Edward E. Smissman-Bristol-Myers Squibb Award Address. The role of concepts in structure-activity relationship studies of opioid ligands.

Various concepts have served as the basis for the development of models to explore the relationship between molecular structure and biological activity. In this presentation I have outlined five concepts that have been useful in our investigation of opioid receptor multiplicity and in the design of selective opioid receptor antagonists. The first of these, the multiple binding modality concept, led to our application of four other concepts in the development of opioid receptor probes. Some of these probes are now standard tools in opioid research. These include the mu-selective affinity label beta-FNA (10), the kappa opioid receptor antagonist norBNI (15), and the delta opioid antagonist NTI (20). These highly selective antagonists have advantages over the universal opioid antagonists naloxone and naltrexone because they are of value in probing the interaction of endogenous opioid peptides with opioid receptor types. Additionally, they are useful in evaluating the selectivity of new opioid agonists. Also, selective opioid antagonists have potential clinical applications in the treatment of a variety of disorders where endogenous opioids play a modulatory role. These include constipation, immune function, drug addiction, and alcoholism, to name only a few.     

Naloxone attenuates voluntary ethanol intake in rats selectively bred for high ethanol preference

The effect of naloxone on voluntary ethanol intake was examined in rats which were selectively bred for oral ethanol preference (High Alcohol Drinking or HAD line). Rats of the HAD line were treated with naloxone in doses of 0.05-18.0 mg/kg b.wt. before access to water alone or to a free-choice between a 10% (v/v) ethanol solution and water. Naloxone suppressed water intake when water was presented as the sole source of fluid. In contrast, naloxone produced a dose-dependent decrease in ethanol consumption, without altering water intake, when rats were given a free-choice between the ethanol solution and water. Selective suppression of ethanol consumption by naloxone was not attributable to changes in blood ethanol concentrations or ethanol elimination rates following naloxone treatment. It appears that although naloxone may attenuate the positively reinforcing properties of both ethanol and water, ethanol drinking is a subset of consummatory behaviors that is particularly sensitive to opioid receptor blockade. The results suggest that activation of the endogenous opioid system may be an important mechanism which serves to maintain continued ethanol drinking.     

Effects of continuous opioid receptor blockade on alcohol intake and up-regulation of opioid receptor subtype signalling in a genetic model of high alcohol drinking

Effects of a continuous naloxone infusion via osmotic pumps on alcohol drinking and opioid receptor density and function in the high-drinking AA (Alko, Alcohol) rats were examined. AA rats were trained to drink 10% (v/v) ethanol in a 1-h limited access procedure and implanted with subcutaneous osmotic pumps delivering either saline, a low dose (0.3 mg/kg per hour), or a high dose (3.0 mg/kg per hour) of naloxone for 7 days. The pumps were then removed and alcohol, food and water intakes were measured for another 4 days. Compared with saline, both naloxone doses significantly suppressed 1-h alcohol intake during the 7-day infusion. The suppression was smaller than that by a bolus injection of the same daily dose 15 min before the session, although a complete blockade of morphine-induced antinociception was achieved even with the smaller naloxone infusion. Significant decreases were also seen in daily food and water intake during the first days, but they quickly returned to their previous baselines. After pump removal, rats of both naloxone-treated groups rapidly increased their alcohol drinking and reached the pretreatment baseline, while their food and water intakes significantly surpassed their baselines. Naloxone infusion at 3.0 mg/kg per hour for 7 days significantly decreased 24-h alcohol drinking without affecting alcohol preference. Twenty-four hours after pump removal, autoradiography with [3H]DAMGO, [3H]DPDPE and [3H]U-69,543 revealed an up-regulation of mu-, delta- and kappa-opioid receptor binding sites in many brain areas of these animals. This receptor up-regulation was functional, because receptor coupling to G-protein activation was enhanced by agonist ligands, as revealed by [35S]GTPgammaS autoradiography. A good correlation existed between ligand binding densities and G-protein activation for mu- and kappa-receptors in control and naloxone-treated brain sections. Furthermore, morphine-induced analgesia in a hot-plate test showed a leftward shift in the morphine dose-response curve after naloxone treatment. These results suggest that the usefulness of a chronic opioid antagonist dosing regime could be limited by nonspecific effects of the antagonist on ingestive behaviour, an up-regulation of opioid receptors with high antagonist doses, and the resulting supersensitivity to opioid agonists after the discontinuation of the treatment.     

