Editors: Spanagel, Rainer; Mann, Karl F.
Title: Drugs for Relapse Prevention of Alcoholism, 1st Edition
Copyright ©2005 Springer
> Table of Contents > Opioidergic compounds: preclinical data
Opioidergic compounds: preclinical data
Department of Mental Health and Alcohol Research, National Public Health Institute, POB 33, FI-00251 Helsinki, Finland
Ethanol-induced activation of the endogenous opioid system has been hypothesized to be one of the mechanisms mediating ethanol reinforcement and enhanced ethanol drinking. This hypothesis is supported by many lines of evidence, including ethanol-induced elevation of extracellular levels of β-endorphin in the nucleus accumbens , and genetically determined differences in the basal levels of β-endorphin, enkephalins, and dynorphins in distinct brain areas of rodent lines that differ in ethanol self-administration behavior [2, 3 and 4]. In accordance with this hypothesis, the non-selective opioid receptor antagonists naltrexone, naloxone and nalmefene suppress ethanol-reinforced behavior in widely different experimental conditions.
Opioid receptors are divided into three major classes, the µ-, δ- and κ-opioid receptors, which have all been cloned and sequenced [5, 6, 7 and 8]. Furthermore, on the basis of pharmacological evidence, subtypes of these receptors have been proposed. β-endorphin recognizes both µ- and δ-receptors with almost equal potency. Also, enkephalins and dynorphins interact with µ-receptors but with a lower affinity as compared to δ- and κ- receptors, respectively . Increasing evidence for the importance of ethanol-induced activation of both β-endorphin and enkephalin systems in ethanol reward has further prompted research on the contribution of the different opioid receptor types, especially the µ- and δ- receptors, in the reinforcing effect of ethanol. Because the commonly used non-selective antagonists (naltrexone, naloxone and nalmefene) bind to all opioid receptor types as a function of the dose administered [10, 11 and 12], the roles of the opioid receptor types in ethanol reinforcement, in recent studies, have been studied with antagonists selective for these receptors.
In the majority of these studies, the selective antagonists have been tested using behavioral models that measure the direct reinforcing effects of ethanol, including various free-choice drinking and operant self-administration models. These models do not permit assessment of the conditioned ethanol effects, which underlie craving and relapse. However, there is evidence showing that the modulation of the reinforcing effects of ethanol by opioid antagonists may
also predict their effects on the appetitive conditioned aspects of ethanol consumption [13, 14].
Effects of selective µ-receptor antagonists on ethanol consumption
Selective µ-opioid receptor antagonists β-funaltrexamine, D-Pen-Cys-Tyr-DTrp-Orn-Thr-Pen-Thr-NH2 (CTOP), and naloxonazine have been shown to reliably suppress ethanol intake in many experimental paradigms. Systemically administered β-funaltrexamine (5-20 mg/kg) produced a dose-dependent decrease in ethanol consumption both in the high alcohol drinking (HAD) rat line on a fluid-deprivation schedule with a 2-h daily access to ethanol and water, as well as in Wistar rats, given limited access 1-h to ethanol with ad libitum water and food [15, 16]. Injections of β-funaltrexamine were administered 16-20 h before the opportunity to drink, because this antagonist has an initial κ-agonist effect lasting 2-3 h, followed by a long (2-4 days) µ-antagonist effect. In both experiments, the highest dose, 20 mg/kg β-funaltrexamine, decreased ethanol intake even during the second post-injection session, probably reflecting the long-lasting µ-antagonist action. Although β-funaltrexamine did not attenuate the 2-h scheduled saccharin intake, decrease in 24-h water consumption suggests that this antagonist displays a general suppressive effect on ingestive behavior .
Another µ-opioid receptor antagonist, CTOP, decreased both limited access ethanol drinking and operant responding for ethanol in alcohol-preferring AA (Alko, Alcohol) and Wistar rats after intracerebroventricular (0.3-3 µg) administration  (Fig. 1). Moreover, systemic administration of this compound
has been reported to suppress ethanol drinking in mice as well . With repeated administration of CTOP across three sessions, a progressive decline in limited access ethanol drinking was observed, with a transient decrease in 24-h water and food intake . It is not clear, however, whether this finding could be interpreted as an extinction-like decrease caused by an antagonist-induced attenuation of ethanol's reinforcing effects or whether it is due to cumulative drug effects. Moreover, the highest dose, 3-µg, had a tendency of slowing the initiation of operant responding for ethanol, suggestive of aversive effects produced by CTOP .
