimulus of the drug is in fact controlling the

responding. Secondly, once an appropriate training drug and dose

are chosen, generalization to other compounds within the same

pharmacologic class can be assessed to provide evidence for the

pharmacological specificity of drug stimulus control. All results

must be interpreted in relation to the training drug dose, however,

since this can markedly affect the results of such testing (Waters

et al. 1972). Thirdly, cross-generalization to drugs belonging to

other pharmacologic classes can be assessed in a similar manner.

Generally, drugs belonging to the same or similar classes tend to

produce training-drug-appropriate responding while drugs from

different classes do not (Colpaert and Rosecrans 1978).

c. Behavioral Toxicity

A broad range of performance procedures has been used to assess both

the acute and chronic decrements which define the behavioral

toxicity of abused drugs. Progress in this area of pharmacology has

closely paralleled laboratory developments in the experimental

analysis of behavior, and has focused most recently upon

psychophysical methods for sensorimotor assessment. The role of

behavioral toxicology has become increasingly important in such drug

evaluation because substances with only minimal (if any) disruptive

behavioral or physiological effects are not generally regarded as

having significant abuse liability even though self-administration

may be widespread (e.g., caffeine in tea or coffee). In contrast,

compounds self-administered even sparingly which produce disruptive

physiological/behavioral changes are considered to have high abuse

liability (e.g., lysergic acid diethylamide). Drugs may fall

anywhere on the continua defined by these parameters and relative

abuse liability is most effectively determined by a comprehensive

assessment of these interactive behavioral/physiological dimensions.

Most procedures employed are based on standard behavioral paradigms

(Weiss and Laties 1975), and,as such, are distinguished from the

more traditional assessment of teratology. Performance decrements

are usually assessed in terms of motor impairment as an index of

behavioral toxicity (e.g., changes in lever pressing to obtain food

or avoid shock), although recent developments in the application of

classical psychophysical procedures have broadened the scope of

sensorimotor evaluation techniques. These refinements in animal

psychophysical methodologies have made available valid and reliable

experimental procedures for precise measurement of sensory functions

in non-human primates, birds, rodents, and carnivores (Masterton et

al. 1969; Stebbins 1973; Fay 1974; Dolling 1980). In general, these

23
advances can be seen to have resulted from a marriage of two

technologies--classical human psychophysics on the one hand, and

animal learning and conditioning on the other. Further, the

training and testing procedures for obtaining valid and reliable

measures of sensory function in such animals have been thoroughly

documented (Blough 1966; Stebbins 1970). providing convincing

demonstrations of their sensitivity and specificity in more general

behavioral pharmacology applications.

Most recently, for example, applications of these psychophysical

procedures to the evaluation of auditory and visual threshold and

reaction time changes in baboons have been reported following

pharmacological treatments involving several drugs of abuse (Brady

et al. 1979; Hienz and Brady 1980; Hienz et al. 1981). Baboons

housed in individual cages and maintained on a 22-hour restricted

feeding schedule were trained to press a lever and hold it depressed

for varying intervals until presentation of a light flash or tone

burst. Correct responding (defined by release of the lever within

1.5 seconds of stimulus presentation) was rewarded with bananaflavored food pellets. A 1-second intertrial interval (ITI) was

then imposed during which no stimuli were presented and any lever

press reinstated the ITI. Experimental sessions were conducted

daily over a Z-hour period with auditory and visual thresholds

determined by randomly varying the intensity of the test stimuli

from trial to trial and examining the number of correct lever

'releases. Four to five separate measures of thresholds and reaction

times were obtained, each based on blocks of 140 trials, thus

providing a measure of the time course of drug effects.

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Schuster, C.R., and Balster, R.L. The discriminative stimulus

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Advances in Behavioral Pharmacoloqy. New York:

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Schuster, C.R.; Dockens, W.S.; and Woods, J.H. Behavioral variables

affecting the development of amphetamine tolerance.

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Siegel, S. Morphine analgesic tolerance: Its situation specificity

supports a Pavlovian conditioning model. Science 193:323-325,

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Stebbins, W.C., ed. Animal Psychophysics: The Design and Conduct

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J Phys Anthrop, 38:357-364, 1973.

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pp. 43-57.

28
6. Procedures for Assessing Opioids

1. Characterization of Opioid Effects

a. Acute Drug Effects

i) Analgesia

Numerous methods are currently available that provide reliable

assessments of the analgesic potency of an opioid. Traditionally,

methods have been segregrated into two classes depending on the

degree to which a non-opioid (e.g., salicylate) can be distinguished

from an opioid analgesic. In the case of the former, Randall and

Selitto (1957) have described a technique in which brewer's yeast

(0.1 ml of a 20% suspension) is injected into a rat's foot. The

ensuing pain and swelling are determined by applying increasing

pressure to the foot. Salicylates elevate the pain threshold in the

inflamed foot only, while opioids elevate pain thresholds in both

hind legs.

Procedures for assessing the analgesic properties of opioids are

mainly conducted with rodents and assess the degree of pain from

heat (e.g., tail flick, hotplate), pressure (tail compression), or

electric shock (e.g., shock titration, stimulation). With the

exception of electric shock, the basic procedures are similar and

 are usually characterized by a group study design, appropriate

controls (e.g., for habituation), operationally defined end-point

responses (e.g., paw lick, vocalization), and a maximal cut-off time

to prevent tissue damage. The tail flick (D'Amour and Smith 1941),

hotplate (Eddy and Leimbach 1953), hot water (Janssen et al. 1963),

electric stimulation (Knowlton and Cross 1943), flinch jump (Evans

1961)) and tail compression (Dandiya and Menon 1963; Geller et al.

1979) methods all include these features. Typically, animals are

first tested for their sensitivity to the system under non-drug

conditions. Then, groups of 8-10 rodents receive either the test

drug or vehicle and are retested at various intervals (e.g., 10

minutes). The "index of analgesia" can then be obtained by

employing the method of Cox et al. (1968) which normalizes the

results from different compounds. Analgesia (as measured by an

increased latency to respond) is characterized not only by the time

to peak effect, but by onset and offset as well. The use of a

maximal cut-off time would generally preclude the quantitation of

analgesic efficacy. However, calculation of a percent of "maximal

analgesia" according to the formula (test)-(control)/(maximum)-

(control) X 100 can circumvent the problem. Collins et al. (1964)

attempted to obviate the problems associated with the artificial

"ceiling" of analgesic effect with opioids by using a rectal

electrode method for delivery of electrical stimulation of

increasing intensity in the range of 0 to 24 V, at a rate of 1 V/set

with rats. Reaction thresholds were measured as a function of the

time from the onset of stimulation until vocalization. Drugs were

given S.C. or i.p. and animals were tested at 15 and 30 minutes, and

then at 30 minute intervals for 3 hours after drug administration.

Weak analgesics such as acetylsalicylic acid were inactive, while

barbiturates were active only at doses that induced deep CNS

29
depression. Using the increase in threshold in volts (rather than %

of control), the dose-effect curve for morphine increased

exponentially with increasing dose while codeine possessed a much

shallower, linear curve.

ii) Spontaneous Locomotor activity

These procedures described above in general methods are commonly

used to assess the pharmacological activity of opioids. While the

measure of locomotor activity does not provide information relevant

to the prediction of dependence potential, it does provide a baseline

for the study of tolerance and cross-tolerance to other drugs.

Opioid-induced changes in locomotor activity are characterized by

both depressant and stimulant actions. In general, the biphasic

response consists of an initial depressant phase which typically

precedes the delayed stimulated phase. Since the overall effect is

species-, dose-, and time-dependent, however, it is important to

obtain frequent measures of locomotor activity after drug

administration rather than single, fixed-time measures (e.g., 30

minutes post injection).

iii) Electroencephalographic (EEG) activity

Visual analysis of the direct EEG has revealed that morphine and its

surrogates slow the predominant frequency and increase high-voltage

delta activity (Khazan 1975); some analogues such as methadone

induce spike-and-dome activity after high doses.. On the basis of

EEG and behavioral profiles and naloxone sensitivities, it is

possible to distinguish between opioids acting at different

receptors (Tortella et al. 1980). While the effect of opioids on

the EEG are readily discernible, the need to quantify the changes in

both voltage and frequency components has resulted in the emergence

of numerous computer-assisted techniques. Currently, power spectral

analysis is used to quantify not only the acute effects of opioids,

but the development of tolerance, cross-tolerance, and drug-seeking

behavior as well (Young et al. 1978).

In the application of EEG procedures for assessing the acute effects

of opioids, as described in general methods above, opioid effects

may appear in biphasic form. In characterizing the EEG profiles of

morphine, methadone, l-alpha-acetylmethadol (LAAM) and its two

demethylated metabolites (NLAAM and DNLAAM), for example, an 8-hour

post-injection evaluation revealed the appearance of distinct EEG

changes 1-2 hours after LAAM which matched those observed

immediately after NLAAM (Lukas et al. 1980). Cross-tolerance has

also been quantified using power spectral analysis (Lukas et al.

1982) with opioid challenges in LAAM-maintained rats producing

attenuation of the EEG voltage and a shift to a higher predominant

frequency. The technique also distinguished between high and low

maintenance doses of LAAM and, in addition, demonstrated that the

development of cross-tolerance to morphine was more complete than

that developed to methadone.

Shifts in the predominant EEG frequency during rapid-eye-movement

(REM) sleep have as well been used to track drug-seeking behavior in

30
dependent rats (Young et al. 1978). The REM sleep predominant

frequency gradually slows during the 2-hour interinjection interval

until the next self-injection of morphine when "reset" to the faster

frequency (i.e., about 1 Hz) is observed. Power spectral analysis

has also been utilized to characterize the EEG profile of the

prototypic mu, kappa, and sigma agonists (Young et al. 1981). The

different compounds are defined by a unique fingerprint-like array

based on the voltage and frequency components of the power spectrum.

Thus, the mu agonist morphine produces a large increase in voltage

over the O-10 Hz band while ketocyclazocine (a kappa agonist)

produces a definite peak in the 5-8 Hz band with lower activity in

the O-4 Hz band. (N-allyl-normetazocine, the putative sigma agonist,

produced .a power spectrum characterized by relatively much lower

voltage in the O-10 Hz band, and a small, sharp peak at 7.5 Hz.

iv) Behavioral performance

Although behavioral methods have been utilized to assess analgesic

activity (e.g., shock titration), conditioning techniques have not

generally been used to identify opioids or distinguish them from

other drug classes. Behavioral paradigms for the assessment of

opioid activity have focused mainly on the measurement of tolerance

and, more recently, in the classification of opioids based upon the

subclasses of opiate receptor activity. Antagonist-induced (i.e.,

naloxone, naltrexone) differential shifts in the dose-response

curves of opioid effects upon lever-pressing in rats on a fixed

interval food reinforcement schedule, for example, have been related

to the several presumed subpopulations of opioids (Harris 1980).

The agonist morphine was completely antagonized by naloxone or

naltrexone while butorphanol, ketocyclazocine, ethylketocyclazocine,

and low doses of cyclazocine were only partially antagonized. The

rate-decreasing effects of SKF-10,047 and high doses of cyclazocine

were not blocked by the opioid antagonists.

v) Seizure threshold

Using the flurothyl-induced seizure assay in rats, opioids have been

shown to either decrease, increase, or have no effect upon seizure

thresholds. On the basis of these differential effects, opioids

have been classified into four distinct subclasses based upon the

observed dose-response. relationships, stereospecificity, naloxone

sensitivity, and development of tolerance-cross-tolerance (Cowan et

al. 1979). The technique for flurothyl administration has been

detailed by Truitt et al. (1960) and Adler (1975). Animals are

placed in a large volume (e.g., 3-4 1) glass jar. A screen holds a

gauze pad inside the jar which is fed by plastic tubing connected to

a syringe containing a 10% v/v solution of flurothyl in ethanol.

Controlled continuous infusion of flurothyl onto the gauze is

maintained by an infusion pump at a rate of 0.103 ml/min. A fullblown generalized seizure usually occurs about 320-350 seconds after

the onset of the infusion.

The profile of flurothyl-induced seizures has been described for

rats as containing four components. The rat's typical exploratory

behavior is interrupted by short periods of "freezing" or

31
immobility. Between 150 and 200 seconds after infusion onset,

jerky, tremor-like movements prevail which are replaced by preclonic

or myoclonic jerks occurring 50-100 seconds later. Clonic movements

of the forelimbs signal the onset of the full seizure involving

both forelimbs and hindlimbs. Test compounds are administered

subcutaneously to individual rats 30 minutes before exposure to

flurothyl. The time interval between the start of the infusion and

the onset of a clonic convulsion is used as the seizure threshold.

vi) Spinal dog preparation

The chronic spinal dog assay (Martin et al. 1976) has been used to

assess the pharmacologic effects of many opioids, determine relative

potencies, and provide a means of establishing pharmacological

equivalence. The technique allows simultaneous measurement of

pupillary diameter, pulse rate, pain reflexes, touch reflexes, and

body temperature among other physiological processes. The

preparation is also capable of differentiating between naive and

opioid-tolerant animals and the measures obtained are sufficiently

sensitive to allow detection of an opiate abstinence syndrome, even

after relatively low doses. Results obtained with this technique

contributed to the formulation of opiate receptor subspecies

accounting for the effects of any given opiate by its action on one

or a combination of three receptors (Gilbert and Martin 1976; Martin

et al. 1976).

The preparation involves transection of the spinal cord at either a

high (e.g., 5th-6th cervical) or a low (e.g., lOth-11th thoracic)

level. Standard measurement of pupillary diameter, pulse rate,

respiratory rate, and body temperature follows recovery from surgery

(Martin and Eades 1964). Somatic reflexes elicited below the level

of spinal cord transection are recorded isotonically (Wikler and

Frank 1948) in the following order: patellar reflex, crossed

extensor reflex, ipsilateral extension thrust, and the ipsilateral

flexor reflex (Martin et al. 1964). The patellar reflex is evoked

using a reflex hammer applied at a rate of about once every 3

seconds. Dorgen forceps are applied across the first and fourth

toes of a hindlimb for 3 seconds in order to evoke the crossed

extensor and ipsilateral flexor reflex. Rapid extension of the toes

or pressure applied to the base of the toe pad elicits the

ipsilateral extensor thrust.

Control observations of all measures and reflexes are made 30

minutes before acute drug administration, and repeated at 15 and 30

minutes post drug and then hourly for 5-7 hours. Total scores are

obtained for each measure by summing the differences between the

before and after drug responses. In this manner, the overall

pharmacologic profile of a compound is obtained and compared to

standard compounds.

vii) Gastrointestinal motility

Acute administration of opioids causes a marked reduction in the

propulsive action of the entire gastrointestinal (G.I.) tract. This

is associated with an increase in tone, the net effect being to

32
delay gastric emptying and slow passage of the contents through the

intestines. The movement of gastric contents through the G.I. tract

can be measured fairly accurately using the charcoal meal method in

rodents (Green 1959; Cowan et al. 1977). Groups of rats are food

deprived for 18 hours, but water is available up until 1 hour before

testing. Twenty minutes after subcutaneous injection or 50 minutes

after oral administration of the test compound or vehicle control,

the rats are given a charcoal meal (5 ml/kg) via gavage. After a

standard time interval (usually 10 minutes) the rats are killed by

cervical dislocation and the distance travelled by the charcoal meal

along the small intestine from the pyloric sphincter is measured.

This distance is then converted to a percentage of the total length.

Opioids significantly slow the passage of the charcoal meal along

the intestines. Furthermore, this effect is dose-related for

morphine with a maximal effect (i.e., no distance traveled) observed

after 3 mg/kg, S.C. (Cowan et al. 1977). Buprenorphine, in doses of

0.01-1.0 mg/kg, s.c., similarly decreases the passage of the

charcoal meal, but higher doses (i.e., 10-30 mg/kg) result in a

reversal of this effect until the distance traveled is the same as

after vehicle control (Cowan et al. 1977). The procedure thus

appears to be sufficiently sensitive to detect both agonist and

antagonist effects of the opioids.

viii) Body temperature

Opiates produce marked changes in body temperature in many species.

The specific effect, however, depends on a number of factors

including species, strain, sex, and age of subject, housing

conditions, ambient temperature, degree of restraint, the particular

drug used, its dose and route of administration. (See reviews by

Burks and Rosenfeld 1979; Clark and Clark 1980; Lotti 1973).

Body temperature is commonly measured by means of a rectal

thermistor probe at regular time intervals after drug administration

(e-g., every 30 minutes for 4-6 hours). Mean changes from pre-drug

baseline and areas under the time-response curves are calculated.

Opioids thought to act on different receptor types have been studied

using variations on this basic technique in several species (e.g.,

mouse, Rosow et al. 1980, 1982a,b; rat, Geller et al. 1983; cat,

Clark and Cumby 1978, Clark and Ponder 1980, Clark et al. 1981).

For continuous measurement, either surgical implantation of a

thermistor or restraint of the animal is generally required. The

latter condition, however, has been shown to influence the

thermoregulatory effects of opiates (Holtzman and Villarreal 1969;

Martin et al. 1977). Whole-body calorimetry is a more sophisticated

technique that allows continuous measurement of several

thermoregulatory-related responses (e.g., oxygen consumption,

respiratory water loss) in addition to simple body temperature (Lin

et al. 1979).

33
ix) Pupillary effects

Change in pupil size is a well-known effect of opiate

administration. As with body temperature, the direction of change

is species-dependent. Man, rabbit, and dog respond to morphine with

miosis (constriction); cat, rat, mouse, and monkey exhibit mydriasis

(dilation). Until recently, pupil size was commonly measured

several times after drug administration by direct observation, with

calipers, or by means of still photographs (Nomof et al. 1968;

Janssen and Jageneau 1956; Gerald et al. 1976; Jasinski and Martin

1967). More precise, almost continuous measurement can now be

achieved through cinematography and infra-red videopupillometry

(Tallarida et al. 1977; Murray and Loughnane 1981; Adler et al.

1981). Frequent sampling methods are highly recommended because of

the fluctuations in pupil size induced by opioids (Henderson and

Graham 1925; Oono 1965; Klemfuss et al. 1979; Tallarida et al.

1977). The entire subject of the pupillary effects of opioids and

the methodology for studying such effects have recently been

reviewed (Murray et al. 1983).

x) In vitro procedures

These procedures deal mainly with the effects of opioids on smooth

muscle tissues such as guinea pig ileum and mouse vas deferens

(Kosterlitz and Waterfield 1975). Tissues from naive, untreated

animals are used to assess the acute effects of opioids. The animal

is sacrificed and the smooth muscle is dissected, washed, suspended

in Tyrode's solution, and affixed to an isotonic gravity lever.

Contractions are recorded kymographically or via a force transducer.

Once stable, the tissue's normal response to electrical stimulation

is determined. The procedure is then repeated in the presence of

various concentrations of opioid agonists and mixed agonists/

antagonists in order to determine relative potencies and efficacies.

The opioid-induced attenuation of the electrically produced

contraction or twitch is blocked by naloxone and serves as the basis

for the determinations of "P

A

2" values (Takemori et al. 1969).

xi) Binding assays

The affinity of opioids for binding sites in the CNS can be

determined in numerous species including rat, cat, pig, and monkey.

Current procedures, based on the original work by Pert and Snyder

radiobeled (e.g, 3H) agonist 'such as etorphine (Simon et al.

(19973) Simon et al. (1973) and Terenius (1973) utilize a

or antagonist nalaxone (Pert and Snyder 1973) which is incubated

with a suspension of whole brain synaptosomes. Alternatively,

regional binding can be determined using only selected areas of the

brain in the incubation medium, or by photoaffinity labeling (Kuhar

and Uhl 1979).

For determination of relative binding affinity, the test compound is

incubated in the medium and its affinity for the binding site is

reflected by the amount of radiolabeled etorphine that is displaced.

The test compound is also studied in the presence and absence of

sodium chloride during the incubation. Sodium ions enhance the

34
binding of agonists while decreasing the binding of antagonists

(Pert and Snyder 1973). This technique has been validated by the

demonstration that extreme values of a ratio of binding affinity in

the presence versus absence of sodium chloride predicts the

characteristics of pharmacologic action in the whole animal (Simon

et al. 1975; Woods et al. 1979).

2. Opioid Physical Dependence Potential

a. Tolerance to Drug Effects

The opioids exert a wide range of effects on numerous organs and

physiological systems. In theory, tolerance to morphine can be

assessed for any of its measurable actions; however, tolerance

develops to the many different effects of morphine, to different

degrees, and at different rates. Furthermore, it is not known

whether the mechanisms of tolerance development for each morphine

effect have a common basis, although this is often tacitly assumed.

Tolerance to morphine after repeated administration can be evaluated

by graded assay methods where a decrease in sensitivity to a given

dose of morphine can be measured, or by quanta1 assays that

determine the increase in the dose of morphine required to produce

an all-or-none effect. In general, chronic administration of

opioids leads to tolerance to both the depressant effects (e.g.,

analgesia, behavioral suppression, EEG effects) and the stimulant

effects (e.g., pupillary diameter, gastrointestinal motility).

However, the stimulant effects are less sensitive to the development

of tolerance. Since it appears that the degree of tolerance

developed is directly proportional to the duration of exposure to

the compound (both dose- and time-related), methods for maintaining

optimal exposure have followed two basic strategies: multiple

injections and continuous exposure.

Multiple injections of morphine have been given intraperitoneally,

subcutaneously, intravenously, and intracerebrally to produce

tolerance, although under certain conditions it is possible to

demonstrate tolerance after a single injection. In general,

however, a high degree of tolerance is attainable only after

frequent repeated injections of high doses of morphine over several

weeks. In recent years, the morphine pellet implantation procedure

(Way et al. 1969) has been widely used to produce a high degree of

tolerance and physical dependence, as well, in the mouse, rat, and

guinea pig within three days. One or more specially formulated

pellets containing 75 mg morphine base (Gibson and Tingstad 1970)

are inserted subcutaneously in the back of the animal after making a

small incision.

After implantation of the morphine pellet(s), animals exhibit the

characteristic signs of acute morphine effects. In mice this

consists of the typical Straub tail and increased motor activity

within 30 minutes after implantation of a single pellet. By 24

hours, the acute effects have subsided and the general behavior of

the morphine-implanted mice resembles that of placebo -implanted

controls. In rats, the typical acute narcotic effects are also

35
observed shortly after implantation of 1 or 2 pellets. These signs

include a cataleptic state, exophthalmos, and shallow respiratory

movements (Wei and Way 1975). In the guinea pig, after implantation

of 4 morphine pellets, the animals become sedated for 24 hours and

have a decreased reaction time to a thermal stimulus when tested by

the hot-plate method. Reaction times return to normal 14 hours

after implantation. A biphasic change in body temperature and some

body weight loss were also observed during the first 2 days after

implantations (Goldstein and Schulz 1973). In all three species,

tolerance and physical dependence are maximal on the third day after

pellet implantation.

In vitro procedures are also available for studying opioid

tolerance. Tolerance development in the ilea of guinea pigs can be

induced by implanting morphine pellets subcutaneously (Goldstein and

Schulz 1973; Huidobro-Toro et al. 1978). A supplementary amount of

agonist should be added to the media to prevent abstinence when the

preparation is washed. Tolerance in vitro can also be produced in

this preparation by incubating segments of excised ileum from naive

animals with the agonist at 37OC for 1 to 4 hours (Rezvani et al.

1983); the effect is stereospecific for the active l-isomers and can

be prevented by naloxone. Development of tolerance to one opioid is

accompanied by cross-tolerance to other opioids of the same

subclass.

b. Withdrawal from Chronic Drug Administration

An abstinence syndrome is represented by a 'constellation of

behavioral signs that is peculiar to a given species. Not all the

signs will appear in one subject and certain of the signs may be

exhibited by a nondependent one. In any assessment, therefore, the

selection of the signs to be observed and their relative rank

ordering of importance are sometimes made arbitrarily. Several

dimensions can be considered, including the frequency of occurrence,

the intensity of each event, and the probability that such a sign

would occur in a given population. For example, withdrawal jumping

is a characteristic abstinence sign in the mouse (Collier et al.

1972). The mean number of jumps per mouse, the height of each jump,

and the incidence of jumping may all be used to quantify the

withdrawal behavior.

In choosing a particular sign of withdrawal for judging dependence

intensity, the possibility that experimental manipulation of the

dependent state might selectively affect only the withdrawal sign

and not the total syndrome must always be kept in mind. This

criticism can be met by using several withdrawal signs. However,

the rates of development of withdrawal signs are not always parallel

and certain intense abstinence signs may suppress the appearance of

other signs (Blasig et al. 1973).

The abrupt withdrawal syndrome is difficult to quantify because the

protracted course of abstinence requires extended observation for

several days. The slow onset and prolonged course of abrupt

morphine withdrawal have led to the increasing use of antagonistprecipitated withdrawal for evaluating physical dependence. The

36
syndrome produced by opiate antagonists, in contrast to abrupt

withdrawal, is a rapid, explosive event that condenses in a short

time period the abstinence signs of abrupt withdrawal.

Withdrawal signs that appear upon termination of chronic exposure to

an opioid fall into three basic classes: autonomic (e.g., blood

pressure, pulse, diarrhea, respiratory rate, pupil diameter, body

temperature), somatomotor (e.g., nociception, various neuromuscular

reflexes, Straub tail, convulsions), and behavioral (e.g.,

irritability, eating and drinking, sleep, degree of alertness). In

general, the autonomic signs appear first and, in the absence of

other signs, suggest a mild abstinence syndrome. The appearance of

somatomotor effects constitutes a more intense syndrome. Behavioral

Changes are observed throughout the course of withdrawal, though

they may make their initial appearance after the early autonomic

signs.

The dependence potential of opioids has been quantified in numerous

species including mice, rats, dogs, and monkeys. The procedures

most commonly used to assess withdrawal effects with opioids involve

rating autonomic signs, seizure threshold, reflexes, body weight,

body temperature, and EEG activity. Body weight in rats falls

following either abrupt or precipitated withdrawal and often amounts

to as much as 10% of the pre-withdrawal weight. Maximum changes are

noted about 1-2 hours after precipitated and 48 hours after abrupt

withdrawal (depending on half-life of the opioid). Other species

show different time courses of effect depending on pharmacokinetic

factors (Akera and Brody 1968; Wei and Way 1975). The antagonistinduced jumping that occurs in opioid-dependent rodents can be

utilized to provide a sensitive, precise measure of one sign of

physical dependence. Utilizing as an index the dose of naloxone

(ED50) to precipitate withdrawal jumping in dependent mice, a

decrease in the naloxone ED50 over time by 70-fold has been

demonstrated as animals become increasingly dependent on morphine

after pellet implantation (Way et al. 1969).

Numerous methods have been used to maximize the animal's exposure to

opioids, and thus induce dependence. Oral administration remains a

favored route because of its ease and minimal maintenance. This is

accomplished either by gavage (Stolerman and Kumar 1970),

intragastric catheter (Lukas et al.1982), or in the drinking water

(McMillan et al. 1974). Continuous exposure to opioids has also

been accomplished by programmed intravenous injection (Weeks 1962),

intraperitoneal injection using a chronic i.p. catheter (Teiger

1974), reservoir implantation (Goode 1971), and pellet implantation

(Wei et al. 1973), with the development of miniature osmotic pumps

(e.g., Alzet) adding to the sophistication of such implantation

procedures (Wei and Loh 1976).

i) Primary physical dependence

Methods for assessing the dependence potential of morphine in rats

were developed by Akera and Brody (1968). While the procedure for

other species varies, the basic method is as follows:

evenly spaced injections should be used, but dependence can also

3 7
easily be produced with two injections per day: 1 in the morning

and 1 in the late afternoon or evening. As tolerance to the

sedative and depressant effects develops, the maintenance dose is

increased. Once the desired dose and duration of exposure are

reached, the withdrawal syndrome can be observed either by stopping

the injections (abrupt or spontaneous withdrawal) or by injecting a

small dose of the opioid receptor antagonist, naloxone (precipitated

withdrawal). In general, spontaneous withdrawal is characterized by

slow onset (hours), relatively moderate intensity, and long duration

(hours-days) while precipitated withdrawal has a rapid onset

(minutes), relatively severe intensity, and a relatively short

duration (1-2 hours).

ii) Single dose substitution

This method was first described for opiates by Seevers (1936) and

has been modified by a number of laboratories (Yanagita 1973).