Effects of naloxone and naltrexone on the increased water intake and drinking duration in phenobarbitone-treated rats

In two experiments, phenobarbitone sodium (30 mg/kg) reliably enhanced water consumption and extended the duration of drinking in 24h water-deprived male rats. The opiate receptor antagonists naloxone (0.1-10 mg/kg) and naltrexone (0.1-10 mg/kg) both decreased water intake and reduced the duration of drinking. When the barbiturate was given in conjunction with either naloxone or naltrexone, phenobarbitone and the opiate antagonist exerted opposite effects on the two measures of drinking. While it was true that both opiate antagonists reduced water intake and drinking duration in barbiturate-treated animals, the barbiturate-induced enhancement of drinking was in no way modified by concurrent opiate antagonist treatment. Hence, the effects of phenobarbitone and of the two opiate antagonists upon the drinking measures appeared to be quite independent. There was no evidence, therefore, that the effects of phenobarbitone upon drinking were related to endogenous opioid mechanisms. The possible contrast between benzodiazepine- and barbiturate-induced hyperdipsia is briefly considered in the light of these results.     

Experience with alcohol and the endogenous opioid system in ethanol analgesia

This study employed naloxone, an opiate antagonist, to explore whether a learned opioid response, mediated by drinking experience, accounts for ethanol and placebo analgesia. Cold pressor pain was evaluated before and after ethanol (0.5 g/kg), placebo, and no-alcohol control treatments (administered in randomized order) and again after double-blind administration (6 mg/kg) of naloxone to 11 men and saline to 9. A triple interaction of treatments, antagonist conditions, and drinking experience indicated that naloxone as compared to saline diminished ethanol and placebo analgesia among experienced drinkers but had opposite effects among the same men in the control treatment. Six men, who reported that the injection of naloxone had an effect on pain, had higher drinking experience scores than the five men who reported naloxone had no effect. The similar pattern of response to both the alcohol and the placebo treatments suggests that the opioid system response to alcohol is learned.     

Effects of delta opioid antagonists on enkephalin-induced seizures

We examined the effect of opioid receptor antagonists on the seizure phenomena induced by specific delta opioid receptor agonist [D-Ser2,Leu5] enkephalyl-Thr (DSLET). The experiments have been performed in the anesthetized rats, and the DSLET-induced seizure phenomena were registered by electrocorticogram and electromyogram. It was demonstrated that two selective delta opioid receptor antagonists, ICI 152,129 and ICI 174,864 inhibited DSLET-induced epileptiform ECoG pattern and myoclonic contractions in a dose-related manner. An equimolar concentration of naloxone failed to antagonize the epileptiform effects of DSLET. It is concluded that delta opioid receptor agonist-induced seizure is mediated by delta receptors, since it can be blocked by delta opioid receptor antagonists. Evidently, delta opioid antagonists can be used as a good tool, in order to demonstrate a delta component in the seizure phenomena induced by other endogenous opioid peptides or their derivatives.     

Lack of renal tubular and hemodynamic effects of non-selective and delta-opioid receptor antagonism

The renal pharmacological actions of the non-selective opioid receptor antagonist naloxone and the selective delta (delta)-opioid receptor antagonist ICI 154,129 were examined in conscious dogs. Neither naloxone nor ICI 154,129 altered glomerular filtration rate, renal blood flow, blood pressure, heart rate, or renal excretion of water, Na+, K+, or Cl-. In addition, urine and plasma osmolality and electrolyte concentrations and hematocrit were unchanged, suggesting that neither agent produced physiologically significant alteration in plasma vasopressin levels. These data suggest that (a) naloxone and ICI 154,129 exert no renal pharmacological effects in dogs and (b) under resting physiological conditions, delta-opioid receptors, as well as other opioid receptor subtypes, probably are not involved in the tonic regulation of renal hemodynamics or tubular function.     

Effect of opioid receptor antagonists on vasodilator nerve actions in the perfused rat mesentery.

