Method of simultaneously enhancing analgesic potency and attenuating dependence liability caused by morphine and other opioid agonists

This invention relates to a method of selectively enhancing the analgesic potency of morphine and other clinically used bimodally-acting opioid agonists and simultaneously attenuating development of physical dependence, tolerance and other undesirable side-effects caused by the chronic administration of said bimodally-acting opioid agonists comprising the co-administration of a bimodally-acting opioid agonist which activates inhibitory opioid receptor-mediated functions of neurons in the nociceptive (pain) pathways of the nervous system and an opioid receptor antagonist which selectively inactivates excitatory opioid receptor-mediated side-effects caused by said bimodally-acting opioid agonists. This invention further relates to a method of detoxifying and treating opiate addicts utilizing said opioid receptor antagonists, as well as to a composition comprising an excitatory opioid receptor antagonist of the invention and a bimodally-acting opioid agonist.

FIELD OF THE INVENTION 
This invention relates to a method of enhancing the analgesic (inhibitory) 
effects of bimodally-acting opioid agonists, including morphine and other 
clinically used opioid analgesics, while at the same time attenuating 
anti-analgesic effects, physical dependence, tolerance, hyperexcitability, 
hyperalgesia, and other undesirable (excitatory) side-effects typically 
caused by chronic use of bimodally-acting opioid agonists. As used herein, 
the term "opioid" refers to compounds which bind to specific opioid 
receptors and have agonist (activation) or antagonist (inactivation) 
effects at these receptors, such as opioid alkaloids, including the 
agonist morphine and the antagonist naloxone, and opioid peptides, 
including enkephalins and dynorphins. As used herein, the term "opiate" 
refers to drugs derived from opium or related analogs. 
In the instant invention, a selective excitatory opioid receptor antagonist 
is combined with a reduced dose of a bimodally-acting opioid agonist so as 
to elicit the desired degree of analgesia (inhibitory effects) and 
attenuate undesired side-effects (excitatory effects). Opioid analgesia 
results from activation (by opioid agonists) of inhibitory opioid 
receptors on neurons in the nociceptive (pain) pathways of the peripheral 
and central nervous systems. The undesirable side-effects, including 
anti-analgesic actions, the development of physical dependence, some types 
of tolerance, hyperexcitability and hyperalgesia, result from sustained 
activation (by bimodally-acting opioid agonists) of excitatory opioid 
receptors on neurons in the nociceptive (pain) pathways of the peripheral 
and central nervous systems. The administration of selective excitatory 
receptor antagonists together with bimodally-acting opioid agonists 
enhances analgesic effects caused by said opioids and attenuates the 
development of physical dependence, tolerance and other undesirable 
side-effects which are also caused by said opioids. In addition, combined 
use of the opioid receptor antagonists and agonists of the invention can 
be used for more effective detoxification and treatment of opiate addicts. 
BACKGROUND OF THE INVENTION 
Morphine or other bimodally-acting opioid agonists are administered to 
relieve severe pain due to the fact that they have analgesic effects 
mediated by their activation of inhibitory opioid receptors on nociceptive 
neurons (see North, Trends Neurosci., Vol. 9, pp. 114-117 (1986) and Crain 
and Shen, Trends Pharmacol. Sci., Vol. 11, pp. 77-81 (1990)). However, 
bimodally-acting opioid agonists also activate opioid excitatory receptors 
on nociceptive neurons, which attenuates the analgesic potency of said 
opioids and results in the development of physical dependence thereon and 
increased tolerance thereto (see Shen and Crain, Brain Res., Vol. 597, pp. 
74-83 (1992)), as well as hyperexcitability, hyperalgesia and other 
undesirable (excitatory) side-effects. As a result, a long-standing need 
has existed to develop a method of both enhancing the analgesic 
(inhibitory) effects of bimodally-acting opioid agonists and limiting the 
undesirable (excitatory) side-effects caused by such opioid agonists. 
The parent Patent Application for the instant invention, Ser. No. 
07/947,690, relates to a specific group of opioid agonists for use as 
low/non-addictive analgesics and for the treatment of opioid addiction. In 
the parent Application, it is stated that this group of opioid agonists 
(which includes etorphine and dihydroetorphine) bind to and activate 
inhibitory but not excitatory opioid receptors. (In contrast, morphine and 
most other opioid alkaloids and peptides elicit bimodal effects by binding 
to and activating both excitatory and inhibitory opioid receptors.) 
To date, no method has been discovered or developed whereby two opioid 
compounds are administered, one of which binds to and activates inhibitory 
opioid receptors to cause analgesia and the other of which binds to and 
inactivates excitatory opioid receptors so as to attenuate undesirable 
side-effects caused by the administration of bimodally-acting opioid 
agonists while simultaneously enhancing the analgesic effects of said 
bimodally-acting opioid agonists. 
It is therefore an object of this invention to provide a method of 
enhancing the analgesic potency of morphine and other bimodally-acting 
opioid agonists by blocking their anti-analgesic side-effects. 
It is a further object of this invention to provide a method of attenuating 
physical dependence, tolerance, hyperexcitability, hyperalgesia and other 
undesirable side-effects caused by the chronic administration of 
bimodally-acting opioid agonists. 
It is another object of this invention to provide a method for detoxifying 
and treating opiate addicts utilizing excitatory opioid receptor 
antagonists. 
It is yet another object of this invention to provide a composition which 
enhances the analgesic effects of bimodally-acting opioid agonists while 
simultaneously attenuating undesirable side-effects caused by said opioid 
agonists, including physical dependence, tolerance, hyperexcitability and 
hyperalgesia. 
It is still a further object of this invention to provide a composition 
which is useful for detoxification and treatment of opiate addicts. 
SUMMARY OF THE INVENTION 
This invention is directed to a method of selectively enhancing the potency 
of morphine and other conventional bimodally-acting opioid agonists and 
simultaneously attenuating undesirable side-effects, including physical 
dependence, caused by the chronic administration of said opioid agonists. 
Morphine and other bimodally-acting (inhibitory/excitatory) opioid 
agonists bind to and activate inhibitory and excitatory opioid receptors 
on nociceptive neurons mediating pain. Activation of inhibitory receptors 
by said agonists causes analgesia. Activation of excitatory receptors by 
said agonists results in anti-analgesic effects, development of physical 
dependence, tolerance, hyperexcitability, hyperalgesia and other 
undesirable side-effects. The co-administration of an opioid antagonist 
which binds to and inactivates excitatory opioid receptors results in the 
blocking of excitatory anti-analgesic side-effects of said opioid agonists 
on these neurons, thereby resulting in enhanced analgesic potency which 
permits the use of lower doses of morphine or other conventional opioid 
analgesics. 