Rhesus monkeys are treated with morphine at a dose of 3.0 mg/kg,

subcutaneously four times a day. These animals are maintained on

this regimen for long periods of time and thus acquire a relatively

stable dependence upon morphine. Minimum exposure duration is 60

days. On a test day, the regularly scheduled morphine injection is

withheld until morphine abstinence signs of intermediate intensity

are present. This is typically 14 hours after injection. Then a

dose of test compound or morphine is 'given S.C. and the abstinence

scores are rated on the degree of change from pre-injection. A drug

with morphine-like dependence potential will suppress the withdrawal

signs while a drug with little or no opioid agonist activity will

not affect the progressively more intense abstinence signs. The

animals are evaluated l/2, 1, 2, and 3 hours after administration or

until they have returned to the pre-injection level. When a lower

maintenance dose of morphine (e.g., 0.3 mg/kg, s.c.) is used,

specific differences between pure agonists and mixed agonist/

antagonist analgesics can often be detected. Low-dose morphineinduced dependence is maintained by mixed agonist antagonists, while

these compounds actually precipitate withdrawal in high-dose

morphine-maintained animals. Martin and co-workers (1974) have

established a standardized bioassay procedure for assessing the

relative potencies for suppressing morphine-abstinence signs.

3. Opioid Abuse Liability

a. Drug self-administration

Procedures for assessing self-administration of opioids have been

described using rats (Weeks 1962; Khazan et al. 1967; Moreton et al.

1976). dogs (Jones and Prada 1973), and non-human primates (Thompson

and Schuster 1964; Deneau et al. 1969; Woods 1980). Most studies

have been conducted using animals that have been prepared with an

indwelling intravenous catheter. The animals are given an injection

which is contingent upon completing a required number of lever

responses (i.e., fixed ratio) or following a single response after a

preset time interval (i.e., fixed interval). Two basic procedures

are employed: continuous or direct, and substitution. A factor

relevant to the conduct of both types of studies is whether opioid

38
dependence must first be present before morphine or its surrogates

will be self-administered.

i) Continuous self-administration

In this procedure (Woods and Schuster 1968) rhesus monkeys are first

prepared with an indwelling intravenous catheter and then restrained

within a cage using an extension arm and harness system (Yanagita et

al. 1965). The availability of morphine for self-injection by lever

pressing is indicated by illuminating a small light above the lever.

The schedule is then changed to a variable interval 2.5 minutes (the

first response after an average interval of 2.5 minutes results in

drug delivery). After a period of 15 days, the drug solution is

replaced by physiologic saline. Another dose of morphine is then

tested in a similar manner. In some instances, to avoid overdose,

the maximal number of self-injections allowed is limited to four.

Using this procedure, the unit dose of 10 mg/kg/inj was observed to

maintain self-injection performance though no signs of abstinence

were observed during saline exposure. In contrast, self-injection

of higher doses resulted in a distinct abstinence syndrome when

saline was substituted for morphine.

An alternative to this procedure permits access to relatively low

doses of an opioid to determine whether drug intake increases with

duration of exposure in the self-administration paradigm. This

procedure is useful in determining whether tolerance develops to the

appetitive or aversive properties of the drug.

ii) Substitution

In this procedure, drug self-administration is initially maintained

by a standard compound (usually cocaine, morphine, or codeine), and

test doses of experimental compounds are substituted upon attainment

of stable baseline performance. As a control, the drug's vehicle or

saline is also substituted for the same duration of time in order to

monitor the extinction of drug-reinforced responding. Comparisons

can then be made between the response rates maintained by the

standard compound, saline, and the test compound (Woods 1980).

Availability of drug for self-injection is usually indicated by

illumination of a light or activation of a tone or buzzer. Upon

completion of the response requirement (usually fixed ratios of 30

or 160 responses), the standard drug is delivered via a peristaltic

or syringe pump. After stable responding for the standard drug has

been maintained, a dose of the test drug is substituted for varying

periods from 1 to 15 days. The number of self-injections and the

response rates for injection are quantified and serve as the basis

for determining abuse liability. Additional doses and test drugs

are evaluated in a similar manner.

iii) Reinforcing efficacy analysis

Extensions of the substitution procedure for evaluating the relative

abuse liability of opioids have included only a few procedures.

Response rate analysis (Woods 1980; Aigner and Balster 1979) and

39
injection patterns remain the major criteria for rank-ordering

compounds with respect to their reinforcing efficacy. Yanagita et

al. (1982) using a progressive ratio procedure demonstrated that

buprenorphine maintained lower break points than pentazocine.

Choice procedures have been used to determine the relative

reinforcing efficacy of various doses of heroin (Griffiths et al.

1981). and a recent report (Mello et al. 1981) has described the use

of second order schedules to demonstrate the reinforcing properties

of buprenorphine.

iv) Tolerance and physical dependence

The relative potency of an opioid is decreased with tolerance

development (i.e., higher doses are necessary to maintain

self-injection performance), but it is not clear whether the

reinforcing properties of the drug have changed, or, alternatively,

the animals must titrate their level of dependence. Two types of

procedures are available to avoid such problems. First, drugs can

be tested in prevously opioid-dependent (but presently nondependent) animals to minimize any differences which may be due to

the animal's drug history (Steinfels et al. 1982). Secondly,

cocaine can be used as the standard drug so that the opioid

tolerance and/or dependence normally associated with codeine or

morphine does not develop. This consideration is important when

mixed agonist/antagonist analgesics are the test compounds, since

withdrawal may be precipitated upon substitution in an opioiddependent animal.

b. Drug discrimination

Procedures for assessing the discriminitive stimulus properties of

opioids have, for the most part, been developed for the rat and

monkey, but the pigeon and gerbil have also been utilized (see

reviews Holtzman 1982, 1983). Two basic methods have been employed:

shock avoidance (Shannon and Holtzman 1976) and food presentation

(Herling and Woods 1981).

In the shock avoidance procedure, animals typically must first press

a "starting" lever after which they must press the appropriate lever

on the other side of the cage in order to prevent shock delivery.

The appropriate lever is determined by whether the animal has

received a saline injection (e.g., left lever appropriate) or an

injection of the training drug - usually morphine (e.g., right lever

appropriate). The similarities in the stimulus properties of other

opioids are then tested over a wide dose range, with 90% drugappropriate responding usually being the accepted criterion for

discrimination.

In the food presentation paradigm, monkeys are usually maintained at

80-9096 of their free-feeding weight and then trained to press one of

two levers in order to receive a food pellet (Herling and Woods

1981; Woods et al. 1982). The response requirement is usually 20 to

30 lever presses (i.e., FR 20-30) and the appropriate lever is

determined by whether the animal has received an injection of an

40
opioid (e.g., either etorphine or morphine) or saline. Training is

continued until 90% or better appropriate responding is maintained.

Using these procedures, a large number of opioids have been

characterized with respect to their discriminative stimulus

properties (Holtzman 1982; Woods et al. 1982). In order to fully

characterize the conglomerate opioid class of drugs (viz a viz, mu,

kappa, and sigma agonists and mixed agonists/antagonists), however,

it is important that the protocol includes animals that have been

trained on drugs other than the pure agonists morphine and etorphine

(e-g., cyclazocine, ethylketocyclazocine, N-allylnormetazocine). In

addition, different doses of these compounds should be used in order

to be able to characterize the full spectrum of a drug's effects

(Holtzman 1982). It is also useful to determine to what extent the

discriminative stimulus properties of these compounds can be blocked

by opioid antagonists such as naloxone or naltrexone. Finally, the

animal species utilized must be taken into account, since distinct

differences have been observed particularly between the rat, pigeon,

and monkey. Drug discrimination procedures have nonetheless been

used effectively to differentiate a number of opioids in three

classes: opioids that generalize to morphine and are antagonized by

naltrexone or naloxone; opioids that generalize to cyclazocine and

are not easily antagonized; and opioids that generalize to

cyclazocine and phencyclidine and are not antagonized (Holtzman

1982).

c. Behavioral Toxicity

To date, no procedures such as those outlined in the general

methodological section have been employed to systematically

investigate the adverse sensory or motor effects of various opioids

in animals. One study, using size selection discrimination of a

visual stimulus, showed that morphine in doses of 1-10 mg/kg

decreased reaction times in rhesus monkeys (Brown and Bass 1967).

This finding is consistent with other reports that morphine

possesses both depressant and stimulant effects which may be

dose related.

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4 7
C. Procedures for Assessing CNS Depressants: Sedative/

Hypnotics, Anesthetics, Aaxiolytics, and Antihistamines

1. Characterization of CNS Depressant Effects

a. Acute Drug Effects

i) Sleep Induction

This procedure determines the hypnotic potency of a compound in

addition to characterizing the nature of the sleep state (e.g.,

light, intermediate, deep). Typically, the fasted mouse is used and

the compound is administered orally, although other species and

routes may be employed. The procedure described by Gruber et al.

(1944) initially begins testing with a dose empirically derived

during pilot studies. If sleep is produced, as evidenced by loss of

righting reflex, a series of decreasing doses is given (ten

different mice per dose) and the ED50 is determined. Additional

data obtained include the onset and duration of sleep (from the loss

to return of the righting reflex) and the duration of symptoms

following hypnosis.

An extension of this procedure may be used to identify compounds

that prolong or potentiate the sleep time of a test compound. Two

groups of fasted mice are used; one group receives 10% of the ED50

dose (determined as above) of a compound identified as a sedative

and the other group receives vehicle. Thirty minutes later both

groups receive pentobarbital sodium at a dose of 6 mg/kg i.p.,and

the onset and duration of hypnosis are observed and comparisons are

made between the two groups.

ii) Spontaneous Locomotor Activity

The procedures described in the general methods section above are

most commonly used to assess the pharmacological activity of these

compounds which decrease spontaneous locomotor activity in a

dose-related manner. No distinction between compounds within the

depressant class can be made using this test, though differences in

duration of action can be detected.

iii) Electroencephalographic (EEG) Activity

Using the procedures outlined in the general methods section above,

CNS depressants have been shown to induce characteristic shifts in

the power of various frequency bands. These shifts have basically

consisted of an increase in the overall power, with the greatest

increase occurring in the 8-13 and 13-25 Hz bands (Gehrmann and

Killam 1976). This contrasts with the narcotics which typically

increase power in the O-4 Hz band (Lukas et al. 1980). Using this

procedure, various compounds including barbiturates,

benzodiazepines, and other depressants have been classified and

compared with respect to the particular shifts in power and

predominant frequency after their administration (Gehrmann and

Killam 1976, 1978; Joy et al. 1971; Schallek and Johnson 1976;

Schallek et al. 1967). Benzodiazepines have a characteristic EEG

48
effect and change the sleep-wakefulness cycles and continuum.

Further, one can couple behavioral effects with EEG changes

(Gehrmann and Killam 1975).

iv) Behavioral Performance

While numerous operant paradigms have been employed to study the

depressant effects of these compounds (e.g., ratio or interval

schedule responding for a reinforcer such as food or water), the

most widely used method for studying the anxiolytic effects of CNS

depressants, particularly the sedative/hypnotics, is the conflict

procedure. First introduced by Geller and Seifter (1960), the model

was further developed and refined (Cook and Davidson 1973) and

subsequently validated by Cook and Sepinwall (1975) as a predictor

of anxiolytic effects. This validation was based upon the good

correlation between the relative potencies of numerous clinically

effective anxiolytic agents (e.g., diazepam, oxazepam,

chlordiazepoxide, phenobarbital, amobarbital, and meprobamate) and

their attenuation of punishment effects upon operant responding.

Basically, the procedure involves imposing a punishment (usually

electric foot shock) contingency upon a stable performance baseline

(usually a variable interval schedule for food). The schedules can

be either alternating or concurrent, with better separation of

performance obtained using the latter. The introduction of shock

suppresses responding for food, and the administration of a minor

tranquilizer attenuates the suppressive effects of the shock. Thus,

responding during shock is actually increased after treatment with a

benzodiazepine or a barbiturate; this selective effect is typically

observed at doses which have little or no effect on non-punished

responding (e.g., Cook and Sepinwall 1975). This procedure appears

to be relatively specific for sedative/hypnotics, while major

tranquilizers (e.g., chlorpromazine), stimulants (e.g.,

amphetamine), antihistamines (e.g., diphenhydramine).

anticonvulsants (e.g., phenytoin), and opioid analgesics (e.g.,

morphine) lack significant effect. The procedure does not, however,

discriminate between the barbiturates, ethanol, and the

benzodiazepines.

v) Binding Assays

While the specific mechanism of action of CNS depressants has not

been completely elucidated, one common feature is that they appear

to interact with gamma-aminobutyric acid (GABA). Currently, no

specific neuronal sites have been shown to bind selectively to

barbiturates. However, they have been shown to increase GABA

activity in a number of in vitro preparations (Eccles et. al. 1971;

Nicoll 1978) and more recently, both anesthetic and convulsant

barbiturates enhance GABA binding to rat brain synaptosomes (Willow

and Johnston 1981).

In contrast, a receptor that specifically binds' benzodiazepines has

been identified (Squires and Braestrup 1977; Mohler and Okada 1977)

and it has been shown that the binding of GABA to its receptor is

enhanced in the presence of benzodiazepines (Haefely 1977).

Furthermore, validation of the benzodiazepine binding assay has been

49
provided by a number of studies (Braestrup and Squires 1978; Mohler

et. al. 1978) in which the affinities of benzodiazepines for the

binding site correlate well with their pharmacological activities

and clinical potencies.

Essentially, the procedure for assessing the affinity of test

substances for the benzodiazepine receptor is conducted using

tritiated diazepam or flurazepam. Appropriate neuronal tissues are

incubated with the radiolabeled ligand, and various concentrations of

the test compound are added. The relative binding affinity of the

test compound is directly related to the amount of labeled diazepam

(or flurazepam) that is displaced. Displacement of the labeled

compound is indicated by a decrease in the disintegrations per unit

time emitted from the tissue after washing.

The discovery of these specific binding sites has led to the search

for an endogenous anxiolytic ligand. Historically, numerous

compounds (e.g., the beta carbolines) have been suggested as likely

candidates, but no convincing evidence exists to date as to the

existence or identity of this substance. A more recent finding that

the benzodiazepine derivative, ethyl-8-fluoro-5,6-dihydro-5-methyl6-oxo-4H-imidazo[1,5a][l,4]benzodiazepine-3-carboxylate (Ro 15-1788)

can bind to the receptor and block numerous pharmacological effects

of benzodiazepines (Hunkeler et al. 1981; Darragh et al. 1981;

Mohler et al. 1981; Polc et al. 1981), however, 'provides a unique

and potentially powerful tool for investigating the acute effects of

benzodiazepines and, ultimately, for characterizing the withdrawal

syndrome (Lukas and Griffiths 1982).

2. CNS Depressant Physical Dependence Potential

a. Tolerance to Drug Effects

Sedation is the, primary pharmacological effect of this class of

compounds. Upon repeated exposure to a standard dose, tolerance

develops as seen by an attenuation of the degree and duration of CNS

depression. According to Kalant et al. (1971), tolerance may be

dispositional or functional. The former is primarily due to an

increased rate of drug metabolism and elimination from the body

while the latter refers to a diminished sensitivity of the CNS. The

extent to which each mechanism contributes to the overall observed

degree of tolerance is unclear. For some drugs, at least,

information on the relative contribution of these factors can be

obtained by conducting pharmacokinetic studies in conjunction with

the pharmacological assays. Thus, drug disappearance curves could

be constructed that would match (or in some cases, not match) the

time-effect curves. While dispositional tolerance may be relatively

important when assessing pentobarbital (see review, Conney 1967), it

cannot account for both the tolerance developed to barbital (which

is only slightly metabolized) and the development of tolerance after

intraventricular administration (Stolman and Loh 1975; Lyness et al.

1979). By employing a "maximally tolerable dose" technique, Okamoto

and co-workers (1975) have been able to separate the two components

of barbiturate tolerance. Essentially, they demonstrated that

50
dispositional tolerance develops very quickly while functional

tolerance develops more gradually.

The degree of tolerance that can be attained is directly related to

the time intervals between exposures to the drug (Gruber and Keyser

1946). Numerous techniques have been employed to maximize the

exposure of the CNS to depressant drug, including: continual

parenteral administration of barbiturates (Kato et al. l964), adding

barbiturates to the drinking water (Crossland and Leonard 1963) or

to food (Belknap et al. 1973). implantation of a barbiturate pellet

(Ho et al. 1975), or delivery of a barbiturate via a subcutaneously

implanted miniature osmotic pump (Siew and Goldstein 1978).

Whichever method is used, it is apparent that tolerance develops to

the mild behavioral effects (e.g., loss of fine motor control, mild

ataxia) of barbiturates but not to the lethal effects (see Okamoto

and Boisse 1981). Thus, as tolerance develops to a CNS depressant,

the therapeutic index decreases.

Numerous animal species have been used to study barbiturate

tolerance and graded rating scales for CNS depression have been

developed and utilized in the rhesus monkey (Yanagita and Takahashi

1970) and the cat (Okamoto et al. 1975). The effects most often

studied include: ataxia. decreased locomotion, decreased

respiratory rate, impaired righting reflex, and changes in cornea1

and linguomandibular reflexes and nictitating membrane tone.

Regardless of the response -studied. it is important that studies of

tolerance include: appropriate dosing schedules and route of

administration, full dose-response evaluations, and appropriate

controls to account for order effects. In addition, measures of

cross-tolerance to another drug within the same class as well as in

a different class are extremely important in characterizing the

overall profile of a drug.

b. Withdrawal from Chronic Drug Administration

Withdrawal from chronic administration of numerous sedative/

hypnotics has been assessed in various animal species including mice

(Ho 1976), cats (Okamoto et al. 1975), dogs (Deneau and Weiss 1968).

and monkeys (Yanagita and Takahashi 1973). While. species

variability generally exists, the sedative/hypnotic abstinence

syndrome is basically characterized by gross behavioral

manifestations which include anorexia, hyperirritability, tremors,

and convulsions. These signs have been grouped into three levels of

withdrawal: Mild - hyperirritability, mild tremor, anorexia,

piloerection; Intermediate - aggravated tremor, muscle rigidity,

impaired motor activities, retching or vomiting, weight loss (10%);

Severe - convulsions, hallucinatory behavior, nystagmus,

unresponsiveness to environmental stimulation, hyperthermia,

morbidity, and mortality.

Another technique that has been utilized to quantify barbiturate

dependence potential is seizure susceptibility. During barbiturate

withdrawal a decrease in seizure threshold has been observed after

the administration of pentylenetetrazol (Jaffe and Sharpless 1965).

and bemegride and picrotoxin (Crossland and Turnbull 1972).

5 1
Barbiturate withdrawal has also been assessed using audiogenic

seizures (Crossland and Turnbull 1972; Gates and Chen 1974) and

flurothyl-induced seizures (Greer and Alpern 1976). Kindled

seizures can also be used for this purpose. Alcohol withdrawal has

been shown to increase the susceptibility to seizures in animals

with previously kindled epileptic foci in the amygdala (Pine1 et al.

1975, 1978).

i) Primary Physical Dependence

Nethods for assessing the dependence potential of barbital in naive

dogs were developed by Seevers and Tatum (1931) and later extended

by Fraser and Isbell (1954). The production of primary dependence

on sedative-hypnotic drugs is directly related to the dose, dosing

interval, duration of treatment, and the route of administration.

Typically, 100 mg/kg of barbital sodium in 4 divided doses is given

orally for 60-90 days. It is sometimes necessary to initiate

treatment with lower doses and gradually increase the dose as

tolerance develops. When comparisons between different drugs are to

be made, it is important that the dosing schedules are adjusted such

that the pharmacological effects of the drugs are equivalent' (e.g.,

see Okamoto et al. 1975). Quantitative measures of withdrawal signs

are equally important when attempting to compare the dependence

potential of numerous compounds. In this regard, the method of

Jones and co-workers (1976), which describes a bioassay for sedative

dependence potential, appears to be the most sophisticated and

perhaps the best validated system to date.

In general, comparative studies indicate that the use of monkeys and

carnivores provides clearer and more profound withdrawal effects,

but that rodents offer the advantage of being cost effective. This

is especially true during preliminary screening procedures where a

large number of experimental subjects is required. Historically,

the assessment of the dependence potential of a compound required

that exposure to the drug be terminated. The recent discovery of

the specific benzodiazepine receptor antagonist (Ro 15-1788), that

can block numeruds pharmacological effects of the benzodiazepines

(Hunkeler et al. 1981; Darragh et al. 1981; Mohler et al. 1981; Pole

et al. 1981), has provided a unique and potentially powerful tool for

studying the benzodiazepine withdrawal syndrome.

Administration of Ro 15-1788 to baboons that had been treated with

diazepam (20 mg/kg/day) via an intragastric catheter for only 7 days

produced numerous signs of withdrawal including bruxism, nose

rubbing, retching and vomiting, abnormal body postures, tremors and

convulsions (Lukas and Griffiths 1982). The withdrawal signs were

more frequent and of greater intensity when the duration of diazepam

exposure was increased to 35 days. Furthermore, when compared with

spontaneous withdrawal from diazepam, the Ro 15-1788 precipitated

abstinence syndrome was characterized by a relatively rapid onset

(i.e., 7-10 minutes), intense signs, and short duration (i.e., 4-8

hours). In contrast, spontaneous withdrawal (obtained by simply

stopping the diazepam) was characterized by a slow onset (i.e., 5-7

days), relatively milder signs, and a long duration (i.e., 8-13

days). Ro 15-1788 has also been used to precipitate withdrawal

52
signs in benzodiazepine-treated mice, rats, cats, and squirrel

monkeys (Rosenberg and Chiu 1982; Cumin et al. 1982; McNicholas and

Martin 1982).

ii) Single Dose Substitution

This method was first described by Deneau and Weiss (1968) and

involved the administration of barbital sodium (100 mg/kg/day) to

dogs for periods of up to 2 years. The termination of drug resulted

in the appearance of withdrawal signs, the severity of which was

assessed using an objective point-scoring system. A test compound

was then administered (at a dose previously determined during pilot,

acute studies) for 5 consecutive days and its effectiveness in

reversing the withdrawal signs was determined. A total of twelve

compounds were evaluated in this manner and all except glutethimide

were found to completely substitute for barbital. After the 5-day

substitution period, the test compound was withheld and the

abstinence signs were quantified. Similar methods have been

developed for the monkey (Yanagita and Takahashi 1973), cat (Okamoto

et al. 1975), and rat (Essig 1966). More recently, Jones et al.

(1976) have developed a quantitative bioassay for comparing the

dependence potential of various CNS depressants in the dog. In

order to effectively compare the dependence potential of various

compounds, it is necessary to incorporate procedures which ensure

that the drugs are functionally equivalent; two drugs are

functionally equivalent only if they produce the same peak,

residual, and total degree of CNS depression.

3. CNS Depressant Abuse Liability

a. Drug Self-Administration

Procedures for assessing self-administration of sedative/hypnotics

have been described using rats (Walton and Deutsch 1978), and

non-human primates (Yanagita and Takahashi 1970, 1973; Griffiths et

al. 1981). While the intravenous route has been employed most

often, the development of other methods including intragastric

(Yanagita and Takahashi 1973) and oral (Ator and Griffiths 1983)

self-administration has recently been reported. Drug injections are

usually contingent upon completion of a fixed number of responses on

a lever (i.e., fixed ratio schedules), but interval and second-order

schedules have also been used effectively. Two basic procedures are

employed: continuous or direct, and substitution.

i) Continuous self-administration

In this procedure (e.g., Yanagita 1976) the intravenous route is

preferred as long as solubility characteristics permit. The

subjects (usually monkeys) are first given the opportunity to

self-inject the drug vehicle alone, and once low and stable response

rates are achieved, a dose of the compound to be tested replaces the

vehicle at one-quarter to one-half the minimal effective dose (as

determined during the acute studies) for 2 to 4 weeks. If, at the

end of this period, the animal still has not increased its rate of

responding, then an automatic injection schedule is superimposed on

53
the self-injection schedule for 2 more weeks. Response rates and

daily self-injections are monitored during this period. Then the

automatic injections are terminated and the self-injection unit dose

is decreased by one-half to one-fourth to determine whether response

rates increase. Finally, the subject is again exposed to 'the

vehicle alone while response extinction is characterized and the

subject is observed for possible withdrawal signs. Using this

procedure, pentobarbital and alcohol were reported to be selfadministered at moderate rates while diazepam, chlordiazepoxide, and

oxazepam were self-injected at lower rates (Deneau et al. 1969;

Yanagita and Takahashi 1973; Yanagita 1976).

ii) Substitution

In this procedure, drug self-administration is initially maintained

by a compound known to reinforce self-injections (e.g., cocaine,

codeine) and test doses of experimental compounds are substituted

upon a stable baseline performance. As another control, saline or

the drug vehicle is also substituted in order to assess extinction

as well as to serve as a measure of the animal's operant level of

responding. Comparisons are then made between self-injection rates

following saline substitution and substitution of an appropriate

range of doses of the test compound.

While the number of responses required for drug- delivery in the

continuous procedure is usually only one, larger numbers (e.g., 30,

160) are typically employed in substitution procedures. In

addition, the opportunity to self-inject a compound is usually

limited either by programming brief sessions (Schuster and Thompson

1969) or by imposing time-out periods between drug injections

(Griffiths et al. 1981) as compared to ad libitum availability in

the continuous self-administration procedure. This latter procedure

tends to reduce the confounding effects of previous drug injections

upon operant behavior, and thus provides a more reliable measure of

the extent to which the "drug-seeking" performance is maintained by

the reinforcing properties of the drug per se.

Using the substitution procedure and cocaine (0.32 mg/kg/injection)

as a baseline drug (fixed ratio 160 schedule of reinforcement and a

3-hour time-out after each drug self-injection); Griffiths et al.

(1981) showed that the three barbiturates secobarbital.

pentobarbital, and amobarbital all maintained rates of responding

that were comparable to those maintained by cocaine. In addition,

numerous benzodiazepines (e.g., diazepam, clonazepam, chlorazepate,

flurazepam, and medazepam) sustained rates of responding that were

higher than vehicle control, but lower than those of cocaine.

iii) Intragastric, oral, and inhalation

self-administration

Yanagita and Takahashi (1973) reported that rhesus monkeys selfadministered pentobarbital and alcohol via an intragastric catheter,

though lever pressing rates were not as high as when the compounds

were given intravenously. In this same study, intragastrically

delivered diazepam maintained more robust self-administration than

5 4
either chlordiazepoxide or oxazepam. Using a similar procedure

Altshuler et al. (1975) found that rhesus monkeys would selfadminister chlordiazepoxide intragastrically, but not diazepam.

Gotestam (1973) and Davis et al. (1978) reported that

intragastrically administered medazepam and chlordiazepoxide,

respectively, were reinforcing to rats while Walton and Deutsch

(1978) found no evidence for intragastric or oral selfadministration of diazepam.