Our previous work suggests that opioid peptides modulate sensory nerves in the perfused rat mesentery. Therefore we tested the hypothesis that opioids are involved in the ongoing regulation of sensory nerve activity using selective opioid receptor antagonists. In the presence of guanethidine and methoxamine, transmural nerve stimulation caused a vasodilator response which was potentiated significantly by naloxone (3 x 10(-7) M). However, naloxone did not affect vasodilator responses to exogenous calcitonin gene related peptide. IC1 174.864 (3 x 10(-7) M), a selective delta receptor antagonist, had no effect on vasodilator responses to transmural nerve stimulation. In contrast CTOP (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Phe-Thr-NH2) (3 x 10(-7) M), a selective mu receptor antagonist, significantly inhibited vasodilator responses to transmural nerve stimulation, effects which were abolished by naloxone treatment. In preparations pretreated with beta-FNA (beta-funaltrexamine HCl), an irreversible mu receptor antagonist, naloxone no longer potentiated vasodilator responses to transmural nerve stimulation. These results suggest that potentiation of vasodilator responses to transmural nerve stimulation by naloxone may be due to blockade of mu receptors, resulting in a reduced inhibitory modulation by endogenous opioids. These findings support the contention that prejunctional opioid receptors on sensory nerves may play a role in modulating activity of the cardiovascular system.     

Endocannabinoid system and alcohol addiction: pharmacological studies

The present paper describes the results of recent pharmacological studies implicating the cannabinoid CB1 receptor in the neural circuitry regulating alcohol consumption and motivation to consume alcohol. Cannabinoid CB1 receptor agonists have been found to specifically stimulate alcohol intake and alcohol's motivational properties in rats. Conversely, the cannabinoid CB1 receptor antagonist, SR 141716, has been reported to specifically suppress acquisition and maintenance of alcohol drinking behavior, relapse-like drinking and alcohol's motivational properties in rats. More recent data indicate that opioid receptor antagonists a) blocked the stimulatory effect of cannabinoids on alcohol intake, and b) synergistically potentiated the suppressing effect of SR 141716 on alcohol intake and alcohol's motivational properties. Consistently, SR 141716 blocked the stimulatory effect of morphine on alcohol intake. These results suggest a) the existence of a functional link between the cannabinoid and opioid receptor systems in the control of alcohol intake and motivation to consume alcohol, and b) that novel and potentially effective therapeutic strategies for alcoholism may come from the combination of cannabinoid and opioid receptor antagonists.     

Naloxone modulates body and organ growth of rats: dependency on the duration of opioid receptor blockade and stereospecificity

Endogenous opioid systems (i.e., endogenous opioids and opioid receptors) act in the trophic regulation of biological development. Opioid antagonist paradigms have served to elucidate the nature of this relationship. If growth is mediated at the level of the opioid receptor, one would expect that this interaction would be stereospecific. This study shows that daily injections of 20-60 mg/kg (-) naloxone, given chronically throughout the preweaning period, depress body weight when monitored at Day 21. Opioid challenge experiments using nociceptive measures show that these dosages of (-) naloxone invoked an opioid receptor blockade for no more than 10-12 hr/day. A dosage of 100 mg/kg (-) naloxone, which blocked the opioid receptor for 12-16 hr/day, did not alter body weight in comparison to control levels. In subsequent experiments, 40 mg/kg (-) naloxone depressed body weight of 21-day-old rats, and the wet weights of the liver, spleen, thymus, heart, and triceps surae muscle from these animals were subnormal. A dosage of 40 mg/kg (+) naloxone did not alter growth. These results show that opioid action in regard to growth is stereospecific and dependent on the duration of opioid receptor blockade, providing additional evidence that endogenous opioid systems play an important role in developmental events.     

Differential involvement of endogenous opioids in sucrose consumption and food reinforcement

Endogenous opioids within the central nervous system are postulated to mediate hedonic aspects of feeding behavior. To identify the relevant endogenous opioid receptor ligands, mice lacking one or two of the opioid peptide families beta-endorphin, enkephalins or dynorphins were tested for sucrose preference in a two-bottle free-choice drinking paradigm under drug-na?ve conditions and following treatment with an opioid antagonist (1 mg/kg naloxone i.p.) or saline. Basal sucrose consumption was unaltered in all of the knockout genotypes compared to their congenic wild-type C57BL/6 littermates during 0.5 and 6 h access to a bottle containing 2, 4, 8, or 16% sucrose and a second bottle containing water. Moreover, all mutant genotypes and wildtype mice exhibited a similar compensatory decrease in overnight food intake following the extra caloric load from 6 h sucrose access. Although these basal responses to sucrose were unaffected by the knockout genotypes, naloxone reduced sucrose consumption by 50% compared to saline treatment during the first 0.5 h in wild-type and beta-endorphin knockout mice, but had no effect in enkephalin knockouts, beta-endorphin and enkephalin double knockouts, or dynorphin knockouts. These data suggest that naloxone reduces sucrose consumption in wild-type mice by blocking endogenous enkephalin and/or dynorphin signaling, but not beta-endorphin. Dynorphin knockouts in the current study had bar-pressing responses for a palatable food reinforcer in an operant procedure under free-feeding conditions similar to wild-type mice while we found in a previous study that beta-endorphin and enkephalin knockout mice had reduced motivation to respond [Hayward MD, Pintar JE, Low MJ. Selective reward deficit in mice lacking beta-endorphin and enkephalin. J Neurosci 2002;22:8251-8258.]. We conclude from these studies directly comparing three strains of opioid peptide knockout mice that enkephalin and dynorphin can modulate sucrose preference but are not necessary to support sucrose consumption. However, dynorphin was not necessary to support wildtype levels of operant responding suggesting that only enkephalin and beta-endorphin modulate conditioned food reinforcement.     