The excitatory opioid receptor antagonists of the invention include 
etorphine, dihydroetorphine, diprenorphine and similarly acting opioid 
alkaloids and opioid peptides. The opioid agonists of the invention 
include morphine or other bimodally-acting (inhibitory/excitatory) opioid 
alkaloids or opioid peptides that are in clinical use as analgesics, 
including codeine, fentanyl analogs and endorphins. 
In addition, combinations of an excitatory opioid receptor antagonist and 
morphine or another conventional bimodally-acting opioid analgesic can be 
used to detoxify and treat opiate addicts.

DETAILED DESCRIPTION OF THE INVENTION 
This invention is directed to a method of selectively enhancing the 
analgesic effect caused by the administration of a bimodally-acting opioid 
agonist and simultaneously attenuating undesirable side-effects caused by 
the chronic administration of said bimodally-acting opioid agonists. This 
is performed by simultaneously inactivating excitatory opioid 
receptor-mediated functions of neurons in the nociceptive (pain) pathways 
and activating inhibitory opioid receptor-mediated functions of 
nociceptive neurons. A bimodally-acting opioid agonist and an excitatory 
opioid receptor antagonist are co-administered. The bimodally-acting 
opioid agonist binds to inhibitory receptors on nociceptive neurons so as 
to activate inhibitory opioid receptor-mediated functions, including 
analgesia, and concomitantly activates excitatory opioid receptors on 
nociceptive neurons. The excitatory opioid receptor antagonist binds to 
excitatory receptors on said neurons and thereby inactivates excitatory 
opioid receptor-mediated functions, including anti-analgesic effects, 
physical dependence and tolerance to the opioid agonist, hyperexcitability 
and hyperalgesia. In addition, this invention is directed to the use of 
said excitatory opioid receptor antagonists and opioid agonists to 
detoxify and treat opiate addicts. Further, this invention is directed to 
a composition comprising an excitatory opioid receptor antagonist and a 
bimodally-acting opioid agonist. 
The inventors have discovered that certain compounds act as excitatory 
opioid receptor antagonists, that is, they bind to and inactivate 
excitatory opioid receptors on neurons in the pain pathways. The 
excitatory opioid receptor antagonists of the invention are preferably 
selected from the group consisting of etorphine, dihydroetorphine and 
diprenorphine. The opioid receptor antagonists of the invention inactivate 
mu, delta, kappa and other subtypes of excitatory opioid receptors. They 
may have varying structures. For example, etorphine and dihydroetorphine 
have very similar chemical structures and are considered to be potent 
analgesics which selectively activate inhibitory but not excitatory opioid 
receptors (see Shen and Crain, Regulatory Peptides, in press (1993)). In 
contrast, diprenorphine has a somewhat different chemical structure than 
etorphine and dihydroetorphine and has been previously considered to act 
as a universal opioid receptor antagonist by inactivating all types of 
inhibitory and excitatory opioid receptors (see Shen and Crain, Brain 
Res., Vol. 491, pp. 227-242 (1989)). Nevertheless, these three compounds 
are all capable of selectively binding to and inactivating excitatory 
opioid receptors on nociceptive neurons when administered at low 
concentrations. 
The bimodally-acting opioid agonists of this invention preferably include 
morphine, codeine, fentanyl analogs, endorphins, and other opioid 
alkaloids and opioid peptides. Typically, the opioid agonists of the 
invention are mu, delta, kappa or epsilon opioid receptor agonists, and 
are capable of binding to inhibitory opioid receptors on neurons in the 
pain pathway. When these bimodally-acting agonists bind to inhibitory 
opioid receptors, they thereby activate inhibitory opioid 
receptor-mediated functions, including analgesia. 
As discussed below, the inventors have discovered that certain compounds 
(the excitatory opioid receptor antagonists of the invention), when 
co-administered with bimodally-acting opioid agonists, are capable at very 
low dosages of enhancing the analgesic effects of the bimodally-acting 
opioid agonists at least 10-1000 fold by inactivating excitatory 
anti-analgesic side-effects of said agonists. In addition, the excitatory 
opioid receptor antagonists of the invention inactivate other excitatory 
receptor-mediated functions, and thereby reduce the undesirable 
side-effects caused by said opioid agonists, including development of 
physical dependence, tolerance to the opioid agonists, hyperexcitability 
and hyperalgesia. In addition, the excitatory opioid receptor antagonists 
of the invention may be administered to detoxify and treat opiate addicts. 
Ordinarily, bimodally-acting opioid agonists are administered clinically in 
pill form and are administered in milligram dosages. By co-administering 
bimodally-acting opioid agonists with the excitatory opioid receptor 
antagonists of the invention, it is possible to achieve an analgesic 
effect with 10-1000 times lower doses of the bimodally-acting opioid 
agonist than when said opioid agonist is administered alone. This is 
because the excitatory opioid receptor antagonists of the invention 
enhance the analgesic effects of the bimodally-acting opioid agonists by 
attenuating the anti-analgesic excitatory side-effects of said opioid 
agonists. Hence, bimodally-acting opioid agonists which are administered 
with the excitatory opioid receptor antagonists of the invention are 
administered in an amount 10-1000 times less than the amount of that 
bimodally-acting opioid agonist which has typically been administered for 
analgesia. 
According to the present invention, the dose of excitatory opioid receptor 
antagonist to be administered is 10-1000 times less than the dose of 
bimodally-acting opioid agonist to be administered, for example, &lt;1 
microgram of said antagonist together with 10-100 micrograms of said 
agonist. These estimates of dosages are based on studies of neurons in 
culture. The excitatory opioid receptor antagonists, as well as the opioid 
agonists, can be administered sublingually, intramuscularly, 
subcutaneously or intraveneously. 
The co-administration of the opioid agonists and excitatory opioid receptor 
antagonists of the invention simultaneously activates inhibitory functions 
of nociceptive neurons in the pain pathway and inactivates excitatory 
functions of the same or other nociceptive neurons. In order to 
demonstrate this, electrophysiologic studies on the effects of opioids on 
mouse sensory dorsal root ganglion neurons in tissue cultures were 
performed. It is shown below that this bimodal modulation is mediated by 
activating putative excitatory opioid receptors in addition to previously 
characterized inhibitory opioid receptors on sensory neurons. 
It is shown that at low pM-nM concentrations, nearly all opioids, including 
morphine, enkephalins, dynorphins, endorphins and specific mu, delta and 
kappa opioid agonists, elicit naloxone-reversible dose-dependent 
excitatory effects manifested by prolongation of the calcium-dependent 
component of the action potential duration (APD) of dorsal root ganglia 
(DRG) neurons. In contrast, the same opioids generally elicit inhibitory 
APD shortening effects when applied at higher concentrations (0.1-1 
.mu.M). 