The difficulty with intragastric self-administration studies appears

to reside in factors relating to drug delivery. Typically,

absorption from the stomach is relatively slow, and previous studies

by Stretch et al. (1976) have shown that response rates for cocaine

self-injection decrease when the delivery of the drug is delayed

after completing the operant task. Thus, the onset of drug effect

may in fact be anywhere from 5-30 minutes (depending on the drug)

after the injection. Controlled fasting prior to test sessions (to

maximize drug absorption) and appropriate stimuli associated with

drug delivery may 'facilitate intragastric self-administration, and

the use of second order schedules may prove helpful.

Because of taste and volume factors associated with drug solutions,

it has been difficult to conduct oral self-administration studies

(Wolf et al. 1978). Forced consumption of chlordiazepoxide in rats

has been accomplished by making the drug solution the only source of

fluid (Harris et al. 1968; Kamano and Arp 1965). This procedure,

however, has not been effective in inducing higher intake of

chlordiazepoxide during testing. An alternative procedure employs a

schedule in order to induce high fluid intake (Falk et al. 1972;

Sanger and Blackman 1978). This paradigm (schedule-induced

polydipsia) has been used to enhance chlordiazepoxide consumption in

rats (Sanger 1977) and ethanol (Henningfield et al. 1981) and

methohexital (Ator and Griffiths 1983) in primates.. During a 3-hour

session in which water was available via a drinking spout, the

entire daily ration of food biscuits was delivered at the end of the

first hour in the study by Ator and Griffiths (1983). This

procedure induced a large increase in drinking during the remaining

2 hours. Then, on subsequent days, increasing concentrations of

methohexital were substituted in place of the water. At a

methohexital concentration of 0.8 mg/ml, the volume consumed in the

first hour of the session (before food delivery) typically exceeded

30% of the session total and overt behavioral effects (e.g., ataxia)

were observed. Drinking and drug intake remained at high levels

after the food-inducing procedure was discontinued and food was made

available at the end of the session.

Administration of CNS depressants via inhalation as a means of

assessing abuse liability has been studied only in the rhesus monkey

(Yanagita et al. 1969). The subjects were fitted with an intranasal

catheter which provided a means by which lacquer thinner fumes could

be forced into the lungs by compressed air. The animals pressed a

lever, received a Z-minute exposure. to chloroform or ether, or 5

minutes of lacquer thinner. Goldstein (1972) has also used the

inhalation route to render mice tolerant and dependent upon ethyl

55
alcohol, but the procedure has not, as yet, been employed routinely

as a self-administration paradigm.

iv) Reinforcing efficacy analysis

Extensions of the substitution procedure for evaluating relative

sedative/hypnotic abuse liability have involved the application of

numerous procedures including progressive ratio performance (Brady

et al. 1975; Yanagita 1976). response rate analysis (Griffiths et

al. 1981), concurrent or choice schedules (Findley et al. 1972), and

second order schedules (Kelleher 1975). Two basic findings have

evolved from these studies. First, barbiturates appear to maintain

higher levels of performance than benzodiazepines, though not enough

comparative data is available to rank-order depressants belonging to

other pharmacological classes. Secondly, compounds with short

durations of action appear to have a greater reinforcing efficacy

than their longer-acting analogues.

v) Tolerance and physical dependence

There is little or no data available on the effects of tolerance and

dependence on the reinforcing efficacy of CNS depressants,

hypnotics, or anxiolytics, nor on the effects of a prior history of

dependence in this regard. Griffiths et al. (1981) have reported,

however, that there was no difference in the benzodiazepinemaintained self-administration performance of baboons before and

after barbiturate self-administration.

b. Drug Discrimination

Procedures for assessing the discriminative stimulus properties of

sedative/hypnotics have been described in pigeons (Herling et al.

1980), gerbils (Jarbe and Holmgren 1977), rats (Overton 1979). and

primates (Ator and Griffiths 1982; Winger and Herling 1982). The

most frequently used procedures in rodents have involved either a

T-maze (e.g., turn left if drugged; turn right if not drugged) or a

two-lever choice situation (e.g., left lever produces food or

terminates shock if drugged; right lever produces food or terminates

shock if not drugged).

The stimulus properties of benzodiazepines have been studied by

Overton (1982) with animals trained to discriminate a number of

benzodiazepines from saline (e.g., chlordiazepoxide, diazepam,

flurazepam, oxazepam). When compared under similar conditions,

training progresses more quickly with barbiturates than with

benzodiazepines. Compared to several other drug classes

(stimulants, hallucinogens, antipsychotics), however, the

benzodiazepines appear more discriminable. If, however, other drugs

are substituted in chlordiazepoxide-trained animals, generalization

(i.e., increased probability of drug-appropriate responses) is

observed for all other benzodiazepines, most other sedative/

hypnotics, but not for neuroleptics. indicating at least some

specificity of effect (Colpaert et al. 1976; Barry and Krinuner

1978).

56
Although pentobarbital- and ethanol-trained animals generalize to

benzodiazepines (but not to stimulants, opioids, or hallucinogens)

animals can be trained to discriminate chlordiazepoxide from

pentobarbital (Barry and Krimmer 1978). A certain asymmetry in

cross-generalization has been noted in that generalization from a

benzodiazepine to a barbiturate has not been demonstrated as

reliably as generalization from a barbiturate to a benzodiazepine.

For example, baboons trained to discriminate pentobarbital from drug

vehicle showed generalization to lorazepam, but not all baboons

trained to discriminate lorazepam showed generalization to

pentobarbital (Ator and Griffiths 1983). These same investigators

have reported that a specific benzodiazepine receptor antagonist, Ro

15-1788, completely blocked the discriminative stimulus effects of

lorazepam but not of pentobarbital. Similarly, Herling and Shannon

(1982) showed that Ro 15-1788 blocked the discriminative stimulus

effects of diazepam but not of pentobarbital.

c. Behavioral Toxicity

Procedures for assessing the acute effects of sedative/hypnotic

compounds on psychophysical processes (i.e., auditory and visual

thresholds and reaction times) have been developed over the past two

decades. Data provided to date suggests that only studies using

primates are sensitive enough to effectively and completely evaluate

the sensory-motor effects of psychotropic drugs. It has been

reported, for example, that barbiturates impair the processing and

interpretation of sensory input (Pragay and Mirsky 1973; Bartus and

Johnson 1977). Rhesus monkeys treated with pentobarbital show a

dose-dependent decrease in correct responding to tachistoscopically

presented stimuli as a function of systematic reductions in stimulus

exposure time. More recently, Hienz et al. (1981) demonstrated that

pentobarbital produces changes in absolute visual thresholds which

may account for the apparent impairment of information processing.

Baboons housed in individual cages and maintained on a 22-hour

restricted feeding schedule were trained to press a lever and hold

it depressed for varying intervals until presentation of a light

flash or tone burst. Correct responding (defined as release of the

lever within 1.5 seconds of stimulus presentation) was rewarded with

a banana flavored food pellet. A l-second intertrial interval (ITI)

was then imposed during which no stimuli were presented and

additional lever presses reinstated the ITI. Experimental sessions

were conducted daily over a 2-3 hour period.

Auditory and visual thresholds were determined by randomly varying

the intensity of the test stimuli from trial to trial and examining

the number of correct lever releases. Using this procedure, 4 to 5

separate measures of threshold and reaction times were obtained,

each of which was based upon 140 trials. In this manner, the time

course of drug effects was also measured. Pentobarbital produced

consistent, dose-related increases in visual thresholds with a

corresponding increase in both visual and auditory reaction time

(Hienz et al. 1981). Over the same dose range, however, no effects

were observed for auditory thresholds, suggesting a specificity in

pentobarbital's psychophysical profile. In contrast,

benzodiazepines such as diazepam produced increases in both auditory

5 7
and visual thresholds which, unlike the effects of pentobarbital,

persisted for several days after injection. This procedure has also

been used to assess the effects of long-term administration of

benzodiazepines to determine whether tolerance develops to these

effects, and whether terminating extended benzodiazepine treatment

affects sensory-motor processes.

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6 1
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6 3
D. Procedures for Assessing CNS Stimulants: Anorectics, Local

Anesthetics, and Antidepressants

1. Characterization of CNS Stimulant Effects

a. Acute Drug Effects

i) Spontaneous locomotor activity

The administration of CNS stimulants in relatively low doses

generally results in an alerting response accompanied by an increase

in locomotor activity and associated behaviors (e.g.. exploration,

grooming, and rearing). This effect is dose dependent in the low

dose range. After higher doses a different spectrum of effects

emerges; stereotypic head-bobbing, gnawing, sniffing, and licking

prevail with little or no increase in locomotion.

The procedures described in the general methods section above are

most commonly used to assess the pharmacological activity of these

compounds. Although the precise mechanisms underlying these actions

may differ for various CNS stimulants, all presumably have the

common effect of increasing activity of neuronal catecholaminergic

systems. Present evidence indicates that the increased locomotor

activity is mediated by both noradrenergic and dopaminergic systems,

while stereotypy is mediated by dopaminergic systems only (Lewander

1977).

ii) Electroencephalographic (EEG) activity

Using the procedures outlined in the general methods section above,

CNS stimulants have been shown to induce characteristic changes in

the EEG. These effects have been studied using two basic

procedures: acute EEG effects, and sleep-awake profiles.

Relatively large doses of amphetamine cause cortical EEG desynchrony

in conscious cats (Bradley and Elkes 1957) correlated with an alert

and excitable behavioral state. Similar results were reported in

the dog (Schallek and Walz 1953). A variant of this acute procedure

involves the use of amphetamine to alter (e.g., reverse) the EEG

effects of CNS depressant. Using this approach, Schallek et al.

(1967) demonstrated that d-amphetamine abolished the pentobarbitalinduced peak in the 8-14 Hz band and reduced the amplitude.

The assessment of changes in the EEG sleep-awake cycle requires a

much slower recording speed and, usually, a concurrent measure of

muscle activity in order to identify the various states of alert,

resting, drowsy, slow-wave sleep, and rapid-eye-movement (REM)

sleep. Nicotine's transient alerting effects have been documented

in the cat (Domino 1979) using this approach.

iii) Behavioral performance

Numerous operant paradigms have been employed to study the effects

of CNS stimulants. These include performance maintained

appetitively on ratio and interval schedules as well as aversively

maintained shock avoidance responses. One of the general

6 4
principles which has emerged from this research is that the effect

of CNS stimulants on operant performance is dependent upon the

ongoing rate of responding. Under a wide variety of experimental

conditions, it has been demonstrated that CNS stimulants increase

low baseline response rates and decrease high baseline response

rates (Sanger and Blackman 1976). The rate dependency analysis of

the actions of CNS stimulant drugs has recently been extended to

behaviors other than schedule-controlled operants (Dews and Wenger

1977), demonstrating that this principle has wide applicability

across a variety of conditioned and unconditioned behaviors in a

variety of species.

iv) Anorexia

While numerous procedures have been employed to assess the appetitesuppressive effects of drugs, the method of Randall et al. (1960)

serves as the standard. Groups of six rats are fasted for 18 hours

but receive water ad libitum. Consumption of a weighed amount of

dry food is then measured over any desired time period (e.g., 4

hours) following injection of saline or of various doses of the test

compound. The potency of the test compound is expressed as the

anorexic dose 50% (AD50) which reduces food consumption by 50%.

Alternatively, anorexic effects can be measured by monitoring an

animal's free feeding behavior using operant techniques. With

nonfasted baboons controlling their food consumption by depressing a

lever for delivery of a 1 g pellet, the relative potency of various

amphetamine analogues has been compared on the basis of the degree

of suppression of the lever-pressing response (Griffiths et al.

1979).

v) Intracranial electrical self-stimulation

Rats are surgically implanted with electrodes in discrete brain

regions (e.g., lateral hypothalamus, medial forebrain bundle) and

then subsequently allowed to depress a lever which delivers constant

current stimulation in 0.5 second trains of a 60 Hz sine wave after

each response. Once stable intermediate response rates are achieved

by adjusting the electrical stimulation parameters, the effects of

various drugs are studied. Drugs are given via an intravenous

catheter during the experiment or via i.p., s.c., or i.m. injection

just prior to session onset. Typically, CNS stimulants such as

amphetamine increase the rate of self-stimulation, presumably by

facilitating the effects of stimulation-induced release of

norepinephrine (Ritter and Stein 1973), though recent experiments

employing discrete brain lesions cast doubt on norepinephrine's role

(Phillips et al. 1977) and suggest that dopamine may be more

important (Hollister et al. 1974).

2. CNS Stimulant Physical Dependence Potential

a. Tolerance to Drug Effects

Tolerance develops to some of the physiological and behavioral

actions of CNS stimulant drugs. For example, tolerance develops to

the anorexigenic actions of these drugs and cross-tolerance between

6 5
certain members of this class has been demonstrated (Woolverton et

al. 1978). Acute tolerance or tachyphylaxis is prevalent with

indirect-acting stimulants (e.g., those that cause the release of

endogenous catecholamines - such as amphetamine), but direct-acting

agents usually are not subject to this phenomenon. The general

methods described in previous sections apply to the assessment of

tolerance to CNS stimulants as well. Tolerance does not seem to

develop to the reinforcing effects of CNS stimulants, however, thus

playing little or no role in the assessment of the abuse liability

of CNS stimulants.

b. Withdrawal from Chronic Drug Administration

A number of investigators have reported the occurrence of withdrawal

effects when repeated administration of amphetamine is abruptly

terminated. The withdrawal profile consists primarily of

non-specific sedation or reduced motor activity (Lewander 1977).

The occurrence of such withdrawal signs has not been used as a

defining characteristic of dependence potential with CNS stimulants

primarily because of a lack of systematic and valid evaluation

procedures. Furthermore, no procedures have been developed to

classify drugs on the basis of their ability to substitute for

amphetamines and prevent the occurrence of the abstinence syndrome.

3. CNS Stimulant Abuse Liability

a. Drug Self-Administration

Procedures for assessing self-administration of CNS stimulants have

been described using rats (Pickens and Thompson 1968; Gotestam and

Andersson 1975), cats (Balster et al. 1976), dogs (Risner and Jones

1976). and primates (Deneau et al. 1969; Balster and Schuster 1973;

Griffiths et al. 1976; Johanson et al. 1976). Though the

intravenous route has been most commonly used, self-administration

has been reported to occur via the intragastric (Altshuler and

Phillips 1977), oral (Magour et al.

al. 1976) routes.

1976), and inhalation (Siegel et

i) Continuous self-administration

Procedures for continuous self-administration have been described in

detail elsewhere, in the general methods section above. Because

animals generally fail to titrate their intake of CNS stimulants

resulting in toxic overdoses (Johanson et al. 1976), continuous

self-administration techniques are generally employed within the

context of time- or dose-limited paradigms (e.g., Gotestam and

Andersson 1975; Risner and Jones 1976).

ii) Substitution

Procedures for the assessment of the abuse liability of CNS

stimulants using the substitution/limited access method have been

described in the general methods section above. Cocaine has most

often served as the baseline drug,

been used.

although amphetamine has also

Using cocaine as the baseline drug and a 3-hour timeout

66
period after each ingestion, Griffiths et al. (1976) compared the

selfiadministration of eight phenylethylamines in the baboon. A

cyclic pattern of self-administration emerged over the 15-day

substitution period in which 2-4 consecutive days of high intake

were followed by 2-3 days of low intake.

iii) Reinforcing efficacy analysis

Extensions of the limited access and substitution procedures for

evaluating relative CNS stimulant abuse liability have involved the

application of numerous procedures including progressive ratio

performance (Griffiths et al 1978; Risner and Silcox 1981), bioassay

(Risner and Jones 1975, 1980), rates of response (Balster and

Schuster 1973). and discrete-trial choice (Brady and Griffiths 1977;

Johanson and Schuster 1975). In general, the results have confirmed

the sensitivity of these methods to detect differences between the

various CNS stimulants with respect to their reinforcing efficacy,

with cocaine—revealed as one of the most potent, efficacious

reinforcers —serving as the standard for comparison.

b. Drug Discrimination

Procedures for assessing the discriminative stimulus properties of

CNS stimulants have been established in a number of animal species

(Schuster and Balster 1977; La1 1977). Rats learned to discriminate

between injection of d,l-amphetamine and saline, and in addition,

generalization was obtained with methylphenidate but not with

atropine (Harris and Balster 1971). In a subsequent study,

d-amphetamine-trained rats generalized to l-amphetamine, but not to

nicotine, mescaline, fenfluramine, or LSD (Schechter and Rosecrans

1973). Additional evidence for the specificity of these procedures

has been provided by Overton and collegues (Overton 1966; Overton

and Lebman 1973), who demonstrated that animals trained to respond to

sedative/hypnotics responded to amphetamine as if it were saline

(i.e., no cross-generalization between drug classes was observed).

Similarly, generalization between d-amphetamine and cocaine has been

demonstrated in the rhesus monkey (Ando and Yanagita 1978).

c. Behavioral toxicity

Procedures for assessing the behavioral toxicity of CNS stimulants

have been refined for use in squirrel and rhesus monkeys, baboons,

and pigeons. Methamphetamine impairs pigeon's performance in visual

discrimination tasks (Dews 1955), while d-amphetamine decreases the

accuracy of visual stimulus duration discriminations in this same

species (Stubbs and Thomas 1974). In both rhesus monkeys (Brown and

Bass 1967) and baboons (Hienz and Brady 1981) d-amphetamine

decreases reaction times to visual and auditory stimuli. This

effect is dose-dependent, however, with increases in reaction time

predominating after higher doses. The decreased reaction times

following d-amphetamine occur in the absence of changes in size

discrimination thresholds (Brown and Bass 1967) or brightness

discrimination (Thurmond 1965) in the monkey. In the squirrel

monkey, however, d-amphetamine and methylphenidate produce modest

increases in masked auditory thresholds (Delay et al. 1979).

6 7
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70
E. Procedures for Assessing Cannabinoids

1. Characterization of Cannabinoid Effects

a. Acute Drug Effects

i) Spontaneous locomotor activity

In general, administration of cannabinoids results in decreased

motor activity over a wide dosage range. Most studies have been

conducted in rats and have used the interruption of photoelectric

beams (Kubena and Barry 1970; Brown 1972). the time to leave a

platform (Cohn et al. 1972), exploration of a shuttle box

(Hendrikson and Jarbe 1971), or merely gross observation (Phillips

et al. 1972) as measures of activity. Mice (Holtzman et al. 1969),

gerbils (Grunfeld and Edery 1969), chickens (Abel et al. 1972). and

dogs (Grunfeld and Edery 1969) have also been the subjects of

studies on the motor effects of cannabinoids.

Davis et al. (1972) demonstrated a biphasic effect of

delta-9-THC in rats over a ten-day period of dosing. Both

excitation (on first two days) and depression (third through the

tenth days) were observed using the photoelectric method of

monitoring locomotor activity. In a study by Hardman et al. (1971)

using dogs and monkeys as subjects, a number of THC analogues were

studied and two were found to decrease activity at low doses (e.g.,

0.1-0.2 mg/kg, i.m.). But high doses induced a biphasic response

characterized by initial CNS stimulation followed by prolonged

depression.

al. (1968).

This finding was confirmed for monkeys by Scheckel et

ii) Analgesic activity

Several laboratories have studied the antinociceptive effects of

cannabinoids using a number of different methods and in a variety of

species. Many pre-1974 studies used rats or mice in the tail-flick

or hot-plate procedures to demonstrate the analgesic activity of

delta-9-THC (Buxbaum 1972; Gallager et al. 1972; Bicher and

Mechoulam 1968). but Harris (1971) and Domino et al. (1971) obtained

inconsistent results. Hill et al. (1974) demonstrated that the

doses required for analgesia in these methods cause decrements in

motor activity. Thus, a retardation in the animal's physical

capacity to respond may partially explain the inconsistent finding

in earlier studies. Other studies using electrical tooth pulp

stimulation with dogs (Kaymakcalan et al. 1974) and shock titration

(Scheckel et al. 1968) have shown that delta-9-THC possesses

analgesic activity. Wilson and May (1975) have shown, however, that

the analgesic activity of delta-&and delta-9-THC is mainly due to

the 11-hydroxy metabolite.

iii) "Hallucinatory" behavior

While no systematic objective procedures have been developed to

assess whether animals experience hallucinations following drug

injection, numerous aberrant behaviors have been observed in higher

71
mammals. For example, Joachimoglu (1965) observed dogs that had

received hashish to be trying to "grab an apparition in the air" and

to direct their eyes toward nonexistent objects in the air. Similar

accounts of abnormal behaviors have been reported to occur in

monkeys (Scheckel et al. 1968), with emphasis on apparent visual

"hallucinations."

iv) Dog ataxia test

Delta-9-THC and other cannabinoids with psychoactive effects in man

have particularly unusual effects on the overt behavior of dogs. A

rating scale for the effects has been developed to allow

quantitation (Dewey et al. 1972; Martin et al. 1975). Test drugs

are given i.v. At low doses a slight reduction in activity can be

seen, but at modest doses a distinctive "prance-like" walk with

exaggerated responses to stimulation is produced. At higher doses,

behaviorally active cannabinoids produce a static ataxia. The dogs

remain standing in one place but sway from side to side. The test

can be used to screen for THC-like activity as well as to estimate

potency differences among cannabinoids.

v) Schedule-controlled behavior

A recent review of the THC effects upon schedule-controlled behavior

has been provided by McMillan (1977). The use of such operant

techniques has been most effective in elucidating numerous

characteristics of the THC pharmacologic profile including their

effectiveness via various routes of administration, the time course

and degree of tolerance development, the long-term or irreversible

cumulative effects after cessation of exposure, and the cumulative

effects after repeated administration (Ferraro 1976).

vi) Electroencephalographic (EEG) activity

As described in the general methods section, two basic procedures

are employed to assess the EEG effects of the cannabinoids: acute

changes in the frequency and voltage characteristics, and

alterations in the sleep-awake cycle. In the former, the data are

quantified using visual inspection, voltage integration, frequency

analysis, or power spectral analysis. Animals prepared with chronic

cerebrocortical or depth electrodes are given acute injections of

either the active constituents of marijuana (e.g., delta-9-THC) or

crude marijuana extract. The resulting EEG effects are then

characterized with respect to the voltage content or predominant

frequency of various bands such as delta (O-4 Hz), theta (4-8 Hz),

alpha (8-13 Hz), or beta (13-25 Hz). Measurements are obtained at

specific time points after injection (e.g., 30 minutes, 1 hour, 2

hours, etc.). Power spectral analysis has been utilizied to more

precisely quantify the voltage and frequency components of the EEG

of rats given delta-9-THC (Buonamici et al. 1982). The EEG voltage

was reduced by 50% of control 1 hour after i.p. administration of 10

mg/kg of delta-9-THC. Return to control levels occurred over the

course of 8 hours. During this period prominent EEG bursts of 6 Hz

appeared to override the continuous EEG training. Similar findings

72
were reported for delta-9-THC administration in the rabbit

(Willinsky et al. 1975).

Characterization of the effects of cannabinoids on sleep-awake

activity has been studied in the rat (Masur and Khazan 1970; Moreton

and Davis 1973). Changes in the distribution of the various phases

of sleep (as measured using the method of van Twyver 1969) have

suggested that time spent in slow-wave sleep is decreased and

wakefulness is increased (Moreton and Davis 1973). In addition,

this study compared REM-sleep-deprived with non-REM-sleep-deprived

rats and found that they differed in the degree of dissociation of

REM sleep phasic and tonic events, and with respect to the profile

of REM sleep rebound.

Both of these techniques provide excellent baselines upon which to

study tolerance development. Rats (Moreton and Davis 1973; Pirch et

al. 1973), pigeons (McMillan et al. 1972), dogs (Dewey et al. 1972),

and monkeys (Stadnicki et al. 1974) all show tolerance to

delta-9-THC or crude marijuana. Two reports, however, have failed

to find EEG tolerance to delta-9-THC in the cat (Lipparini et al.

1969; Barratt and Adams 1973).

2. Cannabinoid Physical Dependence Potential

a. Tolerance to Drug Effects

Tolerance to the analgesic effects of cannabis has been reported to

last up to 64 days after a single injection in the rat (Kaymakcalan

1973). Similar persistence of tolerance to the behavioral

depressant effects of delta-9-THC has been reported in the dog

(Dewey et al. 1972). This prolonged duration of tolerance coupled

with the fact that tolerance to the sedative and ataxic effects

develop slowly relative to other effects suggests that results

obtained with procedures which depend upon locomotor activity to

assess cannabinoid effects must be interpreted with extreme caution.

Such procedures should incorporate 1) controls for ataxia and/or

sedation, 2) standardized dosing and testing schedules, 3) multiple

test doses, and 4) appropriately long "wash-out" periods before

subsequent dosing. Wikler (1976) has reviewed the literature in

this area.

b. Withdrawal from Chronic Drug Administration

Procedures for assessing the abstinence syndrome in chronic

cannabinoid-treated subjects have not been as well standardized as

those for assessing opioids and CNS depressants. Such studies using

rats or mice have reported definite abstinence signs in some cases

(Cutler et al. 1975; Karler and Turkanis 1976), while others using

convulsive thresholds as an indicator (Chesher and Jackson 1974;

Leite and Carlini 1974) failed to demonstrate withdrawal. The most

conclusive evidence for a dependence potential of cannabinoids has

been provided by numerous studies with monkeys, using intravenous

(Kaymakcalan 1972), oral (Stadnicki et al. 1974; Snyder et al.

1975)) and inhalation (Heath 1976) routes of administration to

maximize exposure to the delta-9-THC. Behavioral signs of

7 3
withdrawal from chronic delta-9-THC administration appear after

about 12 hours and continue for about 5 days. They include yawning,

anorexia, piloerection, hyperirritability, aggressiveness,

scratching, biting and licking fingers, pulling hair, tremors,

twitches, penile erection and masturbation, eating feces and

staring, and grasping at invisible objects (Kaymakcalan 1973).

Similar increased aggressiveness was observed by Stadnicki et al.

(1974) with the addition of EEG desynchrony and "hallucinations."

Snyder et al. (1975) reported a disruption of operant performance

for liquid reinforcement upon termination of delta-9-THC.

Precipitated withdrawal procedures have involved two different

strategies. Since the spontaneous withdrawal signs from delta-9-THC

resemble opioid withdrawal signs (Kaymakcalan 1973) and delta-9-THC

alleviates morphine withdrawal signs (Bhargava 1978), naloxone has

been used to precipitate withdrawal in delta-9-THC-maintained rats

(Hirschhorn and Rosecrans 1974; Kaymakcalan et al. 1977). Secondly,

observations suggesting that serotonergic mechanisms may be involved

in the action of delta-9-THC (Sofia et al. 1971) prompted Taylor and

Fennessy (1978) to administer the potent serotonin uptake inhibitor,

clomipramine, in order to precipitate withdrawal signs in

delta-9-THC-treated rats. Withdrawal signs were similar to those

reported to occur in spontaneously withdrawn animals, though Taylor

and Fennessy (1978) failed to observe spontaneous abstinence signs

when delta-9-THC was discontinued.

3. Cannabinoid Abuse Liability

a. Drug Self-Administration

Procedures for assessing self-administration of cannabinoids have

been described using rats (van Ree et al. 1978; Takahashi and Singer

1979) and rhesus monkeys (Kaymakcalan 1972; Pickens et al. 1973;

Harris et al. 1974). Both intravenous and inhalation (Pickens et

al. 1973) routes have been used.

i) Continuous self-administration

Studies using this procedure have failed to show that delta-9-THC

reliably maintains self-injection performance in drug-naive animals,

though subjects previously administered the compound passively do

self-inject (Kaymakcalan 1972; Harris et al. 1974; van Ree et al.

1978). Takahashi and Singer (1979) have also reported that fooddeprived rats self-administered more delta-9-THC than non-fooddeprived rats. Regardless of the procedure used, however, animals

self-administering delta-9-THC show lower response rates than those

reported for opioid, CNS depressant, or stimulant selfadministration.

ii) Substitution

Using cocaine (Kaymakcalan 1972) or phencyclidine (Pickens et al.