Has the sun set on kappa3-opioid receptors?

Mu-opioid receptor agonists are a mainstay of clinical analgesia, despite the significant unwanted effects and dependence liability associated with drugs like morphine. The quest for opioids that produce analgesia with fewer undesirable effects has lead to the putative identification of multiple opioid receptor subtypes, despite the identification of only four opioid-related receptor genes. One such putative receptor subtype is the kappa3 receptor, activation of which supposedly produces analgesia in animals. In the present issue of this Journal, Olianas and co-workers have demonstrated that the prototypic kappa3 agonist naloxone benzoylhydrazone is actually a partial agonist at the cloned mu, delta, and kappa opioid receptors and an antagonist at opioid-like NOP receptors. Together with a recent study that showed that high-affinity naloxone benzoylhydrazone binding is abolished in triple mu/delta/kappa receptor knockout mice, the present study provides strong evidence that in vivo effects attributed to kappa3 receptor activation probably just reflect the combined actions of a particularly nonselective opioid drug. Indeed, molecular identification of any of the proposed subtypes of mu, delta, and kappa opioid receptors has proven elusive, suggesting that it is perhaps time to retire the notion of opioid receptor subtypes until definitive evidence for their existence is provided.     

Involvement of different opioid receptor subtypes in electric shock-induced analgesia and motor suppression in the rat

We have investigated the correlation of electric shock-induced behavioral changes and functional alterations of endogenous opioid receptor subtypes. The degree of electric shock-induced behavioral changes, such as analgesia and motor suppression, was dependent on the duration of and time after electric shock application. The electric shock-induced behavioral changes were completely antagonized by naloxone. The apparent development of tolerance to both behavioral effects as a result of successive daily electric shock was different: Tolerance to electric shock-induced analgesia developed after 2 days' successive electric shock application, while tolerance to motor suppression was not observed even after 7 days' successive electric shock application. There was a decrease of [3H][D-Ala2, Met5]enkephalinamide ([3H]DAMEA, delta agonist) binding and an increase of [3H]naloxone (mu antagonist) binding when potent electric shock-induced analgesia developed. On the other hand, the binding of [3H]DAMEA and [3H]ethylketocyclazocine (kappa agonist) was significantly changed when locomotion was suppressed. These results suggest strongly that different opioid systems may participate in electric shock-induced analgesia and motor suppression: electric shock-induced analgesia and motor suppression may be mediated by mu/delta and kappa/delta receptors, respectively.     

Centrally administered opioid antagonists, nor-binaltorphimine, 16-methyl cyprenorphine and MR2266, suppress intake of a sweet solution

Three opioid antagonists (MR2266, 16-methyl cyprenorphine and nor-binaltorphimine) were tested independently for their ability to suppress the intake of a highly palatable saccharin and glucose (S/G) solution after central administration. MR2266 is an equally potent antagonist at kappa (kappa) and mu (mu) opioid receptors. Nor-binaltorphimine (N-BNI) and 16-methyl cyprenorphine (M80) are two recently developed opioid antagonists that were chosen based upon their ability to act more selectively than naloxone at kappa and delta (delta) opioid receptor types, respectively. Prior research has demonstrated that when dissolved in acid and administered centrally, MR2266 (20 micrograms) fails to suppress S/G intake. Because all three antagonists are rather insoluble in water, they were dissolved in dimethyl sulfoxide (DMSO). Rats with chronic ventricular cannula were allowed to consume S/G for a 0.5 hr bout. They received a single intracerebroventricular (ICV) injection of antagonist (MR2266: 0, 10, 20 and 40 micrograms; M80: 0, 5, 10, 20 and 40 micrograms or N-BNI: 0, 1, 3, and 10 micrograms) 10 min prior to the start of the drinking bout. Administration of DMSO alone failed to alter drinking relative to saline, whereas each antagonist significantly attenuated S/G intake. We conclude that, when dissolved in DMSO, these antagonists suppress drinking by blockade of opioid receptors.     