The excitatory opioid effects on sensory neurons have been shown to be 
mediated by opioid receptors that are coupled via a 
cholera-toxin-sensitive stimulatory GTP-binding protein, Gs, to adenylate 
cyclase/cyclic AMP/protein kinase A-dependent ionic conductances that 
prolong the APD (resembling, for example, beta-adrenergic receptors). (See 
Crain and Shen, Trends Pharmacol. Sci., Vol. 11, pp. 77-81 (1990)). On the 
other hand, inhibitory opioid effects are mediated by opioid receptors 
that are coupled via pertussis toxin-sensitive inhibitory G proteins: Gi 
to the adenylate cyclase/cyclic AMP system and Go to ionic conductances 
that shorten the APD (resembling, for example, alpha.sub.2 -adrenergic 
receptors). Shortening by opioids of the action potential of primary 
sensory neurons has generally been considered to be a useful model of 
their inhibition of calcium influx and transmitter release at presynaptic 
terminals in the dorsal spinal cord, thereby accounting for opioid-induced 
analgesia in vivo. (See North, Trends Neurosci., Vol. 9, pp. 114-117 
(1986) and Crain and Shen, Trends Pharmacol. Sci., Vol. 11, pp. 77-81 
(1990)). Similarly, the delayed repolarization associated with the 
observed opioid-induced prolongation of action potential has been 
interpreted as evidence of excitatory effects of opioids on sensory 
neurons that may result in enhanced calcium influx and transmitter release 
at presynaptic terminals. This could account for some types of 
hyperalgesia and hyperexcitatory states elicited by opioids in vivo (see 
Crain and Shen, Trends Pharmacol. Sci., Vol. 11, pp. 77-81 (1990) and Shen 
and Crain, Brain Res., Vol. 491, pp. 227-242 (1989). 
Chronic treatment of DRG neurons with typical bimodally-acting 
(excitatory/inhibitory) opioids (e.g., 1 .mu.M D-ala.sup.2 -D-leu.sup.5 
enkephalin (DADLE) or morphine for 1 week) results in tolerance to the 
usual inhibitory APD-shortening effects of high concentrations of these 
opioids and supersensitivity to the excitatory APD-prolonging effects of 
these opioid agonists, as well as the opioid antagonist, naloxone (see 
Crain and Shen, Brain Res., Vol. 575, pp. 13-24 (1992) and Shen and Crain, 
Brain Res., Vol. 597, pp. 74-83 (1992)). It has been suggested that the 
latter electrophysiologic effects and related biochemical adaptations are 
cellular manifestations of physical dependence that may underlie some 
aspects of opiate addiction (see Shen and Crain, Brain Res., Vol. 597, pp. 
74-83 (1992) and Terwilliger et al., Brain Res., Vol. 548, pp. 100-110 
(1991)). 
In contrast to bimodally-acting opioids, it has been discovered by the 
inventors that the opioid alkaloids, etorphine (see Bentley and Hardy, 
Proc. Chem. Soc., pp. 220 (1963)) and dihydroetorphine (see Bentley and 
Hardy, J. Amer. Chem. Soc., Vol. 89, pp. 3281-3286 (1967)) uniquely elicit 
dose-dependent, naloxone-reversible inhibitory effects on sensory neurons 
in DRG-spinal cord explants, even at concentrations as low as 1 pM, and 
show no excitatory effects at lower concentrations (see Shen and Crain, 
Regulatory Peptides, in press (1993)). In addition, these potent 
inhibitory opioid receptor agonists also display unexpected antagonist 
effects at excitatory opioid receptors on DRG neurons. Acute pretreatment 
of DRG neurons with etorphine or dihydroetorphine, at low concentrations 
(&lt;pM) which do not alter the APD, block the excitatory APD-prolonging 
effects of morphine and other bimodally-acting opioids and unmask 
inhibitory APD-shortening effects which normally require much higher 
concentrations. The potent inhibitory effect of etorphine and 
dihydroetorphine may be due to their selective activation of inhibitory 
opioid receptor-mediated functions while simultaneously inactivating 
excitatory opioid receptor-mediated functions in sensory neurons. In 
contrast, bimodally-acting opioids activate high-affinity excitatory as 
well as inhibitory opioid receptors on DRG neurons, thereby decreasing the 
net inhibitory effectiveness of these agonists, resembling the attenuation 
of the inhibitory potency of systemic morphine by the "anti-analgesic" 
(excitatory) effect of dynorphin A release in spinal cord in mice (see 
Fujimoto et al., Neuropharmacol., Vol. 29, pp. 609-617, (1990)). 
Acute application of pM-nM etorphine or dihydroetorphine to chronic .mu.M 
morphine-treated DRG neurons elicited marked APD shortening (as in naive 
cells) even when added during naloxone-precipitated APD-prolongation in 
these sensitized cells, whereas 10 .mu.M morphine or DADLE were 
ineffective and in contrast elicited a marked APD prolongation. These 
potent inhibitory effects of etorphine and dihydroetorphine on the action 
potential of chronic morphine-treated sensory neurons show remarkable 
mimicry of the rapid DHE-induced blockade of naloxone-evoked withdrawal 
syndromes in opiate-addicted animals and humans (see Wang et al., Chinese 
J. Pharmacol. Toxicol., Vol. 6, pp. 36-40 (1992) and Qin, Chinese J. 
Pharmacol. Toxicol., Vol. 6 (1992)) and the absence of cross-tolerance to 
etorphine in chronic morphine-treated mice even when the analgesic ED50 
for morphine had increased 15-fold (see Lange et al., Toxicol. Applied 
Pharmacol., Vol. 54, pp. 177-186 (1980)). Furthermore, chronic treatment 
of DRG neurons with 10 nM etorphine for &gt;1 week did not result in opioid 
excitatory supersensitivity, i.e., APD prolongation following acute 
application of fM dynorphin A (1-13) or nM naloxone, nor tolerance to 
opioid inhibitory effects, all of which occur after chronic treatment with 
bimodally-acting opioids, e.g., DADLE or morphine (see Crain and Shen, 
Brain Res., Vol. 575, pp. 13-24 (1992) and Shen and Crain, Brain Res., 
Vol. 597, pp. 74-83 (1992)). 
In vitro studies on sensory neurons suggested that an opioid which can 
selectively activate inhibitory, and inactivate excitatory, opioid 
receptor functions would be a unique analgesic in vivo with high potency, 
low dependence liability, and useful for treatment of opiate addicts. 