1973) as baseline drugs, substituted THC has been reported to

maintain self-injection behavior; however, other substitution

studies with delta-9-THC and other behaviorally active cannabinoids

74
fail to find evidence for reinforcing properties (Harris et al.

1974; Carney et al. 1977; Young et al. 1981). No studies have as

yet been conducted to directly compare the relative reinforcing

efficacy of delta-9-THC and related cannabinoids.

b. Drug Discrimination

Procedures for assessing the discriminative stimulus properties of

cannabinoids have been reported with rats, gerbils, pigeons, and

rhesus monkeys. Using "T" maze performance in rats, it has been

demonstrated that delta-9-THC, delta-8-THC, and hashish' were all

capable of acting as discriminative stimuli resulting in better than

75% correct responses by the 10th trial (Jarbe and Henriksson 1974).

Generalization gradients for various doses of delta-9-THC have been

shown to correlate very well using a number of procedures including

rat two-lever discrimination (Balster and Ford 1978)) T-maze (Jarbe

and Henricksson 1974)) conflict (Barry and Kubena 1972). and

shock-escape maze (Jarbe et al. 1976).

An extension of these procedures provides information relating to

the degree of cross generalization from delta-9-THC to other

cannabinoids. Barry and Krimmer (1975) showed that crude marijuana

extract generalized to delta-9-THC in a two-lever discrimination

task in rats. Further, it has been shown that numerous metabolites

of delta-9-THC generalize to the parent compound. The 11-hydroxy

delta-8- and delta-9-THC compounds both generalize to delta-9-THC at

relatively low doses (Balster and Ford 1978). Cannabinol and

cannabidiol did not generalize to delta-9-THC (Barry and Kubena

1972; Jarbe and Henricksson 1974).

Finally, the discriminative stimulus properties of delta-9-THC are

fairly specific in that many psychoactive compounds do not

generalize to it. Pentobarbital, alcohol, chlordiazepoxide,

chlorpromazine, morphine, methamphetamine, cocaine, atropine, LSD,

mescaline, and phencylidine are among those' tested that fail to

generalize to delta-9-THC (Barry and Kubena 1972; Jarbe and

Henricksson 1974; Jarbe et al. 1975; Barry and Krimmer 1975.

c. Behavioral Toxicity

Procedures for assessing the effects of cannabinoids on sensorymotor performance have encompassed a number of specific techniques

and animal species. Elsmore (1972) demonstrated that delta-9-THC

impairs auditory discrimination in the monkey, while Ferraro and

Grilly (1973) reported that delta-9-THC produced dose-related

changes in response speed on visual matching-to-sample performance

in the chimpanzee. The acute and chronic effects of marijuana on

complex operant performance have been quantified in the squirrel

monkey (Adams and Barratt 1974); In addition, delta-9-THC appears

todisrupt learning differentially, depending on the paradigm used in

rodents (Robichaud et al. 1973). This finding has been expanding to

include both cannabis extract and delta-9-THC. Stiglick and Kalant

(1982, 1983) showed that when these compounds are given chronically

by gavage to rats, impairment of learning on radial-arm maze, DRL

food-reinforced performance, and habituation in an open field

75
develops and persists for long periods of time after drug

administration has ceased. While a number of studies have

characterized the behavioral toxicologic profile of delta-9-THC, no

studies to date have attempted to compare these effects with those

produced by other cannabinoids.

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Pickens, R.; Thompson, T.; and Muchow; D. Cannabis and

phencyclidine self-administration by animals. In: Goldfarb,

L., and Hoffmeister, F., eds. Psychic Dependence. Berlin:

Springer-Verlag, 1973. pp. 78-86.

Pirch, J.H.; Osterholm, K.C.; Cohn, R.A.; and Barratt, E.S. Studies

on EEG tolerance to marijuana in the rat. Arch Int

Pharmacodyn, 203:213-220, 1973.

Robichaud, R.C.; Hefner, M.A.; Anderson, J.E.; and Goldberg, M.E.

Effects of delta-9-tetrahydrocannabinol (THC) on several rodent

learning paradigms. Pharmacology, 10:1-11, 1973

Scheckel, C.L.; Boff, E.; Dahlen, P.; and Smart, T. Behavioral

effects in monkeys of racemates of two bilogically active

marijuana constituents. Science, 160:1467-1469, 1968.

Snyder, E.W.; Lewis, E.G.; Dustman, R.E.; and Beck, E.C. Sustained

ingestion of delta-9-tetrahydrocannabinol and the operant

behavior of stump-tailed macaques. Pharmacol Biochem Behav, 3:

1129-1132, 1975.

Sofia, R.D.; Dixit, B.N.; and Barry, H. The effect of delta-ltetrahydrocannabinol on serotonin metabolism in the rat brain.

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Stadnicki, S.W.; Schaeppi, U.; Rosenkrantz, H.; and Braude, M.C.

Crude marihuana extract: EEG and behavioral effects of chronic

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Stiglick, A., and Kalant, H. Residual effects of prolonged cannabis

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7 9
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8 0
F. Procedures for Assessing Hallucinogens and Anticholinergics

1. Characterization of Hallucinogen and Anticholinergic

Effects

a. Acute Drug Effects

i) Abnormal behaviors

Numerous animal species including mice, rabbits, and cats exhibit

characteristic abnormal postures or behaviors after receiving acute

injections of lysergide-like hallucinogens. Head twitching in mice

and hyperthermia in rabbits have been noted, but these observations

have not evolved into' a standardized testing procedure. The cat

limb-flick model of Jacobs et al. (1977) has received attention as a

more refined procedure for assessing hallucinogenic activity.

Moreover, this abnormal behavior has been shown to be associated

with a decreased firing of serotonin-sensitive neurons in the raphe

nucleus. The success of methods for recording single unit activity

in freely moving animals has been responsible for obtaining these

electrophysiological correlates of behaviors. Cats are given an

acute injection of the test compound and then observed in an openfield environment. The limb-flick is seen in control, non-treated

cats only in response to the presence of a foreign substance on the

paw. In LSD-treated cats, the paw is raised from the ground and

then rapidly shaken or flicked away from the body. The number of

limb-flicks per unit time is then tabulated and used as an index of

a test drug's LSD-like activity.

Another approach to studying psychotomimetics has focused on the

complex relationships within social colonies of non-human primates.

On a test day, only one of 5-10 monkeys is treated and the entire

colony is observed for abnormal or "emergent" behaviors. Each

animal is treated on separate days, and the direct effects on the

treated animal are tabulated,as is the behavior of the untreated

monkeys. Behavioral checklists are used by "blind" observers;

Heinze et al. (1980) utilized this procedure to study the effects of

5-methoxytryptamine in Stumptail macaques. Schlemmer and Davis

(1983) have reviewed the literature comparing various models of

psychotomimetic-induced behaviors.

ii) Spinal dog preparation

This technique (which is described in detail in the opioid section)

has been used to characterize compounds with respect to their LSDlike activity (Nozaki et al. 1977). In general, LSD and related

compounds produce a spectrum of effects including whining, eye

tracking movements, increased respiratory rate, hyperthermia,

increased latency to skin twitch, and facilitation of the flexor

reflex. Thus, it is the entire profile of effects in this procedure

that is utilized to determine a test compound's LSD-like activity.

An extension of this technique involves the use of specific

pharmacologic antagonists (e.g., cyproheptadine, phenoxybenzamine,

chlorpromazine) in order to further differentiate these compounds

with respect to their LSD- vs. amphetamine-like profiles.

8 1
iii) Schedule-controlled behavior

The basic procedures for assessing the effects of hallucinogens on

operant performance are similar to those described under general

methods above. LSD and congeners differentially affect positive

reinforced behavior depending upon the prevailing response rate and

schedule (Appel 1968; Dews and Wenger 1977). These techniques have

been most useful in providing information on relative potencies of

hallucinogens (Appel and Freedman 1965), demonstrating that -LSD is

about 10 times more potent than both psilocybin and d-amphetamine

and 100 times more potent than mescaline. The available data

indicate that the use of aversive control procedures does not

provide information that can be interpreted as unique to this class

of compounds (Appel et al. 1967).

iv) Electroencephalographic (EEG) activity

Procedures for assessing the EEG effects of LSD and related

compounds have been developed for the mouse (Morley and Bradley

1977)) rat (Depoortere and Loew 1971), rabbit (Long 1962; Khazan

and McCash 1965), cat (Killam and Killam 1956)) and monkey (Walter

et al. 1971). Most of these studies have focused on acute effects

described in terms of bursts of spikes, spindling. and

desynchronization. Increased wakefulness and delayed onset and

decreased duration of REM sleep have also been reported (Depoortere

and Loew 1971).

Attempts to quantify the EEG effects of hallucinogens with respect

to frequency and voltage components have been successful in both

mice and cats. Mescaline, dimethoxy-amphetamine (DMA), and related

ring-substituted phenylisopropylamines produce hypersynchronous

slow-wave activity associated with abnormal stereotypic behaviors in

the cat (Fairchild et al. 1967). These effects contrast with

amphetamine which produces desynchrony concomitant with behavioral

alerting and hyperactivity. These data provide a framework within

which relative potencies of test compounds for hallucinogenic

activity can be determined. Winters and Wallach (1970) demonstrated

that cats treated with hallucinogens exhibit hypersynchronous EEG

peaks in the 2.5-3.0 Hz range. Morley and Bradley (1977) have used

power spectral analysis to quantify the EEG effects of N,Ndimethyltryptamine in mice. A dose-dependent hypersynchrony at

2.5-4.5 Hz was observed to persist for up to 60 minutes after

injection. In addition, anticholinergics produce slow wave

hypersynchrony associated with alteration in behavior consistent

with models of hallucinosis.

v) Binding assays

Stereospecific 3H-LSD high affinity binding was demonstrated in the

mid 1970s (Bennett and Aghajanian 1974; Bennett and Snyder 1975) and

has since provided a basis for associating serotonin and LSD with

specific neuronal structures. The physiologic significance of

binding potencies has not been identified because of the poor

correlation of lC50 values and hallucinogenic activity (Freedman and

Boggan 1982). Greater success has been achieved when correlations

82
are made between changes in binding following in vivo treatment with

compounds that produce known functional changes in receptors (e.g.,

5,7-dihydroxytryptamine). Increases in LSD binding in specific

brain areas have been demonstrated in reserpine or parachloroamphetamine-treated rats (Bennett and Snyder 1976).

2. Hallucinogen and Anticholinergic Physical Dependence

Potential

a. Tolerance to Drug Effects

Tolerance appears to develop fairly rapidly to many of the effects

of LSD and related compounds. Most notable have been studies that

employ behavioral procedures (Freedman et al. 1958; Appel and

Freedman 1968). EEG activity (Wallach et al. 1972). and measures of

cardiovascular reflexes in the spinal dog preparation (Martin and

Eades 1972). Because the degree of tolerance developed to almost

all hallucinogens is so complete and occurs so rapidly, these

procedures are not particularly useful in quantifying the dependence

potential per se. Because marked cross-tolerance between compounds

of similar chemical structure is widespread (Martin and Eades 1972;

Vaupel and Nozaki 1975), however, screening for LSD-like activity

using this procedure may be useful.

b. Withdrawal from Chronic Drug Effects

To date no studies have demonstrated that the termination of chronic

administration of hallucinogens results in an abstinence syndrome

(McIsaac et al. 1970).

3. Hallucinogen and Anticholinergic Abuse Liability

a. Drug Self-Administration

i) Continuous self-administration

Deneau et al. (1969) demonstrated that rhesus monkeys would not

initiate self-administration of mescaline either spontaneously or

after 1 month of programmed administration. LSD may, in fact,

possess aversive properties, since Hoffmeister (1975) has

demonstrated that rhesus monkeys will respond in order to terminate

stimuli associated with the delivery of this compound.

ii) Substitution

Using cocaine as the baseline drug, Griffiths et al. (1979)

demonstrated that four hallucinogenic amphetamine-like compounds

(DOET, DOM, mescaline, and PMA) do not maintain self-administration

in baboons.

b. Drug Discrimination

Procedures for assessing discriminative stimulus properties are

perhaps the most well developed of those currently available for

assessing compounds for hallucinogenic activity. The discriminative

83
stimulus properties of LSD, mescaline, and related compounds are

apparent after relatively small doses (Cameron and Appel 1973;

Hirschhorn and Winter 1971). In addition, the number of trials to

criterion is relatively small, suggesting that these compounds

generate strong interoceptive stimulation. Appel et al. (1978) have

investigated the conditions under which such performances occur and

report generalization gradients for doses other than the training

dose as well as for other stimuli (e.g., tones, lights, etc.).

There is good generalization or cross-transfer between the

discriminative effects of LSD, mescaline, psilocybin, and quipazine

(Appel et al. 1978). These procedures thus provide useful

information for classifying these and other compounds with respect

to pharmacologic class (Kuhn et al. 1977).

c. Behavioral Toxicity

Procedures for assessing the degree of behavioral toxicity of the

hallucinogens have been, for the most part, directed at quantifying

the sensory-motor performance of animals. Blough (1956, 1957)

demonstrated that LSD could improve pigeon's accuracy on a complex

visual discrimination task while elevating the absolute visual

threshold. Species differences are apparent, since LSD impairs

auditory frequency discrimination in the cat (Key 1961) but not the

rat (Dykstra and Appel 1974). Auditory generalization gradients

were unaffected by LSD in pigeons (Dykstra and Appel 1972).

Finally, Brown and Bass (1967) showed that LSD increases the visual

reaction time in rhesus monkeys while increased visual thresholds in

this species were reported by Ando and Takada (1979).

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I 29

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Psychopharmacology, 33:117-134, 1973.

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

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Depoortere, H., and Loew, D.M. Alterations in sleep/wakefulness

cycle in rats following treatment with (+)-lysergic acid

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Dews, P.B., and Wenger, G.R. Rate-dependency of the behavioral

effects of amphetamine. In: Thompson, T.,. and Dews, P.B.,

eds. Advances in Behavioral Pharmacology. Vol. 1. New York:

Academic Press, 1977,. pp. 167-227.

Dykstra, L.A., and Appel, J.B. LSD and stimulus generalization:

Rate-dependent effects. Science, 177:720-722, 1972.

Dykstra. L.A.; and Appel, J.B. Effects of LSD on auditory

perception: A signal detection analysis. Psychopharmacologia,

34:289-307. 1974.

Fairchild, M.D.'; Alles, G.A.; Jenden, D.J.; and Mickey, M.R. The

effects of mescaline, amphetamine, and four-ring substituted

amphetamine derivatives on spontaneous brain electrical

activity in the cat. Int J Neuropharmacol, 6:151-167, 1967.

Freedman, D.X.; Aghajanian, G.K.; Ornitz; E.M.; and Rosner, B.S.

Patterns of tolerance to lysergic acid diethylamide and

mescaline in rats. Science, 127:1173-1174, 1958.

Freedman, D.X., and Boggan. W.O. Biochemical pharmacology of

Dsychotomimetics. In: Hoffmeister. F.. and Stille. G., eds.

Handbook of Experimental Pharmacology. Vol 55/III. Berlin:

Springer-Verlag. 1982. pp. 57-88.

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.

Heinze, W.J.; Schlemmer, R.F.; Williams, E.A.; and Davis, J.M. The

acute and chronic effect of 5-methoxytryptamine on selected

members of a primate social colony. Biol Psychiat, 15:829-839,

1980.

8 5
Hirschhorn, I.D., and Winter, J.C. Mescaline and lysergic acid

diethylamide (LSD as discriminative stimuli.

Psychopharmacology, 22:64-71, 1971.

Hoffmeister, F. Negatively reinforcing properties of some

psychotropic drugs in drug-naive rhesus monkeys. J Pharmacol

Exp Ther, 192:468-477, 1975.

Jacobs, B.L.; Trulson, M.E.; and Stern, W.C. Behavioral effects of

LSD in the cat: Proposal of an animal behavior model for

studying the actions of hallucinogenic drugs. Brain Res, 132:

301, 1977.

Key, B.J. The effect of drugs on discrimination and sensory

generalization of auditory stimuli in cats.

Psychopharmacologia, 2:352-363, 1961.

Khazan, N., and McCash, D. Effects of LSD-25, N,N-dimethyltryptamine (DMT), and N,N-diethyltryptamine (DET) on the photic

evoked responses in the unanesthetized rabbit. Arch Int

Pharmacodyn Ther, 154:474-483, 1965.

Killam, K.F., and Killam, E.K. The action of lysergic acid

diethylamide on central afferent and limbic pathways in the

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Kuhn, D.M.; White, F.J.; and Appel, J.B. Discriminative stimulus

properties of hallucinogens: Behavioral assay of drug action.

In: Lal, H., ed. Discriminative Stimulus Properties of Drugs.

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Longo, V.G. Epinephrine and drugs acting on adrenergic receptors;

5-hydroxytryptamine and related compounds. In: Longo. V.G.,

ed. Rabbit Brain Research. Vol. 2. New York: Elsevier,

1962.

Martin, W.R., and Eades, C.G. Cross tolerance to tryptamine in the

LSD tolerant dog. Psychopharmacologia, 27:93-98, 1972.

McIsaac, W.M.; Harris, R.T.; and Ho, B.T. The indole hallucinogens.

In: Harris, R.T., McIsaac, W.M., and Schuster, C.R., eds.

Drug Dependence. Austin, Texas: Capital, 1970. pp. 41-54.

Morley, B.J., and Bradley, R.J. Spectral analysis of mouse EEG

after the administration of N,N-dimethyltryptamine. Biol

Psychiat, 12:757-769, 1977.

Nozaki, M.; Vaupel, D.B.; and Martin, W.R. A pharmacologic

comparison of 3,4-methylenedioxyamphetamine and LSD in the

chronic spinal dog. Eur J Pharmacol, 46:339-349, 1977.

Schlemmer, R.F., and Davis, J.M. A comparison of three

psychotomimetic-induced models of Dsychosis in non-human social

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Vaupel, D.B., and Nozaki, M. A comparison of the effects of

2,5-dimethoxyamphetamine (DMA) and lysergic acid diethylamide

(LSD) in the dog. Fed Proc, 34:767, 1975.

Wallach, M.B.; Friedman, E.; and Gershon, S. 2,5-Dimethoxy-4-

methylamphetamine (DOM), a neuropharmacologial examination. J

Pharmacol Exp Ther, 182:145-154, 1972.

Walter, S.; Balzano, E.; Vuillon-Cacciuttolo, G.; and Naquet, R.

Effects comportementaux et electrographiques du diethylamide de

l'acide d-lysergique (LSD25) sur le Papio papio photosensible.

Electroenceph Clin Neurophysiol, 30:294-305, 1971.

8 6
Winters, W.D., and Wallach, M.B. Drug induced states of CNS

excitation: A theory of hallucinosis. In: Efron, D.H., ed.

Psychotomimetic Drugs. New York:

193-214.

Raven Press, 1970. pp.

8 7
G. Procedures for Assessing Dissociative Anesthetics

1. Characterization of Dissociative Anesthetic Effects

a. Acute Drug Effects

i) Spontaneous locomotor activity

Using an actophotometer, Chen (1959) demonstrated that the

predominant response of rodents to PCP was that of excitant

hyperactivity. This effect, however, was dose and species specific

in that higher doses induced stereotypic and depressed behavior, and

primates were only sedated after PCP administration.

ii) Forced motor activity

Using the rotorod assay described under general methods above, Kalir

et al. (1978) and Vincent et al. (1979) have demonstrated that PCP

and some of its analogues disrupt forced motor activity in a

dose-related manner. In addition, the relative potency for this

effect has been found to be positively correlated with the relative

potency for binding to a neuronal receptor.

iii) Electroencephalographic (EEG) activity

The EEG profile of PCP was first characterized in animals by Domino

(1964). In a comparative study, Gehrmann and Killam (1976)

demonstrated that of all the CNS depressants tested, PCP and

ketamine produced the greatest increase in EEG voltage in all

frequencies of the primate. More recently, Mattia and Moreton

(1981, 1982) have also characterized the EEG profile of PCP and

ketamine using power spectral analysis. Lower doses of PCP produced

increased voltage in the theta band while larger doses produced two

distinct peaks of activity (one at O-5 Hz and the other at 7-8 Hz).

Ketamine demonstrated a similar profile, but was about 10 times less

potent that PCP.

iv) Behavioral performance

Because of the complex nature of PCP's pharmacology, multiple

schedules seem to be the most appropriate paradigms to assess the

effects of PCP on operant performance. This is especially true

since the finding that the effects of psychoactive agents on operant

performance are related to the schedule employed (Dews and De Weese

1977; McKearney and Barrett 1978). In this regard, rodent (Murray

1978; Woolverton and Balster 1979) and primate (Chait and Balster

1978; Brady et al. 1980) data have shown that low doses of PCP

increase low rates of responding (e.g., fixed interval) and decrease

high rates of responding (e.g., fixed ratio). Three analogues, PCE,

TCP, and ketamine, have been shown to produce qualitatively similar

effects on operant performance; the only differences observed were

in the relative potencies of these compounds.

88
v) Binding assays

Using techniques similar to those used to demonstrate the existence

of opiate receptors, a component in brain membrane has been shown to

3 bind [ H] PCP with a high degree of specificity and with a fairly

high affinity (Vincent et al. 1979; Zukin and Zukin 1979). In

addition, PCP and 3 of its analogues have been shown to displace

C

3

H]quinuclidinyl benzylate (muscarinic receptor) and

[

3

H]dihydromorphine (opiate receptor) with a respectable degree of

specificity (Vincent et al. 1979; Su et al. 1980). The "PCP

receptor" appears to be most concentrated in the synaptic, fraction

of cerebral cortex and corpus striatum, while significant, but

somewhat lower, degrees of binding were found in hippocampus and

thalamus.

While there has been some early controversy regarding the specific

binding of PCP to glass wool fibers, the more recent studies have

demonstrated significant correlations between the relative potency

of PCP and its analogues in the rotorod test and binding affinities

(Vincent et al. 1979; Zukin and Zukin 1979). The ultimate impact of

the presence of PCP receptor may reside not in the advancement of

our understanding of PCP's mechanism of action, but in its possible

usefulness in rapid screening of drugs for PCP-like activity and in

the search for a PCP antagonist which would provide a valuable addition to the current strategy of treating PCP intoxication.

2. Dissociative Anesthetic Physical Dependence Potential

a. Tolerance to Drug Effects

Repeated administration of PCP to monkeys and rats results in a

decreased disruption (2-4-fold change) of operant performance

(Balster and Chait 1976; Chait and Balster 1978; Woolverton and

Balster 1979; Woolverton et al. 1980). Various behavioral paradigms

were used including a chain schedule procedure for food

reinforcement and a fixed interval schedule procedure for milk.

Similar degrees of tolerance were observed regardless of whether PCP

was administered before or after the test session. First dose

behavioral tolerance of PCP has also been reported to occur in the

rat (Ruffing and Domino 1980). This phenomenon was found to be

dose related; however, the duration of complete suppression of

lever-pressing activity was used as the dependent variable and not

response rate as reported in the earlier studies. Regardless of the

technique employed, it appears that marked tolerance develops to at

least some of the effects of PCP.

b. Withdrawal from Chronic Drug Administration

Using a continuous access self-administration paradigm, rhesus

monkeys were observed to readily initiate self-injections of PCP

(0.1 mg/kg/inj) and increase their drug intake over the course of 20

days (Balster and Woolverton 1980). Daily intakes of up to 7.4

mg/kg were achieved, and previously depressed food intake had

returned to baseline (indicative of tolerance development). The

animals maintained a state of continuous intoxication which was

89
sustained when the dose was increased to 0.5 mg/kg. Substitution of

saline after 58 days of exposure resulted in the appearance of

numerous abstinence signs and symptoms including increased

vocalizations, bruxism, oculomotor hyperactivity, diarrhea, refusal

of preferred food, piloerection, and tremors. Less common signs

included ear and facial twitches, priapism, abdominal contractions,

emesis, and convulsions. The time course of withdrawal was

characterized by an initial recovery from , the PCP-induced

intoxication at about 4 hours after saline substitution. Onset of

hyperresponsive behaviors became evident at 8-12 hours, with the

maximum number of symptoms occurring 12-15 hours post-substitution.

The syndrome dissipated over 24 hours, however, and all withdrawal

signs were immediately reversed by PCP (0.25 g/kg, i.v.). This

study also showed that PCP blood levels were in the 105-280 mg/ml

range during self-administration, and declined to O-12 mg/ml with

saline substitution.

3. Dissociative Anesthetic Abuse Liability

a. Drug Self-Administration

The rhesus monkey has been shown to self-administer PCP

intravenously (Pickens et al. 1973; Balster et al. 1973; Balster and

Woolverton 1980) and orally (Carroll and Meisch 1980; Carroll 1982).

In the primate studies, not only did PCP maintain self-injection

performance when substituted for cocaine, but animals with no prior

history readily initiated self-injections of PCP (Balster et al.

1973). The self-administration profile of PCP was (like many other

drugs of abuse) characterized by an inverted U dose-response curve

and while the number of self-injections decreased as the dose was

increased, the total intake per day increased. When the fixed ratio

for delivery of an injection was increased from 1 to 5, the number

of self-injections also decreased.

Finally, self-administration studies with PCP analogues such as

ketamine (Moreton et al. 1977) and numerous N-substituted analogues

(Risner 1982) have shown that all of these compounds maintain

self-injection performance similar to that of PCP. The only

differences observed thus far were those of relative potency. Using

a bioassay procedure that predicts relative potencies, parallelism

of curves, and relative maximal effects, Risner (1982) demonstrated

that eight structurally related analogues of PCP were

self-administered by the dog. The thienyl-substituted analogue was

the most potent, followed by PCP, y-substituted alkyl analogues,

monohydroxylated metabolites, and ketamine.

b. Drug Discrimination

Procedures for assessing the discriminative stimulus properties of

phencyclidine and related compounds have been developed and refined

for both the rat and the squirrel monkey. Overton (1975) initially

demonstrated that PCP and ketamine shared similar discriminable

effects which were dissociable from the stimulus effects of

pentobarbital. Similar stimulus generalization curves were observed

for PCP, cyclohexamine, and ketamine, while ditran did not

9 0
generalize to the PCP cue (Jarbe et al. 1975). These procedures

have been used by two laboratories to evaluate a series of PCP

analogues with respect to similar stimulus properties using food

reward in the squirrel monkey (Brady and Balster 1981) and shock

avoidance in the rat (Shannon 1981). Both studies demonstrated that

N-substituted phenylcyclohexylpiperidines generalize to the PCP cue

and, in addition, provided a basis for rank ordering the compounds

by relative potencies. Shannon (1981) studied 17 analogues of PCP

and found that neither the 4-hydroxycyclohexane metabolite nor three

carbonitrile analogues generalized to PCP.

c. Behavioral Toxicity

Procedures for comparing the sensory and motor effects of

dissociative anesthetics have been limited to a single report (Lukas

et al. 1980) showing that both PCP and ketamine disrupt auditory and

visual thresholds and reaction times in unrestrained baboons. The

data demonstrated a definitive difference in time course with PCP

markedly disrupting reaction-time performance throughout the two-tothree hour test session, while recovery from ketamine's effects were

evident after one hour.

Three studies, however, have assessed PCP effects on size

discrimination in rhesus monkeys (Brown and Bass 1967)) matching-tosample task in pigeons (McMillan 1980), and acquisition and

performance of conditional visual discriminations in Patas monkeys

(Thompson and Moerschbaecher 1982). In general, PCP produced dose

related increases in reaction times and percent errors.

References

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phencyclidine. Clin Toxicol, 9:513-528, 1976.

Balster, R.L.; Johanson, C.E.; Harris, R.T.; and Schuster, C.R.