Endogenous opioid mechanisms partially mediate P2X3/P2X2/3-related antinociception in rat models of inflammatory and chemogenic pain but not neuropathic pain

P2X3/P2X2/3 receptors have emerged as important components of nociception. However, there is limited information regarding the neurochemical systems that are affected by antagonism of the P2X3/P2X2/3 receptor and that ultimately contribute to the ensuing antinociception. In order to determine if the endogenous opioid system is involved in this antinociception, naloxone was administered just prior to the injection of a selective P2X3/P2X2/3 receptor antagonist, A-317491, in rat models of neuropathic, chemogenic, and inflammatory pain. Naloxone (1-10 mg kg(-1), i.p.), dose-dependently reduced the antinociceptive effects of A-317491 (1-300 micromol kg(-1), s.c.) in the CFA model of thermal hyperalgesia and the formalin model of chemogenic pain (2nd phase), but not in the L5-L6 spinal nerve ligation model of neuropathic allodynia. In comparison experiments, the same doses of naloxone blocked or attenuated the actions of morphine (2 or 8 mg kg(-1), s.c.) in each of these behavioral models. Injection of a peripheral opioid antagonist, naloxone methiodide (10 mg kg(-1), i.p.), did not affect A-317491-induced antinociception in the CFA and formalin assays, suggesting that the opioid component of this antinociception occurred within the CNS. Furthermore, this utilization of the central opioid system could be initiated by antagonism of spinal P2X3/P2X2/3 receptors since the antinociceptive actions of intrathecally delivered A-317491 (30 nmol) in the formalin model were reduced by both intrathecally (10-50 nmol) and systemically (10 mg kg(-1), i.p.) administered naloxone. This utilization of the opioid system was not specific to A-317491 since suramin-, a nonselective P2X receptor antagonist, induced antinociception was also attenuated by naloxone. In in vitro studies, A-317491 (3-100 microM) did not produce any agonist response at delta opioid receptors expressed in NG108-15 cells. A-317491 had been previously shown to be inactive at the kappa and mu opioid receptors. Furthermore, naloxone, at concentrations up to 1 mM, did not compete for [3H] A-317491 binding in 1321N1 cells expressing human P2X3 receptors. Taken together, these results indicate that antagonism of spinal P2X3/P2X2/3 receptors results in an indirect activation of the opioid system to alleviate inflammatory hyperalgesia and chemogenic nociception.     

Electric footshock-induced changes in behavior and opioid receptor function

The present electric shock (ES) schedule produced significant behavioral changes, such as analgesia and motor suppression, and functional changes in binding capacities for opioid agonist and antagonist. In the naloxone (5 mg/kg, SC 15 min before ES application) pretreated rats, these behavioral and biochemical changes were blocked. In addition, when preincubation (37 degrees C, 30 min) was not carried out in the process of preparation of synaptic membrane, the ES-induced functional changes in hibh affinity binding sites were not observed. Moreover, the present data indicated that preincubation may produce the destruction of [3H]-D-ala2,L-met5-enkephalinamide ([3H]-DAMEA) specific binding sites with the forced dissociation of endogenous delta-type opioid peptides from delta opioid receptors. In addition, the significant decrease of [3H]-DAMEA specific binding in the ES membrane suggested that delta-type opioid peptides were released more than steady state level by ES application and bound to the delta opioid receptors. Therefore, these results suggest that ES-induced behavioral and biochemical changes were mediated by opioid peptides which were released by ES application. In addition, the ES-induced analgesia may be mediated by high affinity delta opioid receptor.     

Naloxone alters alcohol drinking induced in the rat by tetrahydropapaveroline (THP) infused ICV