Figure 1. Effects of subcutaneous naloxone, and intracerebroventricular CTOP and naltrindole injections on ethanol consumption in alcohol-preferring AA and heterogeneous Wistar rats. The animals were allowed to respond for a 0.1 ml drop of 10% ethanol solution on a FR1 schedule during daily 30-min sessions. Data are expressed as mean (±SEM) ethanol intake (g/kg) during the 30-min session. (* p < 0.05, significantly different from the vehicle conditions). Adapted with permission of Lippincott Williams & Wilkins, Baltimore, from Hyytiä P & Kiianmaa K (2001) Suppression of ethanol responding by centrally administered CTOP and naltrindole in AA and Wistar rats. Alcohol Clin Exp Res 25: 25-33.
Systemic naloxonazine blocks central µ1-opioid receptors irreversibly, well over 24 h, but can also block µ2-receptors in a reversible manner during the first hours after administration. Thus, when administered 20 h prior to ethanol and concomitant food access, the dose-dependent (1-15 mg/kg) decrease in both ethanol and food intake by naloxonazine could be attributed to a µ1-blockade . However, naloxonazine suppressed ethanol and food consumption as well, when given 15 min before the session, suggesting that both µ1- and µ2-receptor blockade could modulate ethanol reinforcement . When the 15 mg naloxonazine was administered before three successive ethanol sessions, a decrease was seen only during the first session, after which tolerance to the drug's effect developed. This was probably due to a µ-receptor up-regulation, produced by the prolonged receptor blockade .
Effects of selective δ-receptor antagonists on ethanol consumption
Although δ-opioid receptor antagonists have been shown to attenuate ethanol reinforcement in many studies, the results are generally more inconsistent when compared to those from experiments with µ-antagonists. The first δ-antagonist tested for its effects on ethanol drinking was ICI 174864. When both ethanol and water availability was limited to one 30-min daily access period, ICI 174864 (0.5-3 mg/kg) dose-dependently decreased ethanol but not water intake in the alcohol-preferring HAD rats . Similarly, single injections of this compound (3-8 mg/kg) suppressed ethanol drinking during the first of the three 1-h access periods every 4 h in alcohol-preferring P rats without effects on 24-h water drinking . The effect of naltrindole, another δ-receptor antagonist (5-20 mg/kg) tested in the same model, lasted for the first two access periods, and no effects were seen on water consumption . However, separate control experiments indicated that naltrindole also affected ethanol-saccharin and saccharin solution intake, suggesting that the suppressive effects were not specific for ethanol.
Attenuation of ethanol consumption by naltrindole in rats and mice has been reported in several other studies [17, 18, 24, 25], but these positive findings have not been uniformly replicated [16, 20, 26, 27]. The reasons for the discrepant findings are not clear. The failure to see decreases in ethanol drinking by naltrindole was probably not always caused by insufficient dosing, as negative
findings were also reported from experiments where the naltrindole doses matched those employed in positive reports. The genetic background of the animals cannot easily explain the differences either, as indicated by similar naltrindole-induced decreases in operant responding for ethanol both in the alcohol-preferring AA as well as the heterogeneous Wistar rats  (Fig. 1). Finally, a meaningful comparison of the various behavioral models used for measuring ethanol consumption is complicated by different periods of ethanol and/or fluid deprivation affecting the motivational state of the animals and by the various combinations of conditioned factors present at the time of ethanol access.
Based on pharmacological and behavioral evidence, the existence of two δ-opioid receptor subtypes in the rodent brain, δ1- and δ2-receptors, has been proposed [28, 29]. Naltrindole blocks both subtypes but naltriben is a putative δ2-receptor antagonist. Systemic naltriben has been shown to decrease ethanol drinking in a daily 8-h limited access situation and operant responding for ethanol [30, 31]. The effect of naltriben on ethanol reinforcement was relatively specific, as it did not affect intake of saccharin/ethanol or quinine/ethanol solutions in the limited access paradigm nor did it affect responses for the concomitantly available saccharin solution in the operant model.