Phencyclidine self-administration in the rhesus monkey.

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Balster, R.L., and Woolverton, W.L. Continuous-access phencyclidine

self-administration by rhesus monkeys leading to physical

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Brady. K.T.. and Balster. R.L. Discriminative stimulus properties

of phencyclidine and five analogues in the squirrel monkey.

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Brady, K.T.; Balster, R.L.; and May, E.L. Stereoisomers of

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avoidance behavior in rhesus monkey: A psychophysical

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phencyclidine (PCP) in the rat. Pharmacologist, 23:192, 1981.

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analogues: Effects on behavior, electroencephalogram (EEG) and

EEG power spectra of the rat. Pharmacologist 24:134, 1982.

McKearney, J.W., and Barrett, J.E. Schedule-controlled behavior and

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McMillan, D.E. Effects of drugs on delayed matching to sample in

the pigeon. Pharmacology 22:293, 1980

Moreton, J.E.; Meisch, R.A.; Stark L.; and Thompson, T. Ketamine

self-administration by the rhesus monkey. J Phirmacol Exp

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Murray, T.F. The effects of phencyclidine on operant behavior in

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Overton, D.A. A comparison of the discriminable CNS effects of

ketamine, phencyclidine, and pentobarbital. Arch Int

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phencyclidine on food-rewarded bar pressing behavior in the

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potencies of phencyclidine and analogs in the opiate receptor

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Lazduanski, M. Interaction of phencyclidine ("angel dust")

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the behavioral effects of phencyclidine by repeated drug

exposure and body weight changes. Pharmacol Biochem Behav, 12:

761-766, 1980.

Zukin, S.R., and Zukin, R.S. Specific 3H-phencyclidine binding in

rat central nervous system. Proc Natl Acad Sci, 76:5372-5376,

1979.

93
VI. Human Testing Procedures

A. General Methods

1. Procedures for Characterization of Druq Effects

a. Acute Drug Effects

i) Analgesia

Procedures for the evaluation of the pain-relieving properties of

drugs in humans have developed along essentially two distinct lines

of investigation based on the origin of pain. In one, the empiric

drug trial, the studies are carried out in patients being treated

for clinical pain, usually, though not invariably, in a hospital

setting. In the other, the psychophysiological approach, the

studies are usually of experimentally induced pain and carried out

in normal volunteers or ambulatory chronic pain patients, most often

in a laboratory setting. Although the measurement of pain is common

to both methodologies, there are fundamental differences in concept,

applicability, and results of the two approaches.

In clinical pain, the strength of the stimulus is an unknown, and

inferred only from the medical history, physical examination,

ancillary clinical laboratory data, and the physician's past

experience. In a clinical drug trial meeting the basic requirements

for a well-designed study, the patient's report of pain is accepted

at face value and the effectiveness of any drug in relieving pain is

considered to be meaningful only in relative terms - that is,

relative to an appropriate standard drug or control medication. By

contrast, in psychophysiological experiments in which pain is

experimentally induced, the external stimulus is measurable and

controlled by the investigator or subject whose subjective responses

can be scaled in terms of the strength of the stimulus. Psychophysiological methods more often measure the effects of drugs on one

or more components of the pain complex and express the results in

absolute terms - that is, in terms of the criterion itself.

It has been long recognized that pain is a highly complex,

multifaceted experience with strong affective and motivational

94
components (Beecher 1959; Sternbach 1968; Melzack and Torgerson

1971), and great strides have been made in the development of

laboratory methods for parceling out and measuring these elusive

variables (Clark 1974; Tursky 1975; Gracely 1980; Price et al.

1983). Early psychophysical tests concentrated on the pain

threshold and the dol scale of pain intensity (Wolff et al. 1941;

Hardy et al. 1947, 1952). A wide variety of measurable external

stimuli have been employed, including radiant heat and other forms of

thermal stimulation, electrical shocks, a variety of pressure and

physical devices, chemical methods, and arterial occlusion (Beecher

1957; Stevens et al. 1958; Smith et al. 1966; Yanaura et al. 1977;

Lim et al. 1967; Procacci et al. 1979). But the ability of methods

based solely on measurements of pain threshold to predict the

efficacy of an analgesic in the clinical setting has been

disappointing (Kutscher and Kutscher 1957; Beecher 1959; FDA

Guidelines 1979). 

The results of experimental procedures to measure pain tolerance

have been more concordant with the clinical experience but have

raised ethical issues and, like clinical pain, they are

unidimensional - measuring only pain intensity. More recent

developments of psychophysical measurement procedures have, however,

made it feasible to evaluate some of the qualitative as well as the

quantitative aspects of the pain experience. Most of these

approaches are based on signal detection theory (Green and Swets

1966), magnitude estimation and ratio stimulus scaling (Stevens

1975), or cross-modality matching techniques (Tursky 1975). These

methods have provided means of estimating the contributions of

discrimination sensitivity and response criterion shifts in

evaluating placebo effects and in the diagnostic evaluation of

individual and group expressions of pain, particularly in patients

with chronic benign pain. Measures based on signal detection theory

have also been employed to distinguish analgesics from psychotropic

drugs (Chapman 1977; Wolff et al. 1976; Yang et al. 1979). For

reliable results, however, psychophysical metholds generally require

that the subject submit to large numbers of painful stimuli under

conditions which limit use in seriously ill patients with either

acute or severe pain due to trauma or disease. Thus, in spite of

the more rigorous controls which can be applied in the laboratory,

the well-designed controlled clinical drug study is still regarded

as the crucial test of a drug's analgesic properties (FDA Guidelines

1979).

Psychophysical measurements of pain also include multidimensional

pain descriptor scales and attempts to generate independent

magnitude estimates for each of the pain descriptor categories. The

McGill Pain Questionnaire (Melzack 1975) has been used chiefly to

characterize clinical pain although, more recently, it has also been

used to measure the efficacy of analgesics (Heidrich et al. 1983).

This questionnaire consists of sets of words designed to quantify

three dimensions of the pain experience in terms of the intensity

ratings and number of descriptors chosen by the patient with pain.

Other recent studies employing magnitude estimation, ratio stimulus

scaling, and cross-modality matching procedures have shown that

verbal meanings to an individual may be appropriately quantified

95
either by the individual or a similar group of individuals (Gracely

et al. 1978). These techniques have proven effective in

distinguishing the sensory from the hedonic effects of narcotics and

anxiolytics (Gracely and Dubner 1981). The advantages of these

methods lie in their potential for characterizing how drugs act,

rather than as a means of evaluating the analgesic efficacy or

potency of new drugs.

Scientifically acceptable principles for the conduct of clinical

analgesic drug trials were pioneered and elucidated over 25 years

ago by Beecher (1957). He emphasized that while reliance must be

placed on the verbal report of the patient with "pathological pain,"

reliability and validity can be achieved by appropriate experimental

designs and controls including the double- blind technique,

randomization of treatments, appropriate active drug standards and

placebo controls, and statistical verification of the significance

of the results. On this foundation, a number of different drug

testing models have been developed using either single or repeated,

fixed or graded, doses of the test drug and reference standard, with

or without placebo controls, and administered by one or more of

several different routes to patients with acute or chronic pains of

diverse etiologies (Keats 1956; Houde et al. 1960; Lasagna 1960;

Bellville et al. 1968; Sunshine 1980). In addition, a well-designed

study should include an internal control - a means of demonstrating

within each study that the sample population and the experimental

procedure are capable of discriminating between a.known active drug

standard and a placebo, or between graded doses of the standard - in

terms of the analgesic data generated (Modell and Houde 1958; Houde

et al 1965). There are many factors, however, which enter into the

choice of the appropriate experimental design, standards and

controls, and methods of collecting and analyzing the data. Among

them are the general pharmacological profile based upon laboratory

animal and preclinical data, and the expected role the drug will

fill in medical practice (FDA Guidelines 1979). Pharmacokinetic

characteristics of the test and reference drugs will also influence

the choice of single or repeated doses, and crossover or noncrossover experimental designs, the timing of drug administration

and data collection, and the manner in which the results are

analyzed. Decisions are commonly based on whether a drug 1) is

morphine-like, 2) has associated narcotic antagonist properties, 3)

is aspirin-like or, 4) does not fit any of these stereotypes (FDA

Guidelines 1979).

Because of the great variety of clinical pains for which analgesics

with potential for abuse are employed, there is no single

experimental design or method of conducting a clinical analgesic

drug assay which is appropriate for all situations. The procedure

followed most frequently is to employ fulltime trained observers,

usually registered nurses, who administer the study medication for

complaints of moderate or severe pain. Observations are generally

made prior to and at frequent regular intervals for 4 to 6 hours

after medication or until pain is reported to have returned to its

premedication intensity. For pain which is short-lived, such as

postpartum pain (Bloomfield et al. 1970) and that following many

types of surgery including the extraction of impacted molars (Cooper

96
and Beaver 1976), the patient receives only one treatment or test

medication, and comparisons of the drugs are made in parallel

patient groups. Studies carried out in situations in which the pain

is expected to be more long-lasting, such as in patients undergoing

more formidable surgical procedures, and patients with pain due to

cancer, some form of crossover design may be employed in which each

patient receives single doses of each test drug on separate days

(Houde et al. 1965). The objectives of most of these studies are

not only to test for analgesic efficacy but also to provide an

estimate of the potency of the test drug relative to a known or

established standard analgesic. The methodology for chronic or

repeated dose administration of new drugs has not been as well

characterized, but some. procedures incorporating many of the

features of the single dose studies have been reported (Wang et al.

1981; Ouellette 1982). These methods allow for comparisons of the

effects of the initial doses of drugs with those at proximate

steady-state levels. This can be of considerable importance in

evaluating drugs which are cleared from the body more slowly or have

active metabolites.

Virtually all of the methods of evaluating pain in clinical settings

rely on the patient's own report. While a number of objective

methods - including the measurement of a variety of autonomic signs

(Beecher 1957) and biochemical correlates of pain (Terenius 1978),

of recordings of evoked brain potentials (Chapman et al. 1979)) and

of observations of behavior (Fordyce 1976) - have been employed in a

variety of clinical settings, these methods have had limited

applicability and have been of questionable reliability in

predicting the efficacy of an analgesic (Houde 1982). Most clinical

methods employ some form of categorical or ordinal scale for rating

pain severity. Typically, analgesia is measured indirectly in terms

of changes in pain intensity levels on a 4-5 point scale at

intervals after administration of the test medication in comparison

with the levels of pain before administration. Analgesia is also

measured directly in terms of pain relief estimates. Comparison of

analgesic drugs is generally made in terms of the cusum of

observational period scores and commonly expressed as SPID [sum of

pain intensity differences] or TOTPAR [total pain relief] (Forrest

et al. 1963). Quanta1 measures of pain relief or comfort have also

been employed successfully, often together with the graded rating

1975).

scales, but they tend to be less sensitive (Wallenstein and Houde

Visual analogue scales (VAS) are also being used to measure pain

intensity, pain relief, and mood effects. The VAS is usually

presented in the form of a 100 mm horizontal line with no other

markings except at their ends which are identified as "no pain" and

"worst possible pain, " "complete pain relief" and "no pain relief,"

or "best mood" and "worst mood." The patient places a mark between

these extremes to indicate his subjective impressions. The VAS has

also been presented in the form of a dial, pain thermometer, and

vertical line, but none of these variations provide any advantages

(Huskisson 1974; Littlejohns and Vere 1981). Good agreement has

been found between the ordinal pain scales and the VAS, but the

latter tend to be more sensitive (Ohnhaus and Adler 1975;

97
Wallenstein et al. 1980). Despite some debatable assumptions of the

mathematical properties of the ordinal scales employed (Littlejohns

and Vere 1981), the results of clinical relative analgesic potency

assays by most investigators have, in general, been consistent

whether analyzed by standard parametric tests or by non-parametric

tests (Fucella et al. 1977).

In summary, there are basic differences in the approach to measuring

pain when the objective is the evaluation of an analgesic and when

the objective is the evaluation of the nature of pain. The latter

is recognized as a complex process whose multidimensional features

can best be delineated in a controlled laboratory setting using a

variety of psychophysiological methods. By contrast, the assessment

of analgesic potency and efficacy of a drug is judged in terms of a

unidimensional scale of pain intensity and compared with an

established standard analgesic in a clinical setting. The two

approaches to measuring pain complement eash other, but the

psychophysical techniques are limited in their ability to detect the

effects of simple analgesics and by their often not being applicable

to studies of analgesics in patients with acute pain.

ii) Electroencephalographic (EEG) activity and

sleep

The EEG recorded from multiple scalp electrodes has been utilized in

two basic modes to assess 1) acute effects of a compound on the

electrical activity of the brain and 2) effects of a compound on the

sleep-awake cycle. While the basic equipment is similar, these two

types of studies involve procedural differences that relate to

recording session length, methods of analysis, and in some cases,

recording montage and method of electrode attachment (Fink 1968).

The acute effects of a compound are usually evaluated during a

relatively short (e.g., 3-5 hour) recording session (Itil 1972)

employing an abbreviated method of attaching the scalp electrodes.

Electrodes are usually placed according to the l0-20 system (Jasper

1958) but not all 21 possible electrodes are utilized. Recording

montages are usually selected on the basis of special interest or

because the experiment is designed to study the drug-induced changes

in certain neuronal structures immediately below the electrodes.

Before administering the drug, a sufficiently long period is

customarily devoted to obtaining control, awake recordings under

various conditions, (e.g., eyes closed and relaxed, eyes open and

relaxed, and eyes open and concentrating). These sessions serve as

the basis for assessing drug-induced changes (Longo 1977). The EEG

effects can also be correlated with subjective ratings of euphoria

and other physiological responses (Volavka et al. 1973).

Quantitation of the EEG effects is either by visual inspection,

integrated voltage, period analysis, or power spectral analysis.

The study of effects of drugs on the sleep-awake cycle necessitates

some procedural refinements. Recording sessions typically last 6-8

hours and the EEG train is described by visual inspection in terms

of frequency and amplitude components. These procedures have

provided standardized characteristics that define the various phases

98
of sleep (Rechtshaffen and Kales 1968; Niedermeyer 1982). The sleep

profile has identifiable onset and duration characteristics

involving the various sleep stages I-IV and REM sleep, as well as

amplitude and frequency characteristics involving the clinically

relevant band widths of delta (O-4Hz), theta (4-8 Hz), alpha (8-13

Hz), and beta (13-25 Hz).

Sleep stages have been characterized by their EEG profiles and

rarely by simply observing the subject (Harvey 1980). During stage

0 or awake, beta activity predominates while the eyes are open and

alpha activity while the eyes are closed. Stage I or the decending/

dozing stage is characterized by theta, alpha, and beta activity

with occasional bursts of eye movement. The onset of sleep during

stage II is heralded by the appearance of sleep spindles and K

complexes and the disappearance of alpha and beta activity; theta

waves, however, remain. Stage III or deep sleep transition is

differentiated from Stage II by the appearance of slow delta waves

along with the other waveforms. Deep sleep or Stage IV occurs when

only delta activity prevails with little or no eye movement. Rapideye-movement or REM sleep is a stage that typically follows IV. It

is characterized by many different frequencies, but no spindles or K

complexes are seen. There is a corresponding increase in the amount

of eye movement. Most dreaming occurs during this stage.

Quantification of the EEG activity has been, accomplished by

integrating the amplitude (Goldstein et al. 1963) using a pulse

generator. Temporal frequency changes have also been identified

using period analysis (Burch et al. 1964), by measuring the time

interval between two successive baseline crossings of the EEG

waveforms. EEG power spectral analysis is based upon the

analog-to-digital conversion of the EEG activity which is then

mathematically fitted to a sine/cosine signal. The Fast Fourier

Transform (Grass and Gibbs 1938; Walter 1963), when applied to such

data, results in a fingerprint-like array that characterizes the

amplitude and frequency components simultaneously.

2. Procedures for Assessing Physical Dependence Potential

a. Tolerance to Drug Effects

The development of tolerance to a drug effect is reflected by an

attenuation of the original response with continued exposure to the

drug. The extent of tolerance is determined by the specific drug,

the parameter being assessed, and the dose and duration of

treatment. Thus, once the acute effects of a compound are

characterized, the development of tolerance can be tracked during

treatment.

The assessment of tolerance to a drug's effect must take into

account numerous factors (Martin and Sloan 1977). First, with

chronic administration, the drug's pharmacologic profile may change

due to differential tolerance development to some of the effects and

not others. This complicates subsequent reassessment of the drug's

effects. Secondly, an appropriate baseline of effects must be

established. The time interval from the last dose to the initiation

99
of the procedures for assessing tolerance must be carefully

determined, since certain acute effects may be confounded with the

appearance of early abstinence signs. Thirdly, appropriate doseresponse relationships should be demonstrated before chronic

exposure is begun. Thus, a "tolerance index" can be used to compare

different drugs (Martin and Fraser 1961).

b. Withdrawal from Chronic Drug Administration - -

The assessment of the dependence potential of a class of drugs

generally focuses upon procedures far identification and

quantification of the behavioral and physiological changes following

cessation of chronic drug exposure. Characteristically, such

effects depend primarily on the drug being studied, the maintenance

dose, and the duration of treatment. Two different techniques are

employed to assess the consequences of withdrawal from chronic drug

exposure. Direct or primary dependence assessment involves using a

drug-free (but not necessarily a drug-naive) subject given

increasing doses of a test compound for a predetermined length of

time (Fraser et al. 1957). Upon withdrawing the drug, the subjects

are assessed at regular intervals for changes in physiological

processes, vital signs, and subjective or "mood" changes. These

signs and symptoms are rank-ordered with respect to their degree of

severity and then used as the basis for describing the syndrome as

either mild, moderate, or severe (Martin and Gorodetzky 1965;

Jasinski et al. (1971b).

The substitution procedure uses subjects currently receiving a

compound on a chronic basis. The test compound is then substituted

for a predetermined length of time, usually on a double-blind basis

(Jasinski 1977). The signs and symptoms are recorded and tabulated,

with the presence or absence of signs reflecting the degree of

similarity between the test drug and the standard drug.

3. Procedures for Assessing Abuse Liability

a. Drug Self-Administration

Drug self-administration procedures have evolved over the past 18

years and at present have been refined by the introduction of more

standardized techniques and an adherence to certain basic guidelines

(Bigelow et al. 1976). Assessments of true drug-seeking behavior

must satisfy the following criteria: 1) ingestion or injection of

the drug must be optional (Mello and Mendelson 1965), 2) the

behavior must be observed during a number of experimental

manipulations (Nathan et al. 1970). and 3) the drug selfadministration patterns must be established over an appropriately

long time interval (Cohen et al. 1971).

In general, two basic subject populations have been studied. Normal

volunteers reporting to the lab for only a few hours each day have

been used for short-term determinations of reinforcing efficacy

(Johanson and Uhlenhuth 1978), while residential ward settings have

been used with drug-experienced subjects (Bigelow et al. 1975; Mello

and Mendelson 1978; Fischman and Schuster 1982). In the short-term

100
studies, subjects receive a physical examination and are given drug

or placebo either orally or by injection. Typically, double-blind

conditions are observed. At various time intervals, the subjects

are asked to evaluate their feelings while physiological responses

are monitored. This procedure provides baseline data against which

choice procedures (i.e., between two different drugs) can be

evaluated.

In residential ward settings, subjects usually are given control of

access to drug. In addition, they are required to work (e.g., ride

an exercise bicycle for 15 minutes) in order to obtain drug

(Griffiths et al. 1979). This procedure tends to reduce variability

and increase sensitivity by restricting the drug availability. The

residential ward setting also provides a built-in control for

environmental factors such as social interactions (Bigelow et al.

1974). This procedure is also conducive to studying the patterns of

drug self-administration over time as a basis for the assessment of

tolerance and dependence.

Prior to the initiation of studies with a volunteer participant, an

extensive drug history should be obtained. In addition, the

subjects should be evaluated for current tolerance and/or dependence

to certain drug classes either by giving a test dose or by requiring

a controlled observation period to determine whether withdrawal

signs appear.

b. Self-Report of Drug Effects

Procedures for assessing changes in subjective feelings have been

developed to aid in the identification of drug-induced effects

superimposed on the subject's baseline mood state. The most widely

utilized test, the Addiction Research Center Inventory or ARC1 (Hill

et al. 1963) is a series of 550 questions that are answered "yes" or

"no. " The content of the items reflects a broad range of physical,

emotional, and generally subjective effects of drugs. Additionally,

certain questions are designed to provide some measure of

personality and psychiatric stability (Haertzen et al. 1963). In

some circumstances, an additional 50 questions are asked which

provide information on the sociologic profile of the subject. The

most useful information obtained when applying this test is the

average scores on the 38 recommended scales (see Haertzen 1974),

since these averages can serve as a standard for comparison in the

classification of subjects or groups. Essentially, validation of

the ARCI has resulted in the identification of six psychiatric

groupings and ten drug conditions. The six psychiatric groupings

are: normal, mentally ill, alcoholics, criminals, opiate addicts,

and simulated mental illness. The ten experimental drug conditions

are: no-drug, morphine, cyclazocine and nalorphine, pentobarbital,

chlorpromazine, LSD, benzedrine, alcohol, opiate withdrawal, and

chronic opiate maintained.

Many pharmacological studies have used short forms or single scales

of the ARCI. These forms can be quickly administered and are

especially useful in demonstrating whether a drug effect is present,

its potency, and its duration of action. Three scales in particular

101
have been used to identify sedative-(PCAG), euphoric-(MBG), and

psychotomimetic-(LSD) effects (Martin 1967; Jasinski et al. 1971a).

Morphine and morphine-like drugs elevate MBG scores but not PCAG or

LSD, while nalorphine and nalorphine-like drugs elevate LSD and PCAG

scores but not MBG scores. Central nervous system depressants

elevate PCAG scores and not LSD or MBG scores. The utility and

validity of these short forms have been individually substantiated

by the good correlations of relative potencies and time-action

curves for the resultant assessment of subjective effects and other

physiological responses such as pupillary diameter and nystagmus

(Martin 1967; Jasinski et al. 1971a).

The method of classification of subjective effects inherent in the

ARCI is based on discriminant function procedures described by Rao

(1952). The advantage of this procedure is that it provides a

mechanism that corrects for individual differences and variability.

Full details of the correcting procedures are provided by Haertzen

(1974).

Various studies have shown that ARCI scales may have a factor

analytic or structural significance that is constant in going from

one group to another or in testing from one condition to another.

When individuals are tested under different conditions, however,

different individuals may be placed in the same category. The

successful implementation of the ARCI and resultant predictive power

of the scales are also dependent on the specific test-taking

instructions given, and as such, should be standardized (Haertzen

1974).

c. Behavioral Toxicity

Procedures for assessing drug-induced changes in human performance

have been developed and refined for psychomotor and reaction time

procedures. Little consistent data are available on more complex

behaviors and functions, however (Johnson and Chernik 1982). The

role of such behavioral toxicology assessment has become

increasingly important because substances with little or no

disruptive behavioral or pharmacological effects are not regarded as

having significant abuse liability. In contrast, compounds that are

used even sparingly which produce marked disruptive changes are

considered to have high abuse liability. Since drugs may fall

anywhere on the continuum defined by these parameters, relative

abuse liability is most effectively determined by a comprehensive

assessment of these interactive dimensions.

A relevant issue that must be addressed when conducting these

studies is whether the procedure distinguishes between drug-induced

sensory. decrements (or improvements) on the one hand, and motor

impairment on the other. While it is quite clear that relatively

large doses of CNS depressants will sedate a subject so that he is

incapable of performing, these types of studies do not provide a

complete profile of the drugs being assessed. Rather, the

determination of the threshold dose for performance decrement and

the duration of such decrement more appropriately characterize the

behavioral toxicity of a compound. Recently, performance decrements

102
that occur the day after a hypnotic is used ("hangover effect") have

been studied more closely (Johnson and Chernik 1982; Hindmarch 1980;

Wittenborn 1979). These studies have been generated primarily in

response to the Institute of Medicine's report dealing with the

issues surrounding the relationship of short- versus long-acting

hypnotics and performance decrements (Solomon 1979).

The assessment of performance is also useful in the evaluation of

subjects on chronic methadone maintenance (Rothenberg et al. 1977)

and the enhancement of performance after caffeine and amphetamine

(Weiss and Laties 1962). Basic changes in auditory and visual

thresholds following marijuana administration have also been

documented (Caldwell et al. 1969).

Acute studies are conducted over the course of a few hours with

appropriate collection of pre-drug control performance before

administration. Depending on the procedure employed, the evaluation

is conducted continuously, or in discrete sessions separated by

standard time-out periods until the drug effect has peaked and

waned. In studies designed to assess the "hangover effect," the

subject first undergoes an assessment of performance during the day,

and then receives the drug either later in the day or at bedtime.

Performance is again assessed at various times the following day.

Procedures typically employed include visual tracking (Borland and

Nicholson 1974), auditory vigilance (Jones et al. 1978), and tests

such as continuous arithmetic (Epstein and Lasagna 1968). time

estimation (Pfaff 1968), free recall (Murdock 1962), and digit

symbol substitution (Wechsler.1944). In addition, some procedures

are designed to mimic real life situations such as automobile

driving (Biehl 1979; Hindmarch et al. 1977), while one study has

actually employed real-life driving in downtown traffic (Klonoff

1974).

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109
B. Procedures for Assessing Opioids

1. Characterization of Opioid Drug Effects

a. Acute Drug Effects

i) Analgesia

Methods of assessing the analgesic efficacy and potency of opioids

are greatly influenced by the singular pharmacological properties of

this class of drugs - which includes not only the opiates and their

semisynthetic and synthetic derivatives with both morphine-like and

morphine antagonist properties, but also a large and growing number

of chemically unrelated compounds with similar disparate attributes

- linked together in that they are believed to act upon the same

family of CNS receptors, though with different affinities and

intrinsic activities. Among their most distinctive features are

that they include some of the most potent and universally

efficacious drugs for the control of clinical pain and that their

analgesic properties are greatly influenced by their repeated use

and by prior exposure to drugs of the same class. These factors are

important determinants not only of the setting and subject/patient

population in which opioids are evaluated, but also of how opioids

are evaluated for their analgesic efficacy.

The analgesic properties of most opioids are readily demonstrated in

double-blind, controlled clinical trials employing categorical or

visual analogue scales of pain intensity or pain relief. Moreover,

in clinical assays of graded doses of opioids in patients with

either postoperative or cancer pain, the analgesic response

[measured in terms of pain intensity reduction or pain relief ] has

been observed to be linearly related to the logarithm of the dose

(Houde et al. 1965). Relative analgesic potency assays comparing

graded doses of parenteral morphine to those of other opioids have

shown remarkably consistent results in terms of the cusum scores of

these parameters of pain relief (SPID, TOTPAR) in studies of

patients with postoperative and cancer pain (Wallenstein and Houde,

1975).

Unlike many nonopioid analgesics, the opioid's can be administered by

several parenteral or enteral routes. However, their potencies are

substantially influenced by the route of administration (Beaver et

al. 1977). Flexibility in dose and route of administration provides

an opportunity to evaluate the opioids in a wide variety of pain

models, but, since many have substantial abuse liability, they are

generally tested in hospitalized patients with severe or acute pain.

Graded parenteral doses are usually the most efficient and often the

only feasible method of administering these drugs in this setting.

The most frequently employed pain model used for assaying patient

opioid analgesics is postoperative pain, although analgesic assays

have also been carried out in patients with cancer, biliary or renal

colic, myocardial infarctions, severe or extensive burns, and women

in labor (Lasagna 1960).

110
Although the effects of various opioids can be demonstrated in a

variety of laboratory models employing experimentally induced pain,

these techniques have not consistently and accurately predicted the

clinical efficacy of new analgesics (FDA Guidelines, 1979).