In male rats of the Sprague-Dawley or Long-Evans strain, intracerebroventricular (ICV) cannulae were implanted permanently using stereotaxic techniques. Tetrahydropapaveroline (THP) was infused ICV for up to 14 days either chronically around the clock or acutely once per day in doses previously found to induce an abnormally high intake of alcohol. During these periods, alcohol preference for individual rats was determined by a self-selection procedure in which the concentration of alcohol was increased from 3 to 30% on each day of a 12-day interval. Those rats which displayed a substantial increase in their intake of alcohol were selected for naloxone treatment and subsequently assigned a fixed concentration of alcohol at which maximum consumption occurred. Naloxone was administered subcutaneously two to six times per day for three consecutive days in total daily doses ranging from 1.5 to 3.0 mg/kg. Each rat served as its own control and was given 0.9% saline isovolumetrically according to the same temporal schedule. Naloxone generally suppressed alcohol intake in animals by 20% to 45%, but the reduction in drinking depended upon the injection regimen as well as the prior level of alcohol consumption. In "high drinking" rats, the mean alcohol intake of 6.2 g/kg/day was reduced to 3.7 g/kg/day by naloxone whereas in "low drinkers" the mean intake of 3.5 g/kg/day was suppressed to 2.8 g/kg/day by the opiate antagonist. These results further support the suggestion of a possible opiate receptor link in the pathogenesis and maintenance of aberrant drinking of alcohol, the mechanism of which may involve the endogenous action of an amine-aldehyde condensation product in the brain.     

Effect of nonselective and selective opioid receptors antagonists on antinociceptive action of acetaminophen [part III

The influence of naloxone (NAL), a competitive antagonist of mu, kappa, delta and sigma receptors; D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH(2) (CTOP), selective antagonist of mu-opioid receptors; nor-binaltorphimine (NOR-BNI), a potent and higly selective kappa opioid receptor antagonist; naltrindole (NTI), a delta-opioid receptor antagonist and naltriben (NTB), a highly selective delta(2)-opioid receptor antagonist on antinociceptive action of acetaminophen (ACETA) was studied in rats. NAL administered intraperitoneally (ip) or intracerebroventricularly (icv), and CTOP and NOR-BNI administered icv, markedly decreased the antinociceptive activity of the high dose of ACETA (400 mg/kg). Pretreatment with NTI (sc), as well as with naloxone (it), and NTB (it) slightly but significantly attenuated the ACETA antinociception. The possible involvement of the opioidergic systems in antinociceptive activity of ACETA is discussed.     

Naloxone effects on sucrose-motivated behavior

The opioid system plays an important role in feeding. In general, opioid agonists typically increase feeding and opioid antagonists decrease feeding in nonfood restricted animals. In food restricted animals the effects of these drugs are substantially reduced. Opioid antagonists have shown a marked effectiveness at reducing consumption of sweet foods. Explanations for this robust effect have typically focused on drug induced changes in taste, taste perception, or palatability. The current study relates the effects of the opioid antagonist naloxone on motivation to obtain different sucrose concentrations to the drug's effects on unrestricted sucrose solution consumption. Changes in motivation to respond were assessed under a progressive ratio reinforcement schedule (PR) which required increased response cost for each successive unit of sucrose solution. Motivation, as measured by the PR, increased as sucrose concentration increased and naloxone produced a dose-dependent decrease in motivation to respond for a given sucrose concentration. Thus, the effectiveness of naloxone was indirectly related to strength of the sucrose concentration. Under unrestricted access to sucrose solutions, naloxone reduced consumption greatest under the higher concentrations. The data suggest at least part of naloxone's effects on sweet tasting food may be mediated through endogenous opioid reward systems that are reflected in measures of motivation.     

Central vs. peripheral mediation of opioid effects on alcohol consumption in free-feeding rats

Although there is considerable evidence that pretreatment with low doses of opioid agonists can enhance, and opioid antagonists can reduce alcohol consumption in rats, little is known about the locus or mechanism of these effects. As a first approximation as to where the effect may occur, we compared the effects of an opioid agonist morphine (1, 3 and 10 mg/kg) and antagonist naltrexone (1, 3 and 10 mg/kg) that are known to act within the brain as well as the periphery, to those of an agonist-like drug loperamide (0.3, 1 and 3 mg/kg) and an antagonist methylnaltrexone (3, 10 and 30 mg/kg) that are known to act peripherally only. Free-feeding rats were initially trained to drink alcohol using a limited access paradigm, and when animals were drinking asymptotic amounts of 12% (w/v) alcohol, increasing doses of one of the four drugs or saline were administered IP to separate groups of rats 30 min prior to the hour-long daily drinking session. The results confirmed that the effects of the opioids on alcohol consumption are indeed mediated within the central nervous system in that morphine enhanced alcohol consumption but loperamide did not, naltrexone reduced alcohol consumption but methylnaltrexone did not, and naltrexone was able to block the morphine effect but methylnaltrexone failed to do so. An unexpected finding was that methylnaltrexone alone also increased alcohol consumption. Possible means by which this could occur, also supporting the idea of a central locus for the effect, as well as possible mechanisms by which opioids could influence alcohol consumption generally, are discussed.