Effects of selective opioid receptor antagonists on ethanol seeking
Drug-paired environmental stimuli may acquire powerful incentive-motivational properties through classical conditioning and elicit drug craving and relapse in drug abusers as well as in laboratory animals trained to self-administer drugs [32, 33]. There is both clinical and preclinical evidence showing that the endogenous opioid system is not only involved in the direct reinforcing effects of ethanol, but may also partly mediate the effects of conditioned contextual cues on ethanol seeking. For example, naltrexone was demonstrated to attenuate the efficacy of ethanol-associated environmental stimuli to reinstate extinguished responding for ethanol in laboratory rats . Using the same behavioral model, naltrindole and naloxonazine were used for assessing the contribution of µ- and δ-opioid receptors in ethanol seeking  (Fig. 2). Naltrindole decreased ethanol-seeking behavior at the highest dose (5 mg/kg) under stimuli predictive of ethanol reward, but did not affect responding under the stimulus condition associated with non-reward. In contrast, the effect of naloxonazine was not specific, because the effective dose (15 mg/kg) that suppressed ethanol-seeking behavior also decreased responding under the non-reward stimulus condition.
The conditioned place preference methods have long been used to measure the motivating effects of drug-paired environmental stimuli. The non-selective opioid antagonist naloxone has been shown to attenuate ethanol-induced conditioned place preference in mice [34, 35]. In rats, however, it has been difficult to observe ethanol-induced place preference without special conditioning procedures, including exposure to stress. For example, ethanol produced conditioned
place preference in rats that were given electric foot shocks prior to ethanol injections (0.3 g/kg) and conditioning. In these animals, pre-treatment with the µ-antagonist β-funaltrexamine or the δ-antagonist naltrindole dose-dependently reduced preference for the ethanol-paired compartment at doses that have been shown to decrease ethanol drinking [36, 37].
Figure 2. Effects of naltrindole and naloxonazine on cue-induced reinstatement of ethanol-seeking behavior. Both drugs were tested under the stimulus conditions previously associated with ethanol (S+/CS+) or non-reward/water (S-/CS-). Data are expressed as mean (±SEM) responses at the active lever during 30-min reinstatement sessions. (* p < 0.05, significantly different from the vehicle conditions). Adapted with permission of Nature Publishing Group, London, from Ciccocioppo R, Martin-Fardon R, Weiss F (2002) Effect of selective blockade of µ1 or δ opioid receptors on reinstatement of alcohol-seeking behavior by drug-associated stimuli in rats. Neuropsychopharmacology 27: 391-399.
Opioidergic mechanisms of ethanol reinforcement and conditioned ethanol effects
Reductions produced in ethanol consumption and ethanol-seeking behavior by the selective opioid receptor antagonists, generally suggest a role for both the µ- and δ-receptors in these phenomena. However, there are also discrepant findings, especially with respect to the involvement of the δ-receptors in ethanol drinking behavior. Moreover, the experimental data suggest that in most experimental situations, the suppressive effects of opioid receptor antagonists on ethanol consumption are not specific, but reflect the well-known involvement of opioid receptors with regard to ingestive behavior .
The precise roles of the opioid receptors in ethanol reinforcement and conditioned reinforcement processes are not yet very well known. There is evidence that systemically administered ethanol increases the level of extracellular β-endorphin in the nucleus accumbens . Moreover, intra-accumbal infusions
of methylnaloxonium and naltrindole suppressed ethanol self-administration, suggesting that reductions in ethanol reinforcement by opioid antagonists could be related to their inhibition of the endogenous opioid peptide action in the nucleus accumbens [17, 39]. Infusion of these antagonists into the amygdala also attenuated ethanol reinforcement, which could be related to the role of the amygdala in stimulus-reward associations . Another line of evidence suggests that the suppressive effect of opioid antagonists on ethanol reinforcement could involve interaction with mesolimbic dopamine transmission. For example, ethanol-induced increase in extracellular dopamine level in the nucleus accumbens was attenuated by systemic naltrexone and focal naltrindole administration [40, 41 and 42].
In addition to the direct pharmacological actions of ethanol, ethanol-associated contextual stimuli can increase dopamine levels in the nucleus accumbens as well. This is consistent with the view of the role of the midbrain dopamine neurons in the processing of motivational signals [43, 44]. So far, there are no data on the effects of selective opioid antagonists on cue-induced enhancement in dopamine transmission. Both the µ- and δ-receptors are involved in tonic modulation of mesolimbic dopamine transmission , and blockade of these receptors could therefore blunt the efficacy of the contextual cues in enhancing dopamine transmission and reinstating ethanol seeking.
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