However, some of the psychophysical scaling techniques which were

developed using normal volunteers have been applied to the clinical

setting and have been useful in distinguishing the sensory and

hedonic actions of morphine from those of diazepam (Gracely et al.

1978). Psychophysical procedures which assess pain in its different

dimensions have also provided useful guides to the appropriateness

and usefulness of opioids

(Sternbach 1978).

in certain chronic pain syndromes

The design of a clinical analgesic study of a new opioid will

primarily be determined by the role it is expected to fill - or the

opioid that it is expected to replace - in the therapeutic

armamentarium. Drugs that are expected to substitute for codeine

are commonly tested orally against codeine or propoxyphene, either

alone or in combination with aspirin, acetaminophen or one of the

newer non-steroidal antiinflammatory drugs. These studies are most

frequently carried out in patients with postpartum pain, oral

surgery pain, headaches, or rheumatic and orthopedic pains (Beaver,

1966). A variety of acceptable experimental designs are employed

including single fixed dose comparisons of the test drug with a

known analgesic standard and placebo control, parallel line assays

of graded doses of the test drug and standard, with or without a

placebo control. Factorial studies have also been carried out of

fixed doses of the test drug, standard drug, combined test and

standard drugs, and a placebo control (Houde et al. 1960). Ordinal

and visual analogue scales are most frequently employed, and the

results analyzed using standard parametric and nonparametric

statistical tests (Fucella et al. 1977; Quiding et al. 1981).

Relative oral/parenteral analgesic assays can also be carried out

using graded doses of the two forms of the drug and a double placebo

technique in which each study medication consists of both a capsule

(or tablet) and an injection, one containing the active drug and the

other a placebo (Beaver et al. 1967).

Opioids that are being considered as analgesic substitutes for

morphine or its surrogates are frequently tested first in

parenteral, graded dose, relative to analgesic potency assays with

morphine as the reference standard. The relative potency comparison

allows for an assessment of acute side effects or adverse effects at

equianalgesic doses of the test drug and standard. These assays

have been carried out as non-crossover, twin crossover (incomplete

block) or complete crossover designed studies in hospitalized

patients with moderate to severe pain following major surgery and in

patients with advanced cancer (Wallenstein and Houde 1975).

Categorical and VAS measures of pain intensity are customarily

employed and standard parametric and non-parametric statistical

analyses are carried out on the peak and total (SPIC, TOTPAR)

scores. Multivariate analyses can be performed on the data using

computer programs. Concomitant assessment of the patient's

emotional status can also be carried out using the McGill

Questionnaire (Heidrich et al. 1983), as well as by adjective check

111
lists and visual analogue measures of mood (Kaiko et al. 1981a). In

addition, differential effects on mood can be demonstrated between

morphine and some of the newer nonsteroidal antiinflammatory drugs

in doses which are equianalgesic in terms of measures of pain

intensity (Wallenstein 1981a,b). In similar analgesic assays,

equianalgesic doses of morphine and heroin produced almost identical

changes in mood (Kaiko et al. 1981a,b).

Since the empirical clinical analgesic assay is meaningful only in

terms of comparison between a test drug and a reference standard,

differences between the test and standard drugs in cross-tolerance

to a prior opioid will influence the estimate of their relative

potency. That effect appears to be minimal in most studies of

postoperative and postpartum pain, and even in cancer pain when the

test and standard drugs are different from the prior medication, and

when prior opioid exposure has not been excessive in dose,

frequency, or duration. However, when assaying agonist/antagonist

analgesics this can be a problem because sensitivity to the

antagonist actions is directly proportional to the degree of

physical dependence, and this effect can override both the agonist

actions of the test drug and the diminished efficacy of the standard

drug as a consequence of cross-tolerance (Houde 1979).

Assessment of analgesic potency based on chronic administration of

opioids has been limited and tends to be confounded by a large

number of uncontrollable variables which can develop in the course

of the analgesic assay. Nevertheless, short-term repeated dose

comparisons of fixed doses of opioids have been carried out in a

noncrossover design. In these studies, the patients received the

test or standard drug for periods of up to two to three days, on

request (prn), for moderate or severe pain. Ordinal scale

measurements of pain intensity and pain relief were made on the

first, second, and third days of the study prior to and at 1/2, 1,

2, 3, 4, 5, 6, 7, 8 hours following study medication, or until pain

returned to premeditation level (Wang et al, 1981; Ouellette, 1982).

These studies provide comparisons of the analgesic effects of the

test and standard drugs on the first day with those of the second

and third days. Proximate plateau plasma concentrations of the

drugs will be reached at that time if their terminal elimination

half lives are sufficiently short, as they would be with morphine,

the usual standard in these studies. Graded doses of at least the

standard drug are recommended as an internal control - or measure of

the sensitivity of the study - since placebo controls without rescue

medications would be inappropriate in a double-blind, repeated dose

study of this design.

ii) Electroencephalographic (EEG) activity and

sleep

As mentioned in the General Methods Section, the EEG is used to

measure both the acute effects of drugs and long-term effects on the

sleep-awake cycle. Almost all studies to date have used visual

inspection techniques to characterize the EEG profile of opioid

administration. In general,

to nontolerant,

morphine or heroin acutely administered

nondependent subjects produces a number of changes

112
in the EEG consisting of: slowed alpha frequency, appearance of

delta activity, and an increase in theta activity (for review, see

Martin and Kay 1977). Fink et al. (1968), Zaks et al. (1969), and

Volavka et al. (1970) demonstrated that heroin-induced changes in

the EEG were time dependent and could be divided into an early

response (first 4 minutes) and a later response (5-32 minutes). The

most prominent effect was that the alpha activity was increased

early in the response and subsequently decreased in the late

response. Additionally, these studies showed that these EEG effects

of heroin could be reversed or blocked by naloxone. Similar

techniques have been applied to assess the acute effects of

methadone (Isbell et al. 1947) and cyclazocine (Fink et al. 1969).

The use of power spectral analysis to quantify the effects of

opioids has been relatively limited. Volavka et al. (1974)

demonstrated that the above-mentioned changes in EEG activity were

related to other physiologic measures such as respiratory rate and

heart rate, and the behavioral measures of subject's mood. Power

spectral analysis has also been used to monitor the EEG changes in

heroin addicts during induction and maintenance on methadone

(Feinstein and Hanley 1975). The EEG effects of heroin were

characterized by a prominent, sharply peaked narrow band (9-10 Hz)

of activity. When the subjects were started on methadone, this

alpha peak was substantially attenuated and the predominant

frequency was shifted to 11-12 Hz. In one subject who continued to

use heroin (verified by urinalysis), these shifts did not occur.

Studies on the effects of acute opioid administration on sleep-awake

patterns have been limited to only a few laboratories where such

extensive amounts of data can be gathered. Kay et al. (1979)

compared the effects of acute morphine, heroin, and methadone in

nondependent opioid abusers using double-blind, cross-over design.

Subjects were given single i.m. doses of drug or placebo at weekly

intervals. Each experiment consisted of one adaptation night

followed by an injection night. All three compounds produced

similar changes (as revealed by computer-derived bisector analysis)

in the EEG and muscle activity. These included increased time

awake, increased muscle activity, increased alpha activity, and

decreased theta activity and decreased spindling. Using standard

bioassay techniques (Finney 1964), heroin was about twice as potent

as morphine, which was found to be equipotent with methadone. Other

physiologic measures such as ocular activity and respiration

correlated very well with the EEG sleep stages.

iii) Gastrointestinal motility

The opioids are the most effective antidiarrheal drugs currently

available. Procedures for assessing the passage of intestinal

contents through the G.I. tract have been developed and incorporate

one of three basic designs: 1) monitor the appearance of an orally

ingested dye in the feces (Rothman and Katz 1964); 2) monitor the

progress of a barium-treated meal through the G.I. tract using X-ray

technology (Manousos et al. 1967); and 3) intubation (Hunt and

Spurrell 1951) for sampling gastric contents. The effects of acute

doses of various opioids can then be assessed by the percent of

113
retarded progress of a meal through the G.I. tract. A continual

slow passage of markers through the G.I. tract is indicative of a

lack of tolerance to the constipating effects of opioids. Tolerance

to the effects of opiates on gastrointestinal motility develops only

extremely slowly. In man, tolerance to the constipating effects of

methadone may become apparent after as long as three years of

continual use (Kreek 1973; 1978).

iv) Cough

Procedures for assessing the antitussive effects of opioids have

relied mainly on the use of subjects with pathological cough (e.g.,

obstructive emphysema or chronic bronchitis) rather than

experimentally induced cough (e.g., inhalation of noxious vapors).

The significant developments in this regard relate mainly to methods

utilized to record the incidence and severity of coughs. Sevelius

and Colmore (1966) developed a method that uses a contact throat

microphone coupled to a voice-activated tape recorder via a long

flexible cord. This provided the subject essentially 'unrestrained

movement within the private room. Subjects reside in the room for

at least 6 consecutive days; the first 2 days are used to record

control cough levels. At a specified time (usually in the morning)

subjects receive either codeine, placebo, or a dose of test

substance. Cough rates are then calculated on an hourly basis until

the next morning. The number of coughs after medication is

represented as a percent of the number of coughs after placebo.

One method of experimental induction of cough utilizes citric acid

aerosal spray (Bicker-man 1956). Normal subjects are used, and after

receiving placebo or an antitussive, are exposed to the spray. The

frequency of coughs is recorded and used as an index for the degree

of cough suppression.

2. Opioid Physical Dependence Potential

a. Tolerance to Drug Effects

The nature and characteristics of the development of tolerance to

the effects of opioids in man have been extensively studied. The

basic premise behind the techniques employed to study tolerance

relates to obtaining accurate measures of dose-related drug effect

in the non-tolerant or opioid-free subject first. Then, once the

treatment regimen has been established, controlled reassessments of

the effects of single doses of the same opioid or a test compound

(cross-tolerance evaluation) can begin. This is most effectively

accomplished when multiple doses are studied. Thus, a ratio of the

effective doses in the nontolerant and tolerant state can be

calculated and becomes the "tolerance index" for that compound

(Martin and Fraser 1961).

Numerous problems exist when attempting to quantify the degree of

tolerance to opioids. First, because of different durations of

action, the time between the last maintenance dose and the

administration of the test dose for tolerance assessment can be

critical. Predetermined objective criteria must be established in

114
order to maintain uniformity between test sessions. Another problem

is that with repeated administration of opioids, the profile of

effects changes. For example, non-tolerant subjects report feeling

euphoric and sedate while tolerant individuals report apathy,

anhedonia, and withdrawal from social interactions (Haertzen and

Hooks 1969; Martin et al. 1973).

b. Withdrawal from Chronic Drug Administration - -

Procedures for assessing the dependence potential of opioids in man

have focused upon three main areas: substitution, single dose

suppression, and primary dependence. Typically, individuals with a

history of opioid use are employed since the procedures necessitate

a chronic steady state preparation.

In the substitution procedure, subjects are maintained on morphine

at a dose of 240 mg/day, s.c., given in divided doses. This

treatment is continued for 30 days,after which test procedures can

be initiated. Under double-blind conditions and beginning with the

evening dose (e.g., 10 p.m.), a dose of a test substance, placebo,

or the usual morphine dose is administered. This substitution is

continued for 24 hours during which the subject is observed at 1-

hour intervals. Signs and symptoms such as restlessness, emesis,

fever, hyperpnea, blood pressure, and body weight are scored

according to the Himmelsbach procedure (Jasinski 1977). The

intensity of withdrawal (should these test substances fail to

substitute for morphine) is scored on the basis of the signs present

and include: Mild - yawning, lacrimation, rhinorrhea, perspiration;

Moderate - muscle tremor, dilated pupils, goose flesh, anorexia;

Marked hyperpnea, restlessness, insomnia, elevated blood pressure;

Severe - emesis, diarrhea, weight loss. The original technique as

proposed by Himmelsbach (1937a) was adapted to incorporate a point

system based upon the composite signs of abstinence (Himmelsbach

1937b; Kolb and Himnelsbach 1938)) updated by Fraser and Isbell

(1960), and was reviewed by Jasinski (1977). By systematically

varying the dose of the test substance, the relative degree of its

dependence potential can be determined.

The single dose substitution procedure (Himmelsbach and Andrews

1943) was based on the premise that a compound that possesses

morphine-like activity should be able to suppress abstinence signs

in morphine-dependent subjects. This relatively quick method of

assessing a drug's dependence potential is conducted in subjects

that are stabilized on morphine as described above. At a specific

time, the morphine injections are terminated and the time course of

withdrawal is recorded. At 30 hours after the last morpine

injection, a dose of the test compound, placebo, or the normal

morphine dose is given and the evaluations continued. The relative

dependence potential of a compound is thus defined by its capacity

to ameliorate the signs and symptoms of withdrawal. Morphine

dependence is then reestablished for 1 week, at which time another

experiment is performed.

The conduct of primary dependence studies on opioids has not been

utilized as extensively as the substitution techniques. This is

115
mainly due to the paucity of suitable volunteer subjects and the

concern over the toxicity of the test compounds during long-term

administration. While the chronic administration of a compound is

considered to be the definitive test of its dependence potential,

this procedure is rarely performed first and usually is only

incorporated after the results of substitution or single dose

suppression studies are known. The procedure employs the use of

doses of the test compound that are equivalent to (i.e., similar

analgesia, suppression scores, degree of miosis, etc.) 240 mg of

morphine (Jasinski 1977). Care must be exercised to avoid the

accidental accumulation of drug or metabolites that result in toxic

reactions. Typically, the length of exposure is 18-20 days followed

by 10 days of placebo administration. Alternatively, naloxone can

be administered in order to precipitate withdrawal (Jasinski and

Nutt 1972), althougti this procedure is not widely used. Regardless

of the method used, the appearance of signs and symptoms of

abstinence is graded and the relative degree of dependence is

determined.

3. Opioid Abuse Liability

a. Drug Self-Administration

Because of numerous medical and ethical considerations, unresricted

self-administration of opioids such as morphine or heroin is not

possible. However, procedures that employ limited access have been

developed and provide ample opportunity to study the reinforcing

properties of opioids in human volunteers residing in an inpatient

ward (Mello et al. 1981, 1982). In these studies, heroin was

available every 6 or 8 hours and was self-injected (under

supervision by a physician) at a unit dose of 10 mg, i.v. The

contingency placed upon the subjects was that they work for points

that could then be used to "buy" the heroin dose. This was

accomplished using standard operant procedures that employed a fixed

ratio 300/fixed interval 1 second schedule of reinforcement. As an

added incentive for the subjects to remain on the research ward,

points could also be earned for money which would be paid upon

completion of the study.

Once stable response patterns are established, various manipulations

of the experimental conditions can be employed. To date, no studies

have been conducted that substitute unknown compounds for the

heroin. Rather, studies have been directed at pharmacological

interventions which include observing the changes in heroin

self-administration during chronic naltrexone (Mello et al. 1981) or

buprenorphine (Mello et al. 1982). Both naltrexone and

buprenorphine significantly decreased the number of heroin

self-ingections as compared to placebo conditions. Thus, these

studies satisfy the three criteria defined previously: optional drug

injections, various experimental manipulations were performed, and

the availability for heroin self-injection was over a 30-35 day

period.

An alternative procedure employs subjects that are receiving

methadone as a maintenance therapy for opioid dependence (Stitzer et

116
al. 1979, 1983; McLeod et al. 1983). In the studies by Stitzer et

al. (1979), participating subjects were maintained on an outpatient

basis but were required to attend the clinic each day for their dose

of methadone. On two days of the week, immediately after consuming

their regular dose of methadone, subjects were offered a choice

between an additional dose of methadone (0, 1, 5, 10, 25 or 50 mg)

and money (either $1 or $5). The selected item was given

immediately. In addition, subjects completed a 60-item symptom

check-list four times a week in order to assess signs of opioid

withdrawal. Urines were also analyzed for illicit drugs. These

studies demonstrate that methadone serves as reinforcer in

methadone-maintained subjects and that the relative frequency of

choosing methadone over money increased. as the dose of methadone was

increased. Further, the number of choices for methadone was greater

when the alternative was $1 when compared with $5. This procedure

provides an excellent model of human abuse liability testing and can

be expanded to include the assessment of other opioids, drugs

belonging to different classes, and different alternative choices.

McLeod et al. (1983) utilized methadone-maintained subjects that

resided on a research ward. Subjects regulated their own methadone

dose by completing a fixed number of responses on an operant console

during a 6-hour interval. Both constant and progressive ratio

procedures were used. Non-contingent pretreatment with methadone

reduced methadone self-administration. Subjects who required an

increasing number of responses (i.e., progressive ratio) for a 4 mg

unit dose of methadone, were more likely to detoxify than those

whose response requirements remained constant.

b. Self-Report of Drug Effects

The present instrument used to measure the subjective effects of

acutely administered opioids is the single dose opioid questionnaire

originally developed by Fraser et al. (1961). It consists of four

questions, the first of which simply determines whether the subject

feels the drug or not. Question two asks the subject to identify

the drug by comparing it with prototypic drugs of various classes.

The various physiological sensations are assessed in question three

while question four asks the subject to rate his degree of "liking"

of the drug. Concomitant with this assessment, observers blind to

the experiment rate the subject using similar questions.

The Morphine-Benzedrine Group Scale (or MBG) of the Addiction

Research Center Inventory (ARCI) is a multiple scale questionnaire

that has been used to identify compounds that produce opioid-like

effects (Jasinski et al. 1971). Morphine-like compounds. will

elevate the scores on the MBG scale in a dose-related fashion, so

that bioassay statistics (Finney 1964) can be used to determine

relative potencies between a standard (e.g., morphine) and a test

compound. With the development of opioids that possess mixed

agonist/antagonist effects, the other two questionnaires,

Pentobarbital, Chlorpromazine, Alcohol Group Scale (PCAG), and the

Lysergic Acid Diethylamide Scale (LSD) must be used in order to

characterize the full profile of behavioral effects of a test

compound (Jasinski 1977).

117
Such studies are usually conducted within a crossover design in

which each subject receives two or three doses of the standard drug

(e.g., morphine), an appropriate blank or placebo, and two or three

doses of the test drug. All drugs are given under double-blind

conditions. Double-dummy procedures are employed if the test drug

is given orally and is to be compared to parenterally administered

morphine. This entails giving the subject both an injection and a

liquid (or pill) to consume for each assessment. Thus, on placebo

days, the subject will receive a saline injection and drink a glass

of the liquid vehicle (usually orange-flavored Tang or equivalent).

Drugs are administered in random order with a minimum of 7 days

between sessions. Questionnaires are completed a number of times

after drug administration. The responses are quantified using a

standard scoring system, and dose response curves are constructed

using the mean responses for the standard test drug.

c. Behavioral Toxicity

Procedures for assessing the toxic effects of opioids on behavior

have been limited in scope. The two main studies in the literature

employed subjects currently enrolled in a methadone maintenance

program and compared them with matched non-dependent controls

(Gordon 1970; Rothenberg et al. 1977). Gordon (1970) assessed

single and choice reaction times to a visual stimulus. Subjects

were seated in front of a console that contained 6 neon lights in a

row on a display screen and 6 corresponding buttons on a panel

parallel to the table top. Lights were programmed to remain lit

until a response was made. Three basic conditions were studied:

simple reaction time, multiple-discrimination-multiple-response, and

multiple-discrimination-single-response. Waiting periods (from

completion of response to next stimulus presentation) were varied

randomly. The single reaction time required that the subjects

simply depress the button that corresponded to the illuminated

light. The multiple discrimination-multiple-response condition

required that the subject depress the buttons in the identical

sequence that the lights were illuminated. The multiplediscrimination-single-response condition required that the subject

simply select one of the buttons in the sequence of up to 6 stimuli.

Methadone-maintained subjects achieved the shortest reaction times;

this trend was consistent across the different experimental

conditions.

Rothenberg et al. (1977) also used a visual discrimination task

except that the environment was much more rigorously controlled than

in the previous study. Between 1 l/2 and 2 1/2 hours after their

morning dose of methadone, subjects were placed in the apparatus

(head restraint and chin support) in order to minimize head

movement-induced artifacts. Four different tests were administered

including letter recognition (3 different letters), continuous

performance (interval between test stimuli was 10 msec), reaction

time (the letter X was flashed on the screen and subject's time to

depress a microswitch was recorded), and reaction time with money

incentive (same conditions as previous test except that subjects

were paid for each response that was faster than a predetermined

value). Non-opioid-dependent subjects were similarly tested under

118
non-drug conditions and after receiving various doses of methadone.

Methadone-maintained subjects consistently performed faster than

controls, and money incentive reduced reaction times equally for both

groups. Acute doses of methadone produced dose-related increases in

reaction time.

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122
C. Procedures for Assessing CNS Depressants: Sedative/

Hypnotics, Anesthetics, Anxiolytics and Antihistamines.

1. Characterization of CNS Depressant Effects

a. Acute Drug Effects

i) Anxiolytic and Sedation

Objective measures of anxiety in human subjects are relatively

difficult to obtain. Muscle tension (which can be effectively

measured) is not a direct correlate of anxiety. Thus, almost all

procedures for assessing the effectiveness of these compounds in

reducing anxiety have employed verbal self-reports of a subject's

behavior (McReynolds 1968). One study by Kelly et al. (1969) has

attempted to incorporate concomitant measures of the clinical,

psychological, and physiological components of anxiety and the

effects of chlordiazepoxide on such measures. Chlordiazepoxide was

found to produce improvements in self-ratings and those of clinical

observers on anxiety and depression assessments, to decrease the

dizzy factor score on the Clyde Mood Scale, and to decrease resting

forearm blood flow.

While it is not clear whether the anxiolytic effect of these

compounds is distinctly different from their non-specific effects,

several procedures have been utilized to selectively parcel out the

sedative/hypnotic effects of these compounds as reflected in studies

evaluating sleep (Kay et al. 1976).

ii) Electroencephalographic (EEG) Activity and

Sleep

While procedures for assessing the direct effects of sedative/

hypnotics on human EEG are available, they have not, as yet, been

systematically applied to characterize these compounds. Most

studies have concentrated on subjective ratings of the relative

amounts of various EEG waveforms such as alpha, delta, theta, and

beta activity. In this regard Malpas et al. (1970) compared the EEG

effects of nitrazepam with those of amylobarbitone sodium. Using a

double-blind cross-over design, subjects received either 5 or 10 mg

of nitrazepam, 100 or 200 mg of amylobarbitone, or placebo.

Criteria for scoring EEG activity were established as follows.

0 - Epochs containing 50% or more alpha activity

1 - Epochs containing less than 50% of alpha but

no paroxysmal features of sleep

2 - Epochs containing one paroxysmal feature of sleep

3 - Epochs containing delta activity more than 50% of the

time.

The values were then tabulated as a function of time and then total

scores calculated to determine the sleep-awake profile. These

investigators also used subjective mood ratings and measured

psychomotor performance. Approximately 18 hours after a single

dose, mood and psychomotor performance had returned to normal, but

123
EEG changes were found to persist after the highest dose of

nitrazepam. These effects were indicative of a drowsy state.

Procedures for assessing the effects of these compounds on sleep

have been relatively well developed, refined, and validated in the

clinic. In these types of studies, drugs are assessed with respect

to three major criteria: the effectiveness of inducing and

maintaining sleep, development of tolerance to the hypnotic effects,

and effects upon individual sleep stages manifested by clinical

signs and symptoms. Thus, by employing quantified EEG procedures,

it is possible to replace verbal reports of how well a subject slept

with an objective measure of the sleep stages during the night.

Recently, Kay et al. (1976) reviewed the pharmacology of sleep.

Barbiturates, benzodiazepines, nonbarbiturates, and alcohol have all

been studied extensively using a variety of techniques for their

effects on sleep. Few studies, however, have attempted to provide

comparative information among several compounds. Hartmann (1968)

compared the effects of pentobarbital, chlordiazepoxide, Ro 5-6901,

and amitriptyline on sleep patterns in normal volunteers. All drugs

except amitriptyline increased total sleep. REM sleep was markedly

suppressed by amitriptyline and marginally suppressed by

pentobarbital while chlordiazepoxide and Ro 5-6901 did not affect

REM sleep.

2. CNS Depressant Physical Dependence Potential

a. Tolerance to Drug Effects

Procedures for determining the degree to which tolerance develops to

the effects of these compounds have not been extensively utilized.

Essentially, only the hypnotic effects of the barbiturates,

benzodiazepines, and miscellaneous compounds have been the subject

of tolerance assessment. These techniques are inherent in the

procedures described in the previous section on electroencephalographic activity and sleep.

Tolerance to the hypnotic effects of barbiturates occurs very

rapidly; in fact, blood levels upon awakening from a single dose are

higher than when the subject originally fell asleep (Harvey 1980).

The extent to which tolerance develops to the barbiturates depends,

in part, upon the duration of action and frequency of use. Wikler

(1976) demonstrated that the degree of tolerance to secobarbital or

pentobarbital could be minimized by enforcing a once-a-day treatment

with clinically acceptable hypnotic doses. When such controls over

the subjects' behavior is absent, they may experience less

satisfying sleep (due to the drug's effect on specific sleep stages)

and as a result, increase their dosage. This practice may be

responsible for the rapid development of tolerance.

Thus, while no systematic procedures are currently in use to compare

the degrees of tolerance to the sedative effects of these compounds,

a few points should be followed. First, the doses of the compounds

to be compared should be optimized to produce qualitatively and

quantitatively similar degrees of sedation. This may require using

the pharmacokinetic profile of each drug in preparing the treatment

124
regimen. Secondly, the duration of treatment should be comparable

to ensure similar degrees of exposure. And thirdly, objective

measures of drug effect (e.g., latency to sleep and various sleep

stages, duration of sleep, number of awakenings, etc.) should be

obtained on a regular basis during control and drug phases.

b. Withdrawal from Chronic Drug Administration - -

Procedures for assessing the dependence potential of sedative/

hypnotics in man have evolved over the past 30 years. The early

studies of Isbell et al. (1950, 1955) demonstrated that chronic

administration of intoxicating doses of barbiturates or alcohol

produced a pronounced abstinence syndrome when treatment was

discontinued. In addition, the withdrawal syndrome was

significantly different from that observed following withdrawal from

opiates.

While rating scales for opiate or opioid withdrawal are relatively

well established (Himmelsbach 1941), there is no such uniformity of

procedures for the evaluation of sedative/hypnotic dependence. The

first signs to appear after barbiturate withdrawal include:

anxiety, nervousness, tremors, weakness, anorexia, nausea, vomiting,

and insomnia. Acute weight loss also accompanies these- signs.

Unlike opiate withdrawal, the signs and symptoms of barbiturate

withdrawal may increase in intensity to the extent that grand ma1

convulsions, hallucinations (both auditory and visual), and frank

delirium occur. Hyperpyrexia followed by death has been reported

(Fraser et al. 1953).

The first use of the EEG as a measure of barbiturate withdrawal was

reported by Fraser et al. (1954, 1958). Abrupt termination of a

dose of 0.2 g of pentobarbital per day for as long as six months was

followed by no significant effects. About one third of the subjects

that received 0.4 g for 3 months experienced paroxysmal EEG changes

(e.g., random spikes, fast waves, slow waves) in lieu of other

significant clinical signs of withdrawal. Minor signs of withdrawal

and about 50% abnormal EEG's were characteristic of abrupt

withdrawal from pentobarbital at a dose of 0.6 g for 1 to 2 months.

Seizures, delirium, and all minor abstinence signs were evident in

subjects upon withdrawal from high doses (0.9 to 2.2. g/day X

several months) of pentobarbital.