EXAMPLE 1 
Etorphine And Dihydroetorphine Act As Potent Selective Antagonists At 
Excitatory Opioid Receptors On DRG Neurons Thereby Enhancing Inhibitory 
Effects Of Bimodally-Acting Opioid Agonists 
Methods: The experiments described herein were carried out on dorsal root 
ganglion (DRG) neurons in organotypic explants of spinal cord with 
attached DRGs from 13-day-old fetal mice after 3 to 5 weeks of maturation 
in culture. The DRG-cord explants were grown on collagen-coated coverslips 
in Maximow depression-slide chambers. The culture medium consisted of 65% 
Eagle's minimal essential medium, 25% fetal bovine serum, 10% chick embryo 
extract, 2 mM glutamine and 0.6% glucose. During the first week in vitro 
the medium was supplemented with nerve growth factor (NGF-7S) at a 
concentration of about 0.5 .mu.g/ml, to enhance survival and growth of the 
fetal mouse DRG neurons. 
In order to perform electrophysiologic procedures, the culture coverslip 
was transferred to a recording chamber containing about 1 ml of Hanks' 
balanced salt solution (BSS). The bath solution was supplemented with 4 mM 
Ca.sup.2+ and 5 mM Ba.sup.2+ (i.e., Ca,Ba/BSS) to provide a prominent 
baseline response for pharmacological tests. Intracellular recordings were 
obtained from DRG perikarya selected at random within the ganglion. The 
micropipettes were filled with 3M KCl (having a resistance of about 60-100 
megohms) and were connected via a chloridized silver wire to a neutralized 
input capacity preamplifier (Axoclamp 2A) for current-clamp recording. 
After impalement of a DRG neuron, brief (2 msec) depolarizing current 
pulses were applied via the recording electrode to evoke action potentials 
at a frequency of 0.1 Hz. Recordings of the action potentials were stored 
on a floppy disc using the P-clamp program (Axon Instruments) in a 
microcomputer (IBM AT-compatible). 
Drugs were applied by bath perfusion with a manually operated, push-pull 
syringe system at a rate of 2-3 ml/min. Perfusion of test agents was begun 
after the action potential and the resting potential of the neuron reached 
a stable condition during &gt;4 minute pretest periods in control Ca, Ba/BSS. 
Opioid-mediated changes in the APD were considered significant if the APD 
alteration was &gt;10% of the control value for the same cell and was 
maintained for the entire test period of 5 minutes. The APD was measured 
as the time between the peak of the APD and the inflection point on the 
repolarizing phase. The following drugs were used: etorphine, 
diprenorphine and morphine (gifts from Dr. Eric Simon); dihydroetorphine 
(gift from Dr. B.-Y. Qin, China); naloxone (Endo Labs); DADLE, dynorphin 
and other opioid peptides (Sigma). 
Opioid alkaloids and peptides were generally prepared as 1 mM solutions in 
H.sub.2 O and then carefully diluted with BSS to the desired 
concentrations, systematically discarding pipette tips after each 
successive 1-10 or 1-100 dilution step to ensure accuracy of extremely low 
(fM-pM) concentrations. 
Results: The opioid responsiveness of DRG neurons was analyzed by measuring 
the opioid-induced alterations in the APD of DRG perikarya. A total of 64 
DRG neurons (from 23 DRG-cord explants) were studied for sensitivity to 
progressive increases in the concentration of etorphine (n=30) or 
dihydroetorphine (n=38). Etorphine rapidly and dose-dependently shortened 
the APD in progressively larger fractions of DRG cells at concentrations 
from 1 fM (30% of cells; n=26) to 1 .mu.M (80% of cells; n=16) (see FIGS. 
1 and 2). 
FIG. 1 shows that acute application of low (pM-nM) concentrations of 
etorphine to naive DRG neurons elicits dose-dependent, naloxone-reversible 
inhibitory shortening of the action potential duration (APD). In contrast, 
dynorphin (and many other bimodally-acting opioid agonists, e.g., 
morphine, DADLE) elicit excitatory APD prolongation at these low 
concentrations (see FIG. 2), which can be selectively blocked by &lt;pM 
levels of etorphine or diprenorphine (see FIG. 3). FIG. 1A record 1 shows 
the action potential (AP) generated by a DRG neuron in balanced salt 
solution containing 5 mM Ca.sup.2+ and 5 mM Ba.sup.2+ (BSS). AP response 
in this record (and in all records below) is evoked by a brief (2 msec) 
intracellular depolarizing current pulse. FIG. 1A records 2-5 show that 
APD is not altered by bath perfusion with 1 fM etorphine (Et) but is 
progressively shortened in 1 pM, 1 nM and 1 .mu.M concentrations (5 minute 
test periods). FIG. 1A record 6 shows that APD returns to control value 
after transfer to BSS (9 minute test). FIG. 1B records 1 and 2 show that 
APD of another DRG neuron is shortened by application of 1 nM etorphine (2 
minute test). FIG. 1B record 3 shows that APD returns to control value 
after transfer to 10 nM naloxone (NLX). FIG. 1B records 4 and 5 show that 
APD is no longer shortened by 1 nM or even 1 .mu.M etorphine when 
co-perfused with 10 nM naloxone (5 minute test periods). FIG. 1C records 1 
and 2 show that APD of another DRG neuron is prolonged by application of 3 
nM morphine. FIG. 1C record 3 shows that APD returns to control value by 5 
minutes after washout. FIG. 1C record 4 shows that application of 1 pM 
etorphine does not alter the APD. FIG. 1C record 5 shows that APD is no 
longer prolonged by 3 nM morphine when co-perfused with 1 pM etorphine and 
instead is markedly shortened to a degree which would require a much 
higher morphine concentration in the absence of etorphine. Similar results 
were obtained by pretreatment with 1 pM diprenorphine (see FIG. 3). 
Records in this and subsequent Figures are from DRG neurons in organotypic 
DRG-spinal cord explants maintained for 3-4 weeks in culture. 
FIG. 2 shows dose-response curves demonstrating that etorphine (Et) 
(.quadrature.) and dihydroetorphine (DHE) () elicit only inhibitory 
dose-dependent shortening of the APD of DRG neurons at all concentrations 
tested (fM-.mu.M). In contrast, dynorphin A (1-13) (Dyn) (X) (as well as 
morphine and other bimodally-acting opioids) elicits dose-dependent 
excitatory APD prolongation at low concentrations (fM-nM) and generally 
requires much higher concentrations (about 0.1-1 .mu.M) to shorten the 
APD, thereby resulting in a bell-shaped dose-response curve. Data were 
obtained from 11 neurons for the etorphine tests, 13 for the DHE tests and 
35 for the dynorphin tests; 5, 8 and 9 neurons were tested (as in FIG. 1) 
with all four concentrations of etorphine, DHE and dynorphin, respectively 
(from fM to .mu.M). For sequential dose-response data on the same neuron, 
the lowest concentrations (e.g., 1 fM) were applied first. 