Benzodiazepines have, for the most part, replaced barbiturates as

the treatment of choice for sedation and hypnosis. Consequently,

procedures for assessing their dependence potential have been

reported. Hollister et al. (1961, 1963) demonstrated that the

termination of high doses of chlordiazepoxide or diazepam in

schizophrenics resulted in signs of withdrawal that resembled the

barbiturate withdrawal syndrome. Studies demonstrating the relative

dependence potential of various barbiturates have been reviewed by

Woods et al. (1983). The lack of uniformity of experimental

procedures between the various studies has made it impossible to

distinguish between the various benzodiazepines, if indeed such a

distinction exists.

125
3. CNS Depressant Abuse Liability

a. Drug Self-Administration

Procedures for measuring the self-administration of sedative/

hypnotics by human subjects have been validated by experimental

manipulation of the conditions surrounding drug availability

including such factors as amount of work required to obtain drug and

unit dose size (Bigelow et al. 1976a, 1976b; Pickens et al. 1977)

and choice between different sedatives (Bigelow et al. 1976b;

Griffiths et al. 1979, 1980).

One study by Johanson and Uhlenhuth (1978) used a paired preference

procedure to assess the relative reinforcing efficacy of diazepam as

compared to saline in normal subjects with no history of sedative

abuse. Subjects did not choose any dose of diazepam over saline but

did choose amphetamine over diazepam.

While some studies have assessed self-administration of these

compounds in a treatment facility where patients merely ask for

medication (Rothstein et al. 1976; Kryspin-Exner and Demal 1975),

more recent reports were obtained in research settings employing

more rigorous control over the drug delivery process. These studies

typically use token economies (Bigelow et al. 1976a) in which

subjects are required to perform work for a specific length of time

in order to earn a specific number of tokens. These tokens can then

be exchanged for drug. Surplus tokens are not carried over to the

next day, but rather are cashed in for money to be paid to the

subject upon discharge from the unit.

Most studies have been conducted with subjects that have current or

past histories of sedative abuse. Bigelow et al. (1976b) studied

the effects of varying the number of tokens required to purchase a

single 30 mg dose of pentobarbital or a 10 mg dose of diazepam.

Subjects were required to ride a stationary exercise bicycle for two

minutes for each token. The number of tokens required to purchase

the dose (drugs were available for 7hr/day) was randomly varied

among 1, 3, 5, 8, and 10. Subjects generally worked for and

subsequently consumed all doses of both drugs when the number of

tokens required for purchase was 1 or 3. As the number of tokens

for each dose was increased, however, the amount of drug earned

correspondingly decreased. The results of this study provide a

basic framework upon which to build a more complete program for

assessing sedative self-administration by humans. In subsequent

studies (Griffiths et al. 1979, 1980), it has been shown that

subjects will generally prefer pentobarbital to diazepam over a

fairly wide dose range.

b. Self-Report of Drug Effects

Subjective rating scales remain the primary means of assessing the

effects of psychoactive compounds on mood and feeling states (Fraser

and Jasinski 1977). The PCAG scale scores (Haertzen 1966) are

particularly sensitive to feelings of lethargy, weakness, and loss of

energy that characterize sedative/hypnotics. In order to obtain an

126
overall profile of drug effect, however, liking scores and the MBG

scale scores are also calculated. Using this method, Fraser and

Jasinksi (1977) demonstrated that pentobarbital and secobarbital

produced dose-related increases in the above scale scores.

Phenobarbital, while producing some increases in scores, was only

one-fifth to one-seventh as potent as pentobarbital and

secobarbital, and did not appreciably elevate MBG scale scores.

In a subsequent study, Jasinski et al (1981) compared the effects of

various doses of pentobarbital with diazepam and chlordiazepoxide.

In a double-blind crossover design, both diazepam and

chlordiazepoxide were found to produce pentobarbital-like effects

and euphoria. Diazepam was about ten times more potent than

pentobarbital.

Two other procedures that have been used to characterize the mood

effects are based upon a Mood Adjective Check List (MACL) for

self-report of mood originally introduced by Nowlis and Nowlis

(1956), and Magnitude Estimation (ME) introduced by Ekman (1967).

Svensson et al. (1980) compared the efficiency of these two measures

in evaluating the mood effects of diazepam and caffeine. The result

of both scales indicated that diazepam decreased feelings of

activation and extroversion and increased calmness. While the ME

scale provided slightly better validity, it was harder for the

subjects to complete.

c. Behavioral Toxicity

Numerous procedures for assessing the toxic effects of sedative/

hypnotics on behavior have been developed, refined and validated.

McNair (1973) reviewed the results of 101 studies that assessed the

effects of antianxiety drugs on human performance. He grouped the

43 types of tests into four categories from high to low sensitivity.

Among the highest were digit-symbol substitution test (DSST),

driving accuracy, visual thresholds, and auditory flutter fusion.

Among the low sensitivity tests were problem solving, visual and

auditory vigilance, arithmetic, and learning nonsense syllables.

Studies that compare the psychomotor performance effects of various

benzodiazepines and barbiturates have been reviewed recently by

Johnson and Chernik (1982) with an emphasis on procedures designed

to assess the "day-after" or hangover effect. Card sorting, tapping

rate, symbol copying, and DSST were found to be among the most

sensitive tests for assessing behavioral impairment after

benzodiazepine administration. These effects were dose related, but

inconsistencies with respect to plasma half-lives and duration of

impairment were found (Johnson and Chernik 1982).

References

Bigelow, G.; Griffiths, R.R.; and Liebson, I. Experimental human

drug self-administration: Methodology and application to the

study of sedative abuse. Pharmacol Rev, 27:523-531, 1976a.

127
Bigelow, G.E.; Griffiths, R.R.; and Liebson, I. Effects of response

requirement upon human sedative self-administration and drugseeking behavior. Pharmacol Biochem Behav, 5:681-685, 1976b.

Ekman, G. The measurement of subjective reactions.

Forsvarsmedicin, 3:27-41, 1967.

Fraser, H.F.; Isbell, H.; Eisenman, A.J.; Wikler, A.; and Pescor,

F.T. Chronic barbiturate intoxication. Further Studies. Arch

Intern Med, 94:34-41, 1954.

Fraser, H.F., and Jasinski, D.R. The assessment of the abuse

potentiality of sedative/hypnotics (depressants) (methods used

in animals and man). In: Martin, W.R., ed. Drug Addiction I.

Berlin: Springer-Verlag, 1977. pp. 589-612.

Fraser, H.F.; Shaver, M.R.; Maxwell, E.S.; and Isbell, H. Death due

to withdrawal of barbiturates. Report of a case. Ann Intern

Med, 38:1319-1325, 1953.

Fraser, H.R.; Wikler, A.; Essig, C.F.; and Isbell, H. Degree of

physical dependence induced by secobarbital and pentobarbital.

J Amer Med Assoc, 166:126-129, 1958.

Griffiths, R.R.; Bigelow, G.; and Liebson, I. Human drug

self-administration: Double-blind comparison of pentobarbital.

diazepam, chlorpromazine and placebo. J Pharmacol Exp Ther,

210:301-310, 1979.

Griffiths, R.R.; Bigelow, G.E.; Liebson, I.; and Kaliszak, J.E.

Drug preference in humans: Double-blind choice comparison of

pentobarbital, diazepam and placebo. J Pharmacol Exp Ther,

215: 649-661, 1980.

Haertzen, C.A. Development of scales based on patterns of drug

effects. using the Addiction Research Center Inventory (ARCI).

Psychol-Rep, 18:163-194, 1966.

Hartman. E. The effect of four drugs on sleep patterns in man.

Psychopharmacology, 12:346-353, 1968.

Harvey, S.C. Hypnotics and sedatives, In: Gilman, A.G.. Goodman,

L.S., and Gilman, A., eds. The Pharmacological Basis of

Therapeutic. 6th Edition. New York: McMillan, 1980. pp.

Himmelsbach, C.K. The morphine abstinence syndrome, its nature and

treatment. Ann Intern Med. 15:829-839, 1941.

Hollister, L.E.; Benneth, J.L.; Kimbell, I.; Savage, C.; and

Overall, J.E. Diazepam in newly admitted schizophrenics. Dis

Nerv Syst, 24:746-750, 1963.

Holliz, L.E.; Motzenbecker, F.P.; and Degan, R.O. Withdrawal

reactions from chlordiazepoxide (Librium).

Psychopharmacologia, 2:63-68, 1961.

Isbell H.; Altschul, S.;

H:G.;

Kornetsky, C.H.; Eisenman, A.J.; Flanary,

Fraser, H.F. Chronic barbiturate intoxication. An

experimental study. Arch Neurol Psychiat, 64:1-28, 1950.

Isbell, H.; Fraser, H.F.; Wikler, A.; Belleville, R.E.; and

Eisenman, A.J. An experimental study of the etiology of "rum

fits" and delerium tremens. Quart J Stud Alcohol, 16:1-33,

1955.

Jasinski, D.R.; Haertzen, C.A.; Henningfield, J.E.; Johnson, R.E.;

Makhzoumi. H.M.; and Miyasato, K. Progress report of the NIDA

Addiction Research Center. In: Problems of Drug Dependence.

National Institute on Drug Abuse Research Monograph 41. The

Committee on Problems of Drug Dependence, 1981. pp. 45-52.

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Johanson, C.E., and Uhlenhuth, E.H. Drug self-administration in

humans. In: Krasnegor, N., ed. Self-Administration of Abused

Substances: Methods for Study National Institute on Drug

Abuse Research Monograph 20, Washington, D.C.: U.S. Govt.

Print. Office, 1978. pp. 68-65.

Johnson, L.C., and Chernick. D.A. Sedative-hypnotics and human

performance. Psychopharmacology, 76:101-113, 1982.

Kay, D.C.; Blackburn, A.B.; Buckingham, J.A.; and Karacan, I. Human

pharmacology of sleep. In: Williams, R.L., and Karacan, I.,

eds. Pharmacology of Sleep. New York: John Wiley & Sons,

1976. pp. 83-210.

Kelly, D.; Brown, C.C.; and Shafter, J.W. A controlled

physiological, clinical and pychological evaluation of

chlordiazepoxide. Brit J Ps chiat, 115:1387-1392, 1969.

Kryspin-Exner, K., and e Th use of tranquilizers in the

treatment of mixed drug abuse. Int J Clin Pharmaol, 12:13-18,

1975.

Malpas, A.; Rowan, A.J.; Joyce, C.R.B.; and Scott, D.F. Persistent

behavioral and electroencephalographic changes after single

doses of nitrazepam and amylobarbitone sodium., Brit Med J, 2:

762-764, 1970.

McNair, D.M. Antianxiety drugs and human performance. Arch Gen

Psychiat 29:611-617, 1973.

McReynolds, P., ed. The assessment of anxiety. A survey of

available -techniques. In: Advances in Psychological

Assessment. Chapter XIII. Palo Alto, California: Science and

Behavior Books, 1968.

Nowlis, V., and Nowlis, H.H. The descriptions and analysis of mood.

Ann NY Acad Sci, 65:345-355, 1956.

Pickens, R.; Cunningham, M.R.; Heston, L.L.; Eckert, E.; and

Gustafson, L.K. Dose preference during pentobarbital

self-administration by humans. J Pharmacol Exp Ther, 203:

310-318, 1977.

Rothstein, E.; Cobble, J.C.; and Sampson, N. Chlordiazepoxide:

Long-term use in alcoholism. Ann NY Acad Sci, 273:381-384,

1976.

Svensson, E.; Persson, L.O.; and Sjoberg, L. Mood effects of

diazepam and caffeine. Psychopharmacoloqy, 67:73-80, 1980.

Wikler, A. Review of research on sedative drug dependence at the

Addiction Research Center and University of Kentucky. In:

Thompson, T., and Unna, K., eds. Predicting Dependence

Liability of Stimulant and Depressant Drugs. Baltimore,

Maryland: University Park Press, 1976. pp. 147-163.

Woods, J.H.; Katz, J.L.; and Winger, G. Abuse and dependence

liabilities of benzodiazepines. Pharmacol Rev, 1983

(submitted).

129
D. Procedures for Assessing CNS Stimulants:Anorectics, Local

Anesthetics, and Antidepressants

1. Characterization of CNS Stimulant Effects

a. Acute Drug Effects

i) Electroencephalographic (EEG) Activity and

Sleep

The direct effects of CNS stimulants on EEG activity have been shown

to be alerting or activating as determined by the qualitative

techniques of visual inspection. The desynchronized pattern of EEG

activity, characterized by a slight decrease in amplitude and an

increase in predominant frequency, has been quantified by the use of

voltage integration procedures (Goldstein et al. 1963). Although

the use of power spectral analysis has not been employed extensively

to assess the EEG effects of this class of compounds, a report by

Gibbs and Maltby (1943) did describe a shift in the spectrum to

higher frequencies following benzedrine.

ii) Appetite

Procedures for assessing the effects of stimulants on appetite have

been developed for short-term, single dose experiments. Hoebel et

al. (1975) utilized a procedure that tests a drug's effectiveness in

reducing intake of a diet liquid meal product. Volunteers report to

the laboratory having refrained from eating for 2 hours prior to

lunchtime (12 noon). Using a double-blind cross-over design,

subjects were given either placebo or 25 mg of phenylpropanolamine

30 minutes before lunch. Lunch consisted of a canned chocolateflavored drink which was dispensed via a long straw from a graduated

cylinder. The reservoir was hidden from the subject's view.

Subjects were instructed to drink as much as they wished.. Upon

completion of lunch they filled out questionnaires that related to

the taste of the lunch, the reason for stopping, the amount that

they consumed relative to the previous day, and the taste of the

lunch relative to the previous day.

2. CNS Stimulant Physical Dependence Potential

a. Tolerance to Drug Effects

The assessment of tolerance to the effects of the CNS stimulants has

been directed mainly at two of their pharmacologic effects:

anorexia and mood elevation. Tolerance to the appetite-suppressant

effects has been demonstrated to occur in numerous studies. It

appears; however, that obesity has many complex components, at least

one of which, the 'craving" for food, may interact with the

evaluation of tolerance effects (Wooley and Wooley 1981).

The currently accepted criterion for the appetite suppressant effect

is that weight is lost and not recovered. The fact that the rate of

weight loss rapidly decreases over the course of treatment is taken

as evidence of tolerance. Stunkard (1979) has suggested that these

130
criteria are not valid for assessing tolerance, since a deceleration

in weight loss is common even in more drastic treatments of obesity

such as jejunoileal bypass surgery. Additionally, the metabolic

characteristics of the body change as a result of weight loss, and

these changes should be taken into account when assessing tolerance.

Food that has been ingested during a period of weight loss is more

efficiently metabolized than when the caloric balance is in steady

state (Keesey et al. 1976). A more appropriate determinant of

whether tolerance has developed or not may be the degree of weight

gain following cessation of drug treatment. Tolerance would be

shown by a return to normal weight despite continued administration

of the drug. Rebound hyperphagia and weight gain, upon withdrawal

of the drug, would constitute a withdrawal reaction.

Tolerance to the mood elevating effects of CNS stimulants has been

assessed and, in addition,

effects (Gunne 1977).

is distinct, from the appetite-suppressant

Tolerance to the euphoric effects of

amphetamine (as measured by a subjective questionnaire) was found to

develop rather quickly over 14 days of daily treatment (Rosenberg et

al. 1963). In addition, these authors demonstrated that there was

no cross-tolerance to LSD. To date, no studies have been conducted

to directly compare the degree of tolerance development to the

subjective effects of CNS stimulants.

b. Withdrawal from Chronic Drug Administration

Only limited data are available on withdrawal from chronic CNS

stimulant use, since experimental studies have rarely been

undertaken, and virtually all reported observations have been made

within the context of treatment programs. In addition., the

difficulty in obtaining valid patient histories complicates

determination of the exact profile of the CNS stimulant withdrawal

syndrome. Amphetamine withdrawal has been characterized, however,

by assessing affective states, sleep patterns, and MHPG excretion

(Watson et al. 1972). Suprisingly. patients do riot seem to "crave"

for amphetamine during this abstinence phase as patients withdrawing

from opiates do for morphine.

3. CNS Stimulant Abuse Liability

a. Drug Self-Administration

Procedures for assessing self-administration of' i.v. cocaine have

been reported by Fischman and Schuster (1982) with experienced

cocaine users prepared with two indwelling Angiocath Teflon

catheters, one for drug infusion and one for blood withdrawal.

Subjects were instructed that each of two buttons would be

associated with the same solution (drug or saline) throughout the

study. On the first day, subjects were exposed to each solution and

for the next 8 days were required to press the response button 10

times in order to receive an injection. Once a single response was

made on a given button, the other one was deactivated. A total of

10 injections could be taken during a l-hour session with blood

drawn prior to and after each injection followed by completion of

the POMS and ARC1 questionnaires. After initial sampling on the

131
first day, subjects consistently chose cocaine over saline.

Subjective ratings of drug effect correlated with the self-injection

of cocaine. The most pronounced effects on subjective states, blood

pressure, heart rate, and cocaine levels occurred after this initial

injection.

b. Self-Report of Drug Effects

Studies assessing the subjective effects of CNS stimulants have been

conducted using subjects with drug abuse histories (Martin et al.

1971; Jasinski et al. 1974; Fischman et al. 1976) and normal

volunteers with no history of drug abuse (Johanson and Uhlenhuth

1980). In the former studies, subjects lived on the ward during

their participation in the study. Martin et al. (1971) compared the

subjective effects of several sympathomimetic amines with

physiologic and behavioral effects. Amphetamine, methamphetamine,

ephedrine, phenmetrazine, and methylphenidate were given under

double-blind conditions using a Latin square design. Subjects

completed questionnaires every hour for 6 hours after drug

administration. These scales consisted of the PCAG, MBG, BG, and a

new amphetamine (A) scale which was constructed from significant

items in the other scales. All compounds produced dose-related

increases in MBG, BG. and A scale, as well as subject and observer

liking. PCAG scale scores were also decreased, while the highest

dose of all drugs increased LSD scale scores. These subjective

responses correlated well with physiologic effects and allowed for

valid estimates of relative potency measures.

Johanson and Uhlenhuth (1980) employed a procedure in which the

subjects reported to the laboratory in the morning 3 times a week

for each session. Subjects filled out a POMS form prior to each

amphetamine or placebo ingestion and then again at 1, 3, and 6 hours

post drug. The authors have used this procedure as a basis upon

which to build a choice procedure for different drugs. Subjects

associated certain colored capsules with drug effects, and during the

latter part of the experiment they were allowed to request the

capsule they preferred.

c. Behavioral Toxicity

Studies assessing the effects of CNS stimulants on psychomotor

performance have used a number of strategies to detect increases or

decreases in speed, error rate, and efficiency. In general, most

CNS stimulants have been shown to enhance performance.

Early studies such as that by Seashore and Ivy (1953) and Somerville

(1946) studied the effects of caffeine, amphetamine. and placebo on

perception and motor skills in fatigued army soldiers. Performance

was generally enhanced, but the differences observed between drug

and placebo condition were small, probably because they used trained

individuals that were already performing at peak levels. More

recently Evans et al. (1976) studied the effects of various doses of

dextroamphetamine on quantitative measures of psychomotor

performance. Non-fatigued volunteers refrained from using any drugs

for 18 hours prior to testing. Three measures of psychomotor

132
performance were used: the wobble board (Shipley and Harley 1971),

the pursuit meter (Evans et al. 1973), and delayed auditory feedback

(Hughes et al. 1963). There was some improvement in stability on

the wobble board during the eyes closed condition as well as slight

improvements in performance on the pursuit meter (but only at the 7

cycles/sec mode and not at 5 or 2 cycles/sec) that were dose

related. No change in delayed auditory feedback performance was

observed, however.

References

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Effects of dextroamphetamine on psychomotor skills. Clin

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Fischman, M.W., and Schuster, C.R. Cocaine self-administration in

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

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Goldstein, L.; Murphree, H.B.; and Pfeiffer, C.C. Quantitative

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Gunne, L.M. Effects of amphetamines in humans. In: Martin, W.R.,

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Hughes, F.W.; Forney, R.B.; and Gates, P.W. Performance in human

subjects on delayed auditory feedback after alcohol, a

tranquilizer (benzquinamide)

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Jasinski, D.R.; Nutt, J.G.; and Griffiths, J.D. Effects of

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administration. Clin Pharmacol Ther, 16:645-652, 1974.

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humans: $-amphetamine. Psychopharmacology, 71:275-279, 1980.

Keesey, R.E. The role of lateral hypothalamus in determining the

body weight set point. In: Novin, D., Wyrwicka, W., and Bray,

G.A., eds. Hunger: Basic Mechanisms and Clinical

Implications. New York: Raven, 1976. pp. 243-255.

Martin, W.R.; Sloan, J.W.; Sapira, J.D.; and Jasinski. D.R.

Physiologic, subjective, and behavioral effects of amphetamine,

methamphetamine, ephedrine, phenmetrazine, and methylphenidate

in man. Clin Pharmacol Ther, 12:245-258, 1971.

133
Rosenberg, D.E.; Wolbach, A.B.; Miner, E.J.; and Isbell, H.

Observations on direct and cross tolerance with LSD and

d-amphetamine in man. Psychopharmacologia, 5:1-15, 1963.

Seashore, R.H., and Ivy, A.C.

relieving fatigue.

Effects of analeptic drugs in

Psychol Monogr, 67:1-16, 1953.

Shipley, R.E., and Harley, R. J. A device for estimating stability

of stance in human subjects.

1971.

Psychopharmacology, 7:287-292,

Somerville, W. The effect of benzedrine on mental or physical

fatigue in soldiers.

Stunkard, A.J.

Can Med Assoc J, 55:470-476, 1946.

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The development of tolerance to appetite suppressant

New theory, new hope. In: Smith, D.E., ed.

Amphetamine Use, Misuse and Abuse. Boston: G.K. Hall and Co.,

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Watson, R.; Hartmann, E.; and Schildkraut, J.J.

withdrawal:

Amphetamine

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Affective state, sleep patterns, and MHPG

Amer J Psychiat, 129:263-269, 1972.

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134
E. Procedures for Assessing Cannabinoids

1. Characterization of Cannabinoid Effects

a. Acute Drug Effects

i) Analgesia

Clinical data on analgesic activity have generally been derived from

studies assessing the effects of delta-9-THC or smoked marijuana on

pain which have not yielded clear results. It appears, however,

that delta-9-THC (5-20 mg) is effective in relieving the pain

associated with' advanced stages of cancer (Noyes et al. 1976).

While higher doses were effective in relieving pain, they were

accompanied by significant side effects, including sedation and

verbal impairment. In contrast, Hill et al. (1974) using

experimental electrical stimulation were unable to detect analgesic

activity after 12 mg of delta-9-THC.

While several of the newer synthetic cannabinoids are currently

undergoing clinical evaluations as analgesics, it is clear that the

most difficult problems associated with such assessments continue to

involve the need for parceling out confounding side effects such as

sedation and impaired verbal performances.

ii) Electroencephalographic (EEG) Activity and

Sleep

Studies designed to assess the EEG effects of acute administration

of cannabinoids have been conducted almost exclusively with

delta-9-THC or marijuana via the oral or inhalation (smoked) route.

Most of the past work was performed prior to the widespread use of

computerized, power spectral analysis. In general, orally

administered marijuana or THC has been reported to increase alpha

activity and EEG synchronization (Hollister et al. 1970;

Deliyannakis et al. 1970; Volavka et al. 1971, 1973; Tassinari et

al. 1974). More recently, however, a study by Koukkou and Lehmann

(19.76) utilizing spectral techniques to more accurately quantify the

EEG effects of 200 mg/kg delta-9-THC showed that the predominant

alpha frequency was decreased by THC. but the magnitude of change

was related to the resting state and EEG baseline of each subject.

The effects of cannabis on sleep have been assessed using all-night

EEG recordings in a laboratory setting (Barrett et al. 1974).

Slow-wave sleep increases during the first four nights after smoking

marijuana, though the amount of slow-wave sleep progressively

decreased beyond that point until it was below normal levels. The

suppression of slow-wave sleep persisted well into the post-drug

period. These effects seem to be dose related in that higher doses

actually overstimulate the subjects such that they sleep less.

Pivik et al. (1972) compared the effects of delta-9-THC and synhexl

on sleep patterns in normal subjects. Both compounds selectively

increased stage 4 sleep while rapid-eye-movement or REM sleep was

depressed. When a sleep deprivation procedure was employed,

135
however, subjects recovered from these effects much more rapidly

when treated with synhexl than with delta-9-THC.

2. Cannabinoid Physical Dependence Potential

a. Tolerance to Drug Effects

Tolerance to the subjective and physiological effects of

cannabis as compared to synhexyl was demonstrated by Williams et al.

(1946). In this study measures were collected daily over a 39-day

period, including rectal temperature, pulse, respiratory rate, blood

pressure, body weight, caloric intake, EEG, sleep, and mood states.

Pulse, caloric intake, and expressions of euphoria were all

initially increased, but decreased after a few days of marijuana. A

similar profile was noted for synhexyl. More recently, Fink et al.

(1976) demonstrated tolerance to marijuana produced increases in

pulse rate and alpha activity of the EEG as well as to the

decremental effects of the drug upon short-term memory and reaction

time. After low dose chronic administration of delta-9-THC, it is

difficult to observe tolerance to subjective effects even though

tolerance to the drug-induced tachycardia and dizziness is readily

demonstrated (Hollister and Tinkleberg 1973). Such tolerance

effects are clearly dose related, however, since they are most

apparent after prolonged exposure to high doses.

b. Withdrawal from Chronic Drug Administration - -

Only limited data are available on procedures for assessing the

signs and symptoms attendant upon cessation of chronic cannabinoid

administration. Jones and Benowitz (1976), for example, gave

subjects an oral 30 mg dose of delta-9-THC every 4 hours for 10 to

20 days. Autonomic signs of withdrawal included increased sweating,

salivation, and tremors of the extremities. Milder signs included

irritability, sleep disturbances, and anorexia, while nausea,

vomiting, and diarrhea occurred in some subjects.

3. Cannabinoid Abuse Liability

a. Drug Self-Administration

Procedures for studying self-administration of cannabinoids have

been established using basic operant procedures that employ

marijuana cigarettes as the reinforcer (Mendelson et al. 1974,

1976). Subjects lived on a research ward during all three phases of

the study: 5-day baseline, El-day marijuana smoking, 5-day postmarijuana. During their stay, subjects earned points by pressing a

button on a hand-held portable manipulandum. Subjects could then

"buy" a marijuana cigarette for 1800 points (the equivalent of 30

minutes of sustained button pressing) or save the points until the

end of the study and exchange them for money (about 8.50 per 30

minutes). Once a subject decided to buy a marijuana cigarette, the

points were immediately deducted from his running total and the

cigarette was smoked at that time under staff supervision. Both

"heavy" and "casual" users were studied. All subjects avidly worked

for more points than were necessary to buy the number of cigarettes

136
they used, and as a result they earned a substantial amount of money

as well. The procedure thus serves as' a basis for evaluation of

other constituents of cannabis with respect to the maintenance of

operant performance.

b. Self-Report of Drug Effects

Using the standardized ARCI, Isbell et al. (1967) demonstrated that

subjects could not distinguish between marijuana and delta-9-THC.

Jones and Benowitz (1976) administered the Profile of Mood States

(POMS) questionnaire to subjects participating in a 30-day chronic

marijuana study. Tolerance to the subjective effects (e.g.,

confusion, inertia, and vigor) of 210 mg/day of delta-9-THC was

clearly evident within 3 days. In addition, using the Nurses'

Observation Scale for Inpatient Evaluation (NOSIE-3O), tolerance to

the decrease in social competence was also apparent by the 6th day.

Hollister and colleagues have systematically evaluated many of the

constituents of marijuana, THC metabolites, and synthetic analogues

of THC. In a summary of this work, Hollister (1974) showed that

delta-8-THC differed from delta-9-THC in a quantitative fashion.