Dihydroetorphine was even more effective (n=38; FIG. 2). Naloxone (10 nM) 
prevented the etorphine- and dihydroetorphine-induced APD shortening which 
was previously elicited in the same cells (n=12; FIG. 1B). These potent 
inhibitory effects of etorphine and dihydroetorphine on DRG neurons at low 
concentrations are in sharp contrast to the excitatory APD-prolonging 
effects observed in similar tests with morphine and a wide variety of mu, 
delta and kappa opioids. None of the DRG neurons tested with different 
concentrations of etorphine or dihydroetorphine showed prominent APD 
prolongation. 
The absence of excitatory APD-prolonging effects of etorphine and 
dihydroetorphine on DRG neurons could be due to low binding affinity of 
these opioid agonists to excitatory opioid receptors. Alternatively, these 
opioids might bind strongly to excitatory receptors, but fail to activate 
them, thereby functioning as antagonists. In order to distinguish between 
these two modes of action, DRG neurons were pretreated with etorphine at 
low concentrations (fM-pM) that evoked little or no alteration of the APD. 
Subsequent addition of nM concentrations of morphine, DAGO, DADLE or 
dynorphin to etorphine-treated cells no longer evoked the usual APD 
prolongation observed in the same cells prior to exposure to etorphine 
(n=11; see FIG. 1C). This etorphine-induced blockade of opioid excitatory 
effects on DRG neurons was often effective for periods up to 0.5-2 hours 
after washout (n=4). 
These results demonstrate that etorphine, which has been considered to be a 
"universal" agonist at mu, delta and kappa opioid receptors (see Magnan et 
al., Naunyn-Schmiedleberg's Arch. Pharmacol., Vol. 319, pp. 197-205 
(1982)), has potent antagonist actions at mu, delta and kappa excitatory 
opioid receptors on DRG neurons, in addition to its well-known agonist 
effects at inhibitory opioid receptors. Pretreatment with dihydroetorphine 
(fM-pM) showed similar antagonist action at excitatory opioid receptor 
mediating nM opioid-induced APD prolongation (n=2). Furthermore, after 
selective blockade of opioid excitatory APD-prolonging effects by 
pretreating DRG neurons with low concentrations of etorphine (fM-pM), 
which showed little or no alteration of the APD, fM-nM levels of 
bimodally-acting opioids now showed potent inhibitory APD-shortening 
effects (5 out of 9 cells) (see FIG. 1C and FIG. 3). This is presumably 
due to unmasking of inhibitory opioid receptor-mediated functions in these 
cells after selective blockade of their excitatory opioid receptor 
functions by etorphine. 
EXAMPLE 2 
Diprenorphine At Low Concentration Also Shows Potent Selective Antagonist 
Action At Excitatory Opioid Receptors 
Drug tests: Mouse DRG-cord explants, grown for &gt;3 weeks as described in 
Example 1, were tested with the opioid antagonist, diprenorphine. 
Electrophysiological recordings were made as in Example 1. 
Results: The "universal" opioid receptor antagonist, diprenorphine was 
previously shown to block, at nM concentrations, both inhibitory APD 
shortening of DRG neurons by .mu.M opioid agonists as well as excitatory 
APD prolongation by nM opioids. Tests at lower concentrations have 
revealed that pM diprenorphine acts selectively as an antagonist at mu, 
delta and kappa excitatory opioid receptors, comparable to the antagonist 
effects of pM etorphine and dihydroetorphine. In the presence of pM 
diprenorphine, morphine (n=7) and DAGO (n=7) no longer elicited APD 
prolongation at low (pM-nM) concentrations (see FIG. 3A). Instead, they 
showed progressive dose-dependent APD shortening throughout the entire 
range of concentrations from fM to .mu.M (see FIG. 3B), comparable to the 
dose-response curves for etorphine and dihydroetorphine (see FIG. 2 and 
FIG. 1C). This unmasking of inhibitory opioid receptor-mediated 
APD-shortening effects by pM diprenorphine occurred even in the presence 
of 10.sup.6 -fold higher concentrations of morphine (see FIG. 3A, records 
11 vs. 5). 
FIG. 3 shows that excitatory APD-prolonging effects elicited by morphine in 
DRG neurons are selectively blocked by co-administration of a low (pM) 
concentration of diprenorphine, thereby unmasking potent dose-dependent 
inhibitory APD shortening by low concentrations of morphine. FIG. 3A 
records 1-4 show that APD of a DRG neuron is progressively prolonged by 
sequential bath perfusions with 3 fM, 3 pM and 3 nM morphine (Mor). FIG. 
3A record 5 shows that APD of this cell is only slightly shortened after 
increasing morphine concentration to 3 .mu.M. FIG. 3A records 6 and 7 show 
that after transfer to BSS, the APD is slightly shortened during 
pretreatment for 17 minutes with 1 pM diprenorphine (DPN). FIG. 3A records 
8-11 show that after the APD reached a stable value in DPN, sequential 
applications of 3 fM, 3 pM, 3 nM and 3 .mu.M Mor progressively shorten the 
APD, in contrast to the marked APD prolongation evoked by these same 
concentrations of Mor in the absence of DPN (see also FIG. 1C). FIG. 3B 
dose-response curves demonstrate similar unmasking by 1 pM DPN of potent 
dose-dependent inhibitory APD shortening by morphine (X) in a group of DRG 
neurons (n=7), all of which showed only excitatory APD prolongation 
responses when tested prior to introduction of DPN (X). Note that the 
inhibitory potency of morphine in the presence of pM DPN becomes 
comparable to that of etorphine and diprenorphine (see FIG. 2). 
EXAMPLE 3 
Enhanced Inhibitory Effect of Etorphine and Dihydroetorphine On Chronic 
Opioid-Treated Sensory Neurons Which Become Supersensitive To Opioid 
Excitatory Effects 
Drug tests: Mouse DRG-cord explants, grown for &gt;3 weeks as described in 
Example 1, were chronically exposed to the bimodally-acting 
(excitatory/inhibitory) delta/mu opioid agonist, DADLE (1 .mu.M) or 
morphine (1 .mu.M) for 1 week or longer and tested acutely with etorphine 
or dihydroetorphine. Electrophysiological recordings were made as in 
Example 1. 
Results: Acute application of fM etorphine to chronic .mu.M DADLE- or 
morphine-treated DRG neurons (for &gt;1 week) was still effective in 
shortening the APD in 30% of the treated neurons (n=23) when tested in the 
presence of .mu.M DADLE or morphine (see FIG. 4). Furthermore, pM levels 
of etorphine shortened the APD in 76% of the cells tested (n=21). 