Differences in relative potency were obtained with substitutions on

the alkyl side chain. The metabolites 11-hydroxy-delta-9- and

11-hydroxy-delta-8-THC were both more potent than their respective

parent compounds.

c. Behavioral Toxicity

Procedures for assessing the toxic consequences of cannabinoid

administration have been developed for measures of both- verbal

performances (e.g., "attention," "memory") and sensory-motor

functions (e.g., reaction time, stimulus discrimination). Dornbush

and Kokkevi (1976) assessed the acute effects of various doses of

delta-9-THC or hashish (compared to placebo) upon digit spans,

serial sevens, star tracing, and time estimation. The simple task

(e.g., digit span) was unaffected by cannabinoid administration

while performance on the more complicated tasks was impaired.

Numerous models of human memory assessment have been described

(Delong and Levy 1974; Darley and Tinklenberg 1974) in which

cannabinoids were shown to interfere with attention and decrease

memory efficiency. Vachon and Sulkowski (1976) attempted to dissect

the effects of marijuana on attention, memory, and psychomotor speed

by means of numerous automated testing devices including the

Continuous Performance Test (CPT) developed by Rosvold et al. (1956)

and modified by Mirsky and Kornetsky (1964), and the Automated Digit

Symbol Substitution Test (ADSST). A single dose of delta-9-THC (25

mg in a smoked cigarette) significantly decreased performance scores

as a function of an increase in complex response times and the

emmission of fewer responses.

Finally, the assessment of motor performance using an aviation

instrument flight simulator involved subjects (licensed pilots)

instructed to "fly" through each of four prespecified (and

practiced) holding patterns (Blaine et al. 1976). Using

137
double-blind conditions, marijuana (0.09 mg delta-9-THC per kg) or

placebo cigarettes were smoked during a lo-minute interval. Thirty

minutes later the subjects began the simulations. There was a

significant increase in the number of both major and minor errors,

and an increased average deviation from the assigned flight

sequence. This decrement in performance persisted for at least 2

hours after smoking and returned to control levels by 6 hours post

drug.

References

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on human sleep patterns. Biol Psychiat, 8:47-54, 1974.

Blaine, J.D.; Meacham, M.P.; Janowsky, D.S.; Schoor, M.; and

Bozzetti, L.P. Marihuana smoking and simulated flying

performance. In: Braude, M.C., and Szara, S., ed. The

Pharmacology of Marihuana. New York: Raven, 1976.

445-447.

pp.

Darley, C.R., and Tinklenberg, J.R. Marihuana and memory. In:

Miller, L.L., ed. Marijuana: Effects on Human Behavior. New

York: Academic Press, 1974. pp. 73-102.

Deliyannakis, E.; Panagopoulos, C.; and Huott, A.D. The influence

of hashish on the human EEG.

1:128-140, 1970.

Clin Electroencephalog,

Delong, F.L., and Levy, B.I. A model of attention describing the

cognitive effects of marijuana. In: Miller, L.L., ed.

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Press, 1974.

Dornbush, R.L., and Kokkevi, A. The acute effects of various

cannabis substances on cognitive, perceptual, and motor

performance in very long-term hashish users. In: Braude,

M.C., and Szara, S., eds. The Pharmacology of Marihuana. New

York: Raven, 1976. pp. 421-428.

Fink, M.; Volavka, J.; Panayiotopoulos, C.P.; and Stefanis, C.

Quantitative EEG studies of marijuana, delta-gtetrahydrocannabinol. and hashish in man. In: Braude, M.C.,

and Szara, S., eds. The Pharmacology of Marihuana. New York:

Raven, 1976. pp. 383-391.

Hill, S.Y.; Schwin, R.; Goodwin, D.W.; and Powell, B.J. Marihuana

and pain. J Pharmacol Exp Ther, 188:415-418, 1974.

Hollister, L.E. Structure-activity relationships in man of cannabis

constituents and homologs and metabolites of delta-gtetrahydrocannabinol. Pharmacology, 11:3-11, 1974.

Hollister, L.; Sherwood, S.; and Cavasino, A. Marijuana and the

human electroencephalogram. Pharmacol Res Commun, 2:305-308,

1970.

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Jones, R., and Benowitz, N. The 30-day trip: Clinical studies of

cannabis tolerance and dependence. In: Braude, M.C., and

Szara, S., eds. The Pharmacology of Marihuana. New York:

Raven, 1976. pp. 627-645.

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Koukkou, M., and Lehmann, D.: Human EEG spectra before and during

cannabis hallucinations. Biol Psychiat, 11:663-677, 1976

Mendelson, J.H.; Kuehnle, J.C.; Greenberg, I.; and Mello , N.K.

Operant acquisition of marihuana in man. J Pharmacol Exp Ther,

198:42-53, 1976.

Mendelson, J.H.; Rossi, A.M.; and Meyer, R.E., eds. The Use of

Marihuana: A Psychological and Physiological Inquiry. New

York Plenum, 1974.

Mirsky A.F., and Kornetsky, C. On dissimilar effects of drugs on

the digit symbol substitution and continuous performance tests.

Psychopharmacologia, 5:161-177, 1964.

Noyes. R.; Brunk, S.; Baram, D.; and Canter, A. Analgesic effects

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The Pharmacology of Marihuana. New York: Raven,

Pivik, R.T.; Zarcone, V.; Dement, W.C.; and Hollister,. L.E.:

pp. 833-836.

Delta-9-tetrahydrocannabinol and synhexl: Effects on human

sleep patterns: Clin Pharmacol Ther, 13:426-435, 1972.

Rosvold, H.E.; Mirsky, A.F.; Sarason, I.; Bransome, E.D.; and Beck,

Consult Clin Psychol 20:343-350

A continuous performance test of brain damage. J

1956.

Tassinari, C.A.; Peraita-Adrados, M.R.; Ambrosetto, G.; and Gastaut,

A. Effects of marijuana and delta-9-THC at high doses in man.

Electroenceph Clin Neurophys, 36:94, 1974.

Vachon, L., and Sulkowski. A. Attention, learning, and speed in

psychomotor performances after marihuana smoking. In: Braude.

M.C., and Szara, S., eds. The Pharmacology of Marihuana.

New York: Raven. 1976.

Volavka, J.; Crown, P.; Dornbush, R.; Feldstein, S.; and Fink, M.

EEG, heart rate, and mood change ("high") after cannabis.

Psychopharmacologia, 32:11-25, 1973.

Volavka, J.; Dornbush, R.; Feldstein, S.; Clare, G.; Zaks, A.; Fink,

M.; and Freedman, A.M. Marijuana, EEG and behavior.

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Lloyd, B.J. Studies in marihuana and pyrahexyl compound. Pub

Health Rep, 61:1059-1083, 1946.

139
F. Procedures for Assessing Hallucinogens

1. Characterization of Hallucinogen Effects

a. Acute Drug Effects

i) Electroencephalographic (EEG) Activity

In general, LSD has been shown to increase the alpha frequency in

patients with sensory defects (Korein and Musacchio 1968) and normal

individuals (Shagass 1967). Reduction in the overall amplitude of

the EEG has also been reported after LSD and mescaline (Monroe et

al. 1957) and LSD-25 (Rodin and Luby 1966; Shagass 1967), while both

frequency and power shifts have been quantified using spectral

analysis after ditran and LSD administration (Itil and Fink 1966).

In a study using integrative techniques, Goldstein et al. (1963)

demonstrated that a 1 microgram/kg dose of LSD decreased

electrogenesis of the EEG of normal male volunteers, but not in

chronic schizophrenic patients. The successive waves of the EEG

were full-wave rectified and an oscillator was used to generate a

train of pulses the frequency of which was directly proportional to

the area under the rectified EEG. Thus, pulses per unit time were

used to quantify the amount of energy contained in the corresponding

epoch of EEG activity. Recordings from the occipital area were

analyzed because of alpha rhythm prominence. After 10 minutes of

control recording, the subjects were allowed to rest for 20 minutes.

This cycle was repeated to obtain five lo-minute sessions at 10, 30,

60, 90, and 120 minutes. Drug or placebo was administered orally

after the first control session.

Doses of LSD (0.001-0.002 mg/kg) were administered intravenously

over a period of 5 minutes. Twenty-to 60-second epochs of EEG

activity were sampled and subjected to computer analysis. Samples

were taken before injection and 15 to 25 and 40 to 50 minutes

post injection. LSD caused an increase in all frequencies, a

decrease of slow activity, and an increase in beta activity.

2. Hallucinogen Physical Dependence Potential

a. Tolerance to Drug Effects

Physiologic measures of pupil diameter, patellar reflexes, blood

pressure, heart rate, and body temperature are typically used for

assessing the development of tolerance to the hallucinogens.

Subjective effects are assessed using the 70 item LSD questionnaire

(Isbell et al. 1956) and a clinical ordinal scale sensitive to

expressions of anxiety at the low end, and perceptual distortions

as well as hallucinations at the high end.

have been found,

While some exceptions

chronic treatment with LSD leads to a rapidly

developing tolerance to these effects (Isbell et al. 1956).

Cross-tolerance in LSD-treated subjects to the acute effects of

N,N-dimethyltryptamine (DMT) has also been found to occur for the

above physiologic effects as well as subjective reports after drug

administration (Rosenberg et al. 1964).

140
Wolbach et al. (1962) studied the effects of chronically

administered mescaline and found tolerance to both the subjective

effects and the mydriasis. In addition, marked cross-tolerance to

the effects of LSD on subjective response, pupillary diameter, and

blood pressure were found. The reverse situation was also observed

(i.e., subjects tolerant to the effects of LSD were cross-tolerant

to mescaline).

b. Withdrawal from Chronic Drug Administration - -

While the degree of tolerance that develops to the effects of these

compounds is quite extensive, no signs of abstinence have been

reported in subjects treated chronically with LSD.

3. Hallucinogen Abuse Liability

a. Drug Self-Administration

To date, no systematic studies have been undertaken to characterize

the profile of hallucinogen self-administration in man. There are

obvious difficulties in conducting such studies, which require

rigorous control of the subject's environment during

self-administration, including lighting conditions and visual and

auditory stimuli as well as social contacts.

b. Self-Report of Drug Effects

Procedures for assessing the subjective effects of the hallucinogens

have centered around the use of questionnaires that are completed at

regular intervals after drug administration. Using the MMPI,

Belleville (1956) found that LSD elevated the Psychasthenia,

Schizophrenia, Paranoia, and Taylor Anxiety scales. Klee et al.

(1961) found that the clinical grades of LSD reaction were dose

related. These were found to range from expressions of anxiety and

nervousness to perceptual distortion or hallucinations without

recognition of the drug-related nature of these effects.

The LSD Significant Scale of the ARCI (Hill et al. 1963a) contains

items that are related to expressions of euphoria and sedation,

anxiety, restlessness and dysphoria, disruption of thought

processes, paresthesia, perceptual changes, changes in self-image,

and autonomic changes. The specific questions that relate to the

above effects are outlined by Martin and Sloan (1977). Lower doses

of LSD were found to produce expressions of euphoria, while higher

doses were associated with nervousness and anxiety (Hill et al.

1963b).

Snyder et al. (1974) studied the stereospecific actions of DOET in

man using the Lon Outpatient Mood Scale, Outpatient Symptom Check

List, Hildreth Current Reaction Scales, and the Block Design and

Digit Span subtests of the Wechsler Adult Intelligence Scale.

Subjects were also evaluated using a modified clinical grade scale.

All drugs and placebo were administered under double-blind

conditions while the subjects resided in a quiet comfortable room.

A variety of art objects as well as records were available. The (-)

141
or "R" isomer was found to be four times more potent than the (+) or

"S" isomer. This study represents a novel approach to confirming

the active chemical conformation of a hallucinogen at its receptor

site.

C. Behavioral Toxicity

Procedures for assessing the effects of hallucinogens on psychomotor

performance have mainly measured visual acuity and reaction time.

Edwards and Cohen (1961) studied the effects of LSD on visual

illusion, tactile sensibility, and reaction time in normal

volunteers. Subjects were tested for their ability to detect colors

while seated 10 feet from a translucent light filter (12.5 cm

diameter) surrounded by a black panel. By randomly presenting

different colors at various intensities using the Method of Constant

Stimuli, the threshold for detection of- each color could be

obtained. LSD had no effect on color detection, but size constancy

(assessed by maneuvering two triangles until they appeared to be the

same size) was affected, with the subjects becoming more susceptible

to illusions of similar sizes. LSD also significantly elevated the

difference threshold in a warmth detection discriminator procedure

and increased visually determined reaction times. When a buzzer was

used instead of light for reaction time determinations, no

significant drug effects were observed.

Hartman and Hollister (1963) compared three hallucinogens

(mescaline, LSD, psilocybin) for their effect on color perception

using the Farnsworth-Munsell hue discriminator which consists of 85

colored buttons arranged in four banks of graded hues. The buttons

are mixed thoroughly and the subjects are required to replace them

in the correct sequence of graded hue. After-images were also

determined by having the subjects look at a bright light for 5

seconds, close their eyes, and then name and pick out the colors that

they perceived. All three compounds impaired the subjects' ability

to distinguish the colors. LSD produced more disruptive effects in

the red-yellow and yellow-orange range, while mescaline increased

errors throughout a wider range. Psilocybin increased errors in all

areas except green-blue. LSD and mescaline caused more colors to be

evoked by flicker, but only psilocybin significantly increased the

duration of after-image. Thus, the results of this study indicate

that tests requiring subjective reports of color evoked artificially

are more sensitive to hallucinogens than tests of color perception

based upon hue discrimination procedures.

References

Belleville, R.E. MMPI score changes induced by lysergic acid

diethylamide (LSD-25). J Clin Psychol 12:279-282 -1956.

Edwards, A.F., and Cohen, S. Visual illusion, tactile sensibility

and reaction time under LSD-25. Psychopharmacologia,

2:7297-303. 1961.

Goldstein, L.; Murphree, H.B.; Sugerman, A.A.; Pfeiffer, C.C.; and

Jenny, E.H. Quantitative electroencephalographic analysis of

naturally occurring (schizophrenic) and drug-induced psychotic

states in human males. Clin Pharmacol Ther, 4:10-21, 1963.

142
Hartman, A.M., and Hollister, L.E. Effects of mescaline, lysergic

acid diethylamide and psilocybin on color perception.

Psychopharmacologia, 4:441-451, 1963.

Hill, H.E.; Haertzen, C.A.; Wolbach, A.B.; and Miner, E.J. The

Addiction Research Center Inventory: Standardization of scales

which evaluate subjective effects of morphine, amphetamine,

pentobarbital, alcohol, LSD-25, pyrahexyl, and chlorpromazine.

Psychopharmacologia, 4:167-183, 1963a.

Hill, H.E.; Haertzen, C.A.; Wolbach. A.B.; and Miner, E.J. The

Addiction Research Center Inventory: Appendix I. Items

comprising empirical scales for seven drugs. II. Items which

do not differentiate placebo from any drug condition.

Psychopharmacologia, 4:184-205, 1963b.

Isbell, H.; Belleville, R.E.; Fraser, H.F.; Wikler, A.; and Logan,

C.R. Studies on lysergic acid diethylamide (LSD-25). I.

Effects in former morphine addicts and development of tolerance

during chronic intoxication. Arch Neurol Psychiat, 76:468-478,

1956.

Itil, T.M., and Fink, M. Anticholinergic drug-induced delerium:

Experimental modification, quantitative EEG, and behavioral

correlations. J Nerv Ment Dis, 143:492-507, 1966.

Klee. G.D.; Bertino, J.; Wentraub. W.; and Callaway, E. The

influences of varying dosage on the effects of lysergic acid

diethylamide (LSD-25) in humans. J Nerv Ment Dis, 132:404-409,

1961.

Korein, J., and Mussacchio, J.M. LSD and focal cerebral lesions.

Behavioral and EEG effects in patients with sensory defects.

Neurology, 18:147-152, 1968.

Martin, W.R., and Sloan, J.W. Pharmacology and classification of

LSD-like hallucinogens. In: Martin, W.R., ed. Drug

Addiction. Vol. 2. Berlin: Springer-Verlag, 1977. pp.

305-368.

Monroe, R.R.; Heath, R.G.; Mickle, W.A.; and Llewellyn, R.C.

Correlation of rhinencephalic electrograms with behavior. A

study of humans under the influence of LSD and mescaline.

Electroenceph Clin Neurophysiol, 9:623-642, 1957.

Rodin, E., and Luby, E. Effects of LSD-25 on the EEG and photic

evoked responses. Arch Gen Psychiat, 14:435-441, 1966.

Rosenberg, D.E.; Isbell, H.; Miner, E.J.; and Logan, C.R. The

effects of N,N-dimethyltryptamine in human subjects tolerant to

lysergic acid diethylamide. Psychopharmacologia, 5:217-227,

1964.

Shagass, C. Effects of LSD on somatosensory and visual evoked

responses and on the EEG in man. In: Wortis. J., ed. Recent

Advances in Biological Psychiatry. Vol. 9. New York: Plenum

Press, 1961.

Snyder, S.H.; Unger, S.; Blatchley, R.; and Barfknecht, C.F.

Stereospecific actions of DOET (2,5-dimethoxy-4-

ethylamphetamine) in man. Arch Gen Psychiat, 31:103-106, 1974.

Wolbach, A.B.; Isbell, H.; and Miner, E.J. Cross tolerance between

mescaline and LSD-25 with a comparison of the mescaline and LSD

reactions. Psychopharmacologia, 3:1-14, 1962.

143
6. Procedures for Assessing Dissociative Anesthetics -

1. Characterization of Dissociative Anesthetic Effects

a. Acute Drug Effects

i) Anesthesia

The characteristics of the various stages of anesthesia following

both inhalation and intravenous administration of dissociative

anesthetics have been defined by Gillespie (1943) and remain in use

today. Four stages are delineated in the progression from loss of

consciousness through delirium and surgical anesthesia, to medullary

depression and cardiovascular collapse. While early clinical

studies showed that phencyclidine (PCP) readily induced and

maintained anesthesia, the incidence of delirium, hallucinations,

and disruptive reactions discouraged further use in surgical

procedures (Johnstone et al. 1959; Luby et al. 1959; Domino 1968).

Some of these effects are also seen with ketamine, but the incidence

and severity are lower.

Stella et al. (1979) determined the relative potencies of ketamine

with respect to a number of other anesthetics using standardized

probit analysis. Drugs were administered via a large antecubital

vein 20 seconds after releasing a sphygmomanometer cuff. A 10 ml

volume was injected over a 15-second interval during which the

subjects were asked to count out loud. When they stopped counting and

did not respond to a request to open their eyes, they were considered

sleeping. Using a total of 450 subjects, dose-respose relationships

were obtained by considering the percentage of positive responses

(i.e., asleep) in groups of subjects given different doses of each

agent. In this manner, the UD30, UD50, and UD80 (unconscious dose

30%, 50%. and 80%, respectively) were determined. Ketamine was

found to be about 4.4 times more potent than thiopentone and about

equipotent with diazepam. The UD95 of each drug was calculated and

is considered to be the clinically relevant dose.

ii) Analgesia

Sadove et al. (1971) determined the analgesic effects of ketamine

both by an experimentally induced pain technique and by questioning

individuals during recovery from surgery. Painful stimuli were

produced using an earlobe algesimeter (Siker et al. 1966). While

the earlobe contains undifferentiated bare nerve endings, subjects

have easily distinguished between heat, cold prick, and touch

(Sinclair et al. 1952). The current delivered to the earlobe is

adjusted from 8 to 30 microamperes and after 3 determinations of

control responses (time to release button), separate measures of

pain threshold were obtained at 15-minute intervals after drug

injection. Ketamine was compared with meperidine and saline and was

found to increase the pain threshold to a level higher than

meperidine, but its duration was shorter (about 75 minutes vs. 120

minutes for ketamine and meperidine, respectively). In the clinical

double-blind study of post-operative pain, subjects were asked to

rate their pain on a 0 to 3 scale ("none," "mild," "moderate," and

144
"severe"). Beginning with a pain scale score of 2, they were

re-evaluated at 5-and lO-minute intervals after receiving 0.44 mg/kg

of ketamine, i.m., or placebo. Ketamine significantly reduced the

pain scores, but the duration of this analgesic effect was limited

to 1 hour.

iii) Electroencephalographic (EEG) Activity

Procedures for assessing the EEG effects of dissociative anesthetics

in man using visual inspection techniques have shown that doses of

2-3 mg of PCP produce slowing of the EEG that is most pronounced in

the theta band. Higher doses produce more diffuse, slow activity

mostly in the occipital, temporal, and parietal areas (Greifenstein

et al. 1958). The EEG effects of ketamine have been shown to be

similar in nature to PCP (Schwartz et al. 1974) using conventional

EEG tracings and a single-channel cerebral function monitor (CFM)

(Sully and Scott 1972) which graphs the mean amplitude of the EEG

over a 2-50 Hz band on slowly moving paper. The CFM consistently

detected changes 1O-15 seconds before the direct EEG when injections

were given over both a 15-and 60-second interval. The differences

in onset of effect were less for the second ketamine injection (same

dose) given 10 minutes after the first.

Most recently, power spectral analysis has been used to quantify the

frequency and voltage components of the EEG after PCP intoxication

in one patient (Stockard et al. 1976). A prominent theta peak (5.75

Hz) persisted during the period when the patient was unconscious and

unresponsive to stimulation. Intravenous administration of diazepam

produced a shift to a lower predominant frequency accompanied by a

return of spontaneous movements, though normal alpha frequencies

(i.e., 8.5 Hz) were recovered only after 72 hours.

2. Dissociative Anesthetic Physical Dependence Potential

a. Tolerance to Drug Effects

Numerous case reports of tolerance to the anesthetic and analgesic

doses of ketamine, particularly in pediatric medicine, have appeared

stating the approximate percent increase in dose required to

maintain adequate anesthesia (Cronin et al. 1972; Stevens and Hain

1981). No attempts to control for duration between treatments in

these studies were reported, however, and no systematic quantitative

studies of tolerance are available.

b. Withdrawal from Chronic Drug Administration

While drug-seeking behavior has been noted to occur, no signs of

withdrawal were noted in individuals who reported smoking 100 mg or

more of PCP daily for several months (Burns and Lerner 1981). To

date, however, no quantitative methodology has been applied to

determine the nature and extent of the abstinence syndrome following

withdrawal from chronic administration of dissociative anesthetics.

145
3. Dissociative Anesthetic Abuse Liability

a. Drug Self-Administration

There are currently no accepted procedures for assessing the ability

of PCP or its analogues to maintain self-injection performance in

humans.

b. Self-Report of Drug Effects

Numerous studies of PCP have described verbal reports of subjective

effects involving impairments of thought processes, changes in body

image and sensory perception, and depersonalization concomitant

with impaired attention and concentration (Burns and Lerner 1981).

Occasionally delusions and hallucinations have been reported, as

after-effects usually associated with the recovery following drug

withdrawal. No reports are available describing the use of

standardized psychological testing procedures (e.g., MMPI, ARCI).

c. Behavioral Toxicity

Morgenstern et al. (1962) administered a battery of seven tests to

normal volunteers in order to study the effects of PCP on sensory

processes. After swallowing a 7.5 mg tablet, the battery of tests

(which took 10-12 minutes to complete) was repeated every 15 minutes

for 2 hours. Phencyclidine produced a general impairment in sensory

function as evidenced by the elevated thresholds on all seven tests.

Two-point discrimination, aesthesiometry, and perimetry thresholds

showed the greatest increase, while taste was the least affected. In

addition, the PCP-induced sensory disturbances always occurred

before any verbal reports of subjective changes in feelings or mood.

PCP's effects on reaction time, rotary pursuit, and weight

discrimination have also been compared with LSD-25 and amobarbital

sodium in normal volunteers (Rosenbaum et al. 1959). PCP and

amobarbital were given i.v., while LSD was given orally. The tests

were administered imediately after PCP and amobarbital, and 3 hours

after LSD. A marked increase in reaction time was observed after

PCP with a lesser increase observed after amobarbital. LSD did not

affect reaction time. The ability to trace a rotating disc was also

impaired by PCP, though amobarbital had little or no effect, while

LSD actually improved performance on this task. PCP significantly

disrupted performance on the weight discrimination tasks as well,

though amobarbital and LSD had no such effect.

References

Burns, R.S., and Lerner, S.E. The effects of phencyclidine in man:

A review. In: Domino, E.F., ed. PCP (Phencyclidine):

Historical and Current Perspectives. Ann Arbor, Michigan: NPP

Books, 1981. pp. 449-469.

Cronin, M.M.; Bousfield, J.A.; Hewett, E.B.; McLellan, I.; and

Boulton, T.B. Ketamine anaesthesia for radiotherapy in small

children. Anaesthesia, 27:135-142, 1972.

146
Domino, E.F. Neurobiology of Phencyclidine (Sernyl), a drug with an

unusual spectrum of pharmacological activity. Int Rev

Neurobiol, 6:303-347, 1964.

Gillespie, N.A. The signs of anaesthesia. Curr Res Anesth Analg,

22 1275-282. 1943.

Greifenstein, F.E.; DeVault, M.; Yoshitake, J.; and Gajewski, J.E.

A study of a 1-aryl cycle hexyl amine for anesthesia. Anesth

Analg, 37:283-294, 1958.

Johnstone, M.; Evans, V.; and Brigel, S. Sernyl (CI-395) in

clinical anaesthesia. Brit J Anaesth, 31:433-439, 1959.

Luby, E.D.; Cohen, B.D.; Rosenbaum, G.; Gottlieb, J.S.; and Kelley,

R. Study of a new schizophrenomimetic drug - Sernyl. AMA Arch

Neurol Psychiat, 81:363-369, 1959.

Morgenstern, F.S.; Beech, H.R.; and Davies, B.M. An investigation

of drug-induced sensory disturbances. Psychopharmacologia, 3:

193-201, 1962.

Rosenbaum, G.; Cohen, B.D.; Luby, E.D.; Gottlieb, J.S.; and Yelen,

D. Comparison of Sernyl with other drugs. Arch Gen Psychiat,

1:651-656, 1959.

Sadove, M.S.; Shulman, M.; Hatano, S.; and Fevold, N. Analgesic

effects of ketamine administered in subdissociative doses.

Anesth Analg, 50:452-457, 1971.

Schwartz, M.S.; Virden. S.; and Scott, D. Effects of ketamine on

the-electroencephalograph. Anaesthesia, 29:135-140, 1974.

Siker, E.S.; Wolfson, B.; and Stewart, W.D. Further experience with

the earlobe algesimeter. Anesthesiology, 27:88-92, 1966.

Sinclair, C.D.; Weddel, G.; and Zander, E. Relationship of

cutaneous sensibility to neurohistology in human pinna. J

Anat, 86:402, 1952.

Stella, L.; Torri, G.; and Castiglioni, C.L. The relative potencies

of thiopentone, ketamine, propernidid, alphaxalone and

diazepam. Brit J Anaesth, 51:119-122, 1979.

Stevens, R.W., and Hain, W.R. Tolerance to rectal ketamine in

paediatric anaesthesia. Anaesthesia, 36:1089-1093, 1981..

Stockard, J.J.; Werner, S.S.; Aalbers, J.A.; and Chiappa, K.H.

Electroencephalographic findings in phencyclidine intoxication.

Arch Neurol, 33:200-203, 1976.

Sully, M., and Scott, D.F. Testing a new anaesthetic drug. J

Electrophys Technol ASSOC. 19:53, 1972.

147
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77 ADOLESCENT DRUG ABUSE: ANALYSES OF TREATMENT RESEARCH.

Elizabeth R. Rahdert, Ph.D. and John Grabowski, Ph.D., eds.

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Alcohol, Drug Abuse, and Mental Health Administration

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