FIG. 4 shows that after chronic exposure to morphine (Mor), or other 
bimodally-acting opioids, DRG neurons become supersensitive to the 
excitatory APD-prolonging effects of these opioids, whereas etorphine (Et) 
becomes even more effective in eliciting inhibitory shortening of the APD 
of the same DRG neurons when tested acutely in the presence of the chronic 
opioid. Dose-response curves show that after chronic treatment of DRG 
neurons with 3 .mu.M Mor for 2-3 weeks in culture, the magnitude of APD 
shortening elicited by acute application of Et () is markedly enhanced at 
all test concentrations (fM-.mu.M); see typical records in Inset), thereby 
shifting the dose-response curve sharply to the left, as compared to data 
obtained from naive DRG neurons (.quadrature.). In contrast, after washout 
of the chronic morphine with BSS, retests of sequentially increasing 
concentrations of Et from fM to .mu.M result in less prominent APD 
shortening (X), comparable to, or even weaker than, Et effects on naive 
cells (.quadrature.). These results suggest that the apparent enhancement 
in inhibitory potency of Et on chronic Mor-treated neurons is actually due 
to unmasking of inhibitory APD-shortening effects of chronic Mor following 
Et-antagonist action at excitatory opioid receptors, as occurs in tests on 
naive DRG cells (see FIG. 1C). As shown in the inset, FIG. 4 record 1 
shows AP generated by a DRG neuron treated for 3 weeks in culture with 3 
.mu.M Mor and then tested in BSS in the presence of the chronic Mor. FIG. 
4 record 2 shows that 1 fM Et does not alter the APD in the presence of 3 
.mu.M Mor. FIG. 4 records 3-5 show that sequential increases in the 
concentration of Et from 1 pM to 1 .mu.M progressively shortens the APD in 
the presence of 3 .mu.M Mor (whereas dynorphin dose-dependently prolonged 
the APD of the same chronic opioid-treated cell -- not shown; see FIG. 2). 
In contrast, morphine, DADLE and other chronic bimodally-acting 
opioid-treated DRG neurons showed supersensitive excitatory APD-prolonging 
effects when tested with low (fM-pM) concentrations of dynorphin (1-13) 
before (n=13) or after (n=6) washout of the chronic DADLE or morphine. The 
effectiveness of etorphine in eliciting inhibitory APD-shortening in 
chronic opioid-treated DRG neurons appeared to be significantly enhanced 
relative to naive cells. Whereas nM etorphine moderately shortened the APD 
of naive DRG neurons (mean decrease to about 86+6%; n=18; see FIGS. 1A and 
2), this low concentration was much more effective on chronic 
morphine-treated DRG neurons (mean decrease to 66%+7% (n=9)), when tested 
in the presence of morphine (FIG. 4). Dose-response tests of etorphine on 
chronic DADLE- or morphine-treated DRG neurons showed that the magnitude 
of the APD was progressively shortened when the acute etorphine 
concentration was tested sequentially from 1 fM to 1 .mu.M in the presence 
of .mu.M DADLE or morphine (FIG. 4). On the other hand, after washout of 
the chronic morphine, acute application of etorphine to chronic .mu.M 
opioid-treated DRG neurons no longer showed greater inhibitory 
effectiveness as compared to tests on naive cells (n=10) (FIG. 4). These 
results suggest that the apparent enhancement in inhibitory effectiveness 
of etorphine (and dihydroetorphine), when tested during chronic exposure 
to bimodally-acting opioid-treated DRG neurons, is due to their dual 
synergistic action as agonists at inhibitory opioid receptors and 
antagonists at excitatory opioid receptors. The latter property results in 
unmasking of the inhibitory actions of the chronic DADLE or morphine, as 
occurs in similar tests on naive neurons (see FIG. 1C and FIG. 3). Acute 
application of dihydroetorphine to chronic .mu.M morphine-treated DRG 
neurons indicated that this opioid showed even greater inhibitory potency 
than etorphine. fM concentrations shortened the APD in 80% of the treated 
cells (n=10) and pM (or higher) levels were effective on all cells tested 
in the presence of the chronic opioid (n=10). 
Thus, etorphine and dihydroetorphine show similarly remarkable 
effectiveness as diprenorphine in antagonizing excitatory opioid receptors 
even when tested in the presence of 10.sup.6 -10.sup.9 higher 
concentrations of morphine or DADLE. As a result of these unusual 
properties, etorphine and dihydroetorphine showed no cross-tolerance in 
tests on chronic DADLE- or morphine-treated DRG neurons, just as chronic 
morphine-treated mice showed no cross-tolerance to etorphine even when the 
analgesic ED for morphine had increased 15-fold (see Lange et al., 
Toxicol. Applied Pharmacol., Vol. 54, pp. 177-186 (1980)). The absence of 
cross-tolerance to etorphine in chronic morphine-treated DRG neurons is in 
sharp contrast to the attenuated inhibitory effects (i.e., tolerance) and 
the enhanced excitatory effects (i.e., "dependence") displayed by all 
bimodally-acting mu, delta and kappa opioid agonists when tested acutely 
on these chronic opioid-treated cells. 
EXAMPLE 4 
Etorphine Or Dihydroetorphine Can Block Naloxone-Precipitated 
Supersensitive Excitatory Effects In Chronic Opioid-Treated DRG Neurons 
Drug tests: Mouse DRG-cord explants, grown for &gt;3 weeks as described in 
Example 1, were chronically exposed to the bimodally-acting 
(excitatory/inhibitory) delta/mu opioid agonist, DADLE (1 .mu.M or 
morphine (1 .mu.M) for 1 week or longer. The excitatory opioid 
supersensitivity of chronic opioid-treated DRG neurons was precipitated by 
naloxone (1 nM). Electrophysiological recordings were made as in Example 
1. 
Results: The opioid antagonist, naloxone (1 nM-1 .mu.M) does not alter the 
APD of naive DRG neurons. In contrast, after chronic opioid treatment (as 
well as after acute GM1 ganglioside treatment) excitatory opioid receptor 
functions become so supersensitive that acute application of low 
concentrations of naloxone prolonged the APD of the treated sensory 
neurons, presumably due to weak partial agonist properties of naloxone at 
excitatory opioid receptors (see Crain and Shen, Brain Res., Vol. 575, pp. 
13-24 (1992) and Crain and Shen, J. Pharmacol. Exp. Ther., Vol. 260, pp. 
182-186 (1992)). Naloxone (1 nM) elicited excitatory APD-prolonging 
effects in 92% of the chronic .mu.M DADLE- or morphine-treated DRG neurons 
tested in the present study (n=12) (see FIG. 5). 
These results provide a novel cellular model to account for 
naloxone-precipitated withdrawal supersensitivity in opiate addicts in 
vivo. It should be emphasized that naloxone was not simply blocking the 
inhibitory effect of residual DADLE or morphine since the treated DRG 
neurons were already tolerant to these APD-shortening effects. It is 
therefore of great interest that acute application of remarkably low 
concentrations of etorphine (fM-nM) to chronic .mu.M morphine or 
DADLE-treated cells could effectively block naloxone-induced APD 
prolongation in all of the treated DRG neurons (n=18) (see FIG. 5), 
thereby mimicking the potent effects of the related etorphine analog 
dihydroetorphine in suppressing naloxone-evoked withdrawal symptoms in 
opiate addicts (see Wang et al., Chinese J. Pharmacol. Toxicol., Vol. 6, 
pp. 36-40 (1992)). 
FIG. 5 shows that after chronic exposure to morphine (Mor) or other 
bimodally-acting opioids, acute application of low concentrations of 
etorphine (Et) can dose-dependently block the excitatory APD-prolonging 
effects of naloxone (NLX) on these supersensitive DRG neurons. Histogram 
shows that acute application of 1 nM NLX to DRG neurons chronically 
treated with 3 .mu.M Mor for 1-4 weeks prolonged the APD by about 50%, 
tested in the presence of 3 .mu.M Mor. See also Inset: records 1 and 2. In 
contrast, NLX (nM-.mu.M) does not alter the APD of naive DRG neurons. 
Sequential co-perfusions with 1 fM to 1 .mu.M Et elicit dose-dependent 
attenuation of the NLX-induced APD prolongation in these treated neurons, 
resulting in shortening of the APD to about 70% of the control value in 1 
.mu.M Et. In contrast, NLX-pretreatment of naive DRG neurons blocks 
Et-induced APD shortening (see FIG. 1B). Inset records 1 and 2 show that 1 
nM NLX prolongs the APD of a DRG neuron after chronic 3 .mu.M Mor 
treatment for 2 weeks and tested in the presence of 3 .mu.M Mor (5 minute 
test). Inset record 3 shows that acute addition of 1 fM Et attenuates the 
NLX-induced APD prolongation (5 minute test). Inset record 4 shows that 
increasing the concentration of Et to 1 pM almost completely blocks the 
NLX-induced APD prolongation. Inset records 5 and 6 show that sequential 
application of 1 nM and 1 .mu.M Et in the presence of Mor and NLX results 
in progressive shortening of the APD well below the initial magnitude in 
chronic Mor (as shown in FIG. 4). 
These results suggest that the potent dose-dependent effects of etorphine 
and dihydroetorphine in blocking naloxone-evoked APD prolongation in 
chronic opioid-treated DRG neurons in vitro and withdrawal syndromes in 
opiate addicts in vivo is due to the strong antagonist actions of these 
opioids at supersensitive excitatory opioid receptors which have become 
responsive to the weak agonist properties of naloxone. This is in sharp 
contrast to the blockade of etorphine-induced APD shortening in naive DRG 
neurons by naloxone under conditions where it acts primarily as an 
antagonist at inhibitory opioid receptors (see FIG. 5 and FIG. 1B). 
EXAMPLE 5 
In The Presence Of Etorphine, Chronic Morphine Treatment Of DRG Neurons No 
Longer Results In Development Of Opioid Excitatory Supersensitivity And 
Tolerance 
Drug tests: Mouse DRG-cord explants, grown for &gt;3 weeks as described in 
Example 1, were chronically exposed to bimodally-acting opioid agonist, 
morphine (1 .mu.M) and opioid excitatory receptor antagonist, etorphine (1 
pM) for &gt;1 week and tested for opioid excitatory supersensitivity of DRG 
neurons at low concentrations of naloxone or dynorphin A (1-13) and 
tolerance to the opioid inhibitory effects of APD with high concentrations 
of morphine. Electrophysiological recordings were made as in Example 1. 
Results: Co-administration of low (pM) concentrations of etorphine during 
chronic treatment of DRG neurons with .mu.M levels of morphine was 
remarkably effective in preventing development of the opioid excitatory 
supersensitivity and tolerance that generally occurs after sustained 
exposure to bimodally-acting opioids. Acute application of 1 fM dynorphin 
(1-13) (n=10) or 10 nM naloxone (n=8) to DRG neurons chronically exposed 
to 3 .mu.M morphine together with 1 pM etorphine (for &gt;1 week) did not 
evoke the usual excitatory APD prolongation observed in chronic 
morphine-treated cells, even when tested up to 6 hours after return to 
BSS. Furthermore, there was little or no evidence of tolerance to the 
inhibitory effects of .mu.M morphine: 6 out of 10 cells still showed APD 
shortening following acute application of .mu.M morphine similar to tests 
on naive DRG cells. If etorphine was acting simply as an agonist at 
inhibitory opioid receptors, one might predict that the addition of 1 pM 
etorphine together with a 10.sup.6 -fold higher concentration of morphine 
would have a negligible effect on chronic morphine-treated DRG neurons or 
would augment development of cellular signs of dependence. On the other 
hand, the results are readily accounted for by the potent antagonist 
action of etorphine at excitatory opioid receptors during chronic morphine 
treatment, thereby preventing development of opioid excitatory 
supersensitivity and tolerance, just as occurs during chronic opioid 
treatment of DRG neurons in the presence of cholera toxin-B subunit (see 
Shen and Crain, Brain Res., Vol. 597, pp. 74-83 (1992)). This toxic 
subunit selectively interferes with GM1 ganglioside regulation of 
excitatory opioid receptor functions (see Shen and Crain, Brain Res., Vol. 
531, pp. 1-7 (1990) and Shen et al., Brain Res., Vol. 559, pp. 130-138 
(1991)). 
EXAMPLE 6 
Chronic Etorphine-Treated DRG Neurons Do Not Develop Opioid Excitatory 
Supersensitivity Or Tolerance 
Drug tests: Mouse DRG-cord explants, grown for &gt;3 weeks as described in 
Example 1, were chronically exposed to etorphine (nM) for &gt;1 week and 
tested for opioid excitatory supersensitivity of DRG neurons to low 
concentrations of naloxone or dynorphin A (1-13) and tolerance to the 
inhibitory effects of higher concentrations of etorphine. 
Results: Chronic treatment of DRG neurons with etorphine alone even at a 
relatively high concentration (10 nM) for &gt;1 week in culture also did not 
result in opioid excitatory supersensitivity when tested acutely with 1 fM 
dynorphin (1-13) (26 out of 28 cells) or 1-10 nM naloxone (13 out of 14 
cells), either before or after withdrawal of the chronic etorphine (FIG. 
6). 
FIG. 6 shows that after chronic exposure to nM etorphine (Et), DRG neurons 
do not become supersensitive to the excitatory effects of dynorphin 1-13 
(Dyn) and naloxone (NLX), nor do they develop tolerance to the inhibitory 
effects of etorphine or other opioid agonists. Record 1 shows the action 
potential (AP) generated by a DRG neuron after chronic 10 nM Et treatment 
for 20 days and tested in balanced salt solution (BSS) shortly after 
washout of the Et. Record 2 shows that APD is not altered by 1 fM Dyn. 
Records 3-6 show that APD is not altered by bath perfusion with 1 fM Et 
but is progressively shortened by 1 pM and 1 .mu.M concentrations (5 
minute test periods). Records 7 and 8 show that APD of the same neuron is 
not altered by application of 10 nM NLX. Record 9 shows that APD is no 
longer shortened by 1 .mu.M Et when co-perfused with 10 nM naloxone (5 
minute test period). 
After etorphine withdrawal for about 1 hour, 1 nM dynorphin (1-13) 
shortened the APD in 3 cells or did not show typical APD prolongation (10 
out of 11 cells), resembling the attenuation of opioid excitatory effects 
and unmasking of opioid inhibition observed in acute tests on naive cells 
after washout of low (pM) concentrations of etorphine (see FIG. 1C). 
Furthermore, after washout of the chronic 10 nM etorphine, acute 
application of pM, nM and .mu.M etorphine elicited similar APD shortening 
(in 3 out of 4 cells tested at each concentration) as observed in naive 
cells. Thus chronic 10 nM etorphine treatment of DRG neurons did not 
result in the characteristic cellular signs of physical dependence and 
tolerance that occurred after chronic exposure of these cells to morphine 
and other bimodally-acting opioids. 
Estimates Of Specific In Vivo Dosages Of Excitatory Opioid Receptor 
Antagonists That May Enhance Analgesic Potency And Reduce Dependence 
Liability (And Other Side-Effects) Of Morphine Or Other Conventional 
Opioid Analgesics When Administered In Combination 
Electrophysiological studies on DRG neurons indicated that pre-treatment 
with low fM-pM concentrations of etorphine, dihydroetorphine and 
diprenorphine are remarkably effective in blocking excitatory 
APD-prolonging effects of morphine bimodally-acting opioid agonists by 
selective antagonist actions at mu, delta and kappa excitatory opioid 
receptors. The potency of these three excitatory opioid receptor 
antagonists is clearly shown by their ability to unmask inhibitory opioid 
receptor-mediated APD-shortening effects, even in presence of 10.sup.6 
-fold higher concentrations of morphine other bimodally-acting opioid 
agonists (FIGS. 3-5). 
In the presence of these selective excitatory opioid receptor antagonists, 
morphine and other clinically used opioids showed markedly increased 
potency in evoking the inhibitory effects on the action potential of 
sensory neurons which are generally considered to underly opioid analgesic 
action in vivo. These bimodally-acting opioid agonists became effective in 
shortening, instead of prolonging, the APD at pM-nM (i.e., 10.sup.-12 
-10.sup.-9 M) concentrations, whereas 0.1-1 pM (i.e., 10.sup.-7 -10.sup.-6 
M) levels were generally required to shorten the APD (FIG. 2). Selective 
blockade of the excitatory side-effects of these bimodally-acting opioid 
agonists eliminates the attenuation of their inhibitory effectiveness that 
would otherwise occur. Hence, according to this invention, the combined 
use of a relatively low dose of one of these selective excitatory opioid 
receptor antagonists, together with morphine or other bimodally-acting mu, 
delta or kappa opioid agonists, will markedly enhance the analgesic 
potency of said opioid agonist, and render said opioid agonist comparable 
in potency to etorphine or dihydroetorphine, which, when used alone at 
higher doses, are &gt;1000 times more potent than morphine in eliciting 
analgesia. 
Co-administration of one of these excitatory opioid receptor antagonists at 
low concentration (about 10.sup.-12 M) during chronic treatment of sensory 
neurons with 10.sup.-6 M morphine or other bimodally-acting opioids (&gt;1 
week in culture) prevented development of the opioid excitatory 
supersensitivity, including naloxone-precipitated APD-prolongation, as 
well as the tolerance to opioid inhibitory effects that generally occurs 
after chronic opioid exposure. This experimental paradigm was previously 
utilized by the inventors on sensory neurons in culture to demonstrate 
that co-administration of 10.sup.-7 M cholera toxin-B subunit, which binds 
selectively to GM1 ganglioside and thereby blocks excitatory GM1-regulated 
opioid receptor-mediated effects, but not opioid inhibitory effects (see 
Shen and Crain, Brain Res., Vol. 531, pp. 1-7 (1990)) during chronic 
opioid treatment prevents development of these plastic changes in neuronal 
sensitivity that are considered to be cellular manifestations related to 
opioid dependence/addiction and tolerance in vivo (see Shen and Crain, 
Brain Res., Vol. 597, pp. 74-83 (1992)). Hence, according to this 
invention, the sustained use of a relatively low clinical dose of one of 
these selective excitatory opioid receptor antagonists, e.g., &lt;1 microgram 
of etorphine, dihydroetorphine or diprenorphine, in combination with 
10-100 micrograms of morphine or other conventional bimodally-acting 
opioid analgesics will result in analgesia comparable to that elicited by 
said analgesics when administered alone in milligram doses and will 
attenuate or even prevent development of physical dependence and other 
undesirable excitatory side-effects generally associated with said 
analgesics. 
Estimates Of Specific In Vivo Dosages That Provide Improved Detoxification 
And Treatment Of Opiate Addicts 
Acute application of one of these excitatory opioid receptor antagonists, 
e.g., etorphine or dihydroetorphine, at a low concentration (about 
10.sup.-12 M) to chronic opioid-treated sensory neurons prevented the 
excitatory APD-prolonging effects precipitated by naloxone (10.sup.-9 M) 
(FIG. 5). The latter effects provide a novel cellular model to account for 
naloxone-evoked withdrawal supersensitivity in opiate addicts in vivo. The 
potent effects of etorphine and dihydroetorphine in blocking 
naloxone-evoked APD prolongation in chronic opioid-treated sensory neurons 
in vitro (FIG. 5) is due to their blockade of supersensitive excitatory 
opioid receptors which have become responsive to the weak agonist 
properties of naloxone. A similar mechanism may account for the efficacy 
of dihydroetorphine in suppressing naloxone-evoked withdrawal syndromes in 
opiate-addicted animals and humans (see Lange, Toxicol. Applied 
Pharmacol., Vol. 54, pp. 177-186 (1992) and Qin, Chinese J. Pharmacol. 
Toxicol., Vol. 6, (1992)). Hence, according to this invention, 
appropriately low doses of one of these selective excitatory opioid 
receptor antagonists, e.g., about 1 microgram in combination with a 
10-1000 fold lower than standard dose of one of the opioids currently used 
for the treatment of opioid dependence, e.g., methadone, buprenorphine, 
will provide an improved method for detoxifying and weaning addicts from 
dependence on opiates. 
Although the invention herein has been described with reference to 
particular embodiments, it is to be understood that these embodiments are 
merely illustrative of various aspects of the invention. Thus, it is to be 
understood that numerous modifications may be made in the illustrative 
embodiments and other arrangements may be devised without departing from 
the spirit and scope of the invention.