15. Baclofen - preclinical data

Editors: Spanagel, Rainer; Mann, Karl F.

Title: Drugs for Relapse Prevention of Alcoholism, 1st Edition

Copyright 2005 Springer

> Table of Contents > Baclofen: preclinical data

Baclofen: preclinical data

Mauro A.M. Carai¹

Roberta Agabio¹

Giovanni Addolorato²

Gian L. Gessa^{1, 3}

Giancarlo Colombo³

¹ Department of Neuroscience, University of Cagliari, Cagliari, Italy

² Institute of Internal Medicine, Catholic University, Rome, Italy

³ C.N.R. Institute of Neuroscience, Section of Cagliari, Viale Diaz 182, I-09126 Cagliari, Italy

Introduction

Alcohol interacts, as a receptor modulator capable of selectively altering specific neurochemical processes, with multiple brain receptor systems. The behavioral consequences of alcohol ingestion are thought to be the summation of its effects at these receptor systems. The contribution of each receptor system to the behavioral effects of alcohol varies as the alcohol dose/concentration is increased, providing a neurochemical basis for the dose-dependent nature of alcohol effects. Accordingly, it may be conceived that at a given alcohol dose/concentration, a specific receptor system is more sensitive to alcohol than others, thus resulting in a more prominent contribution to a particular behavioral effect of alcohol.

Recent experimental results (reviewed in the present chapter) as well as preliminary clinical data (reviewed in another chapter included in this book [1]) suggest that the GABA_B receptor may be considered a novel player among the receptor systems controlling different aspects of alcohol drinking behavior, including alcohol consumption, alcohol relapse and alcohol appetitive properties. These preclinical and clinical findings feature GABA_B receptor agonists as promising agents in the pharmacotherapy of alcoholism.

Effect of GABA_B receptor agonists on alcohol intake and alcohol motivational properties in rats

Previous studies on the ability of GABA_B receptor agonists to alter alcohol intake (under the non-operant, 2-bottle choice regimen) and alcohol self-administration (under operant procedures) have yielded mixed results. Indeed, it has been reported that the prototype GABA_B receptor agonist, baclofen, reduces [2, 3], produces no change on [4], or even stimulates [5, 6] voluntary alcohol intake in rats, when given a choice between two bottles containing an alcohol solution and water, respectively. In the self-administration studies (where rats were trained to press a lever to gain access to alcohol), and depending on the drug

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dose or the experimental procedure used, baclofen has been found to stimulate [7] or decrease [7, 8 and 9] operant responding for alcohol in rats. Furthermore, when observed, the inhibition of alcohol-motivated lever pressing was not selective, being accompanied by a proportional decrease in lever pressing for an alternative reinforcer such as sucrose [8]. In a study using the sipper tube model of alcohol access, which permits some separation between the appetitive and consummatory aspects of alcohol self-administration, baclofen decreased alcohol-seeking behavior (measured as the time spent in achieving the response requirement to gain access to the alcohol solution), while it increased alcohol-consummatory behavior (the amount of alcohol actually consumed) in rats [10].

More consistent results have been obtained in Sardinian alcohol-preferring (sP) rats, one of the few rat lines selectively bred worldwide for high alcohol preference and consumption. Indeed, recent work (as reviewed below) has demonstrated that the pharmacological activation of the GABA_B receptor in sP rats resulted in the suppression of a) acquisition and maintenance of alcohol drinking behavior, b) alcohol motivational properties, and c) alcohol relapse-like drinking.

In the acquisition study [11], the GABA_B receptor agonists, baclofen and CGP 44532, were administered to alcohol-naive sP rats, i.e., rats which had never consumed alcohol before the start of the experiment. Adult, male sP rats were singly housed and injected intraperitoneally with either baclofen (0, 1 and 3 mg/kg) or CGP 44532 (0, 0.1, 0.3 and 1 mg/kg) once a day, for a total of 10 consecutive days. Alcohol (10%, v/v) and water were offered under the standard, homecage 2-bottle alcohol versus water choice with unlimited access for 24 h/day, immediately following the first injection of baclofen or CGP 44532. Food was available ad libitum.

In vehicle-treated rats, acquisition of alcohol drinking behavior had a rapid onset, reaching an average daily intake of 5-6 g/kg (i.e., the amount of alcohol ordinarily consumed on a daily basis by sP rats) within 4-7 days. In contrast, daily alcohol intake was dose-dependently suppressed in baclofen- and CGP 44532-dosed rats throughout the 10-day treatment period. Interestingly, in the rat groups treated with the highest dose of each drug (3 mg/kg baclofen and 1 mg/kg CGP 44532), daily alcohol intake was on average lower than 1.5 g/kg. Reduction in alcohol intake, induced by both GABA_B receptor agonists, was associated with a full compensatory increase in daily water intake, so that the total daily fluid intake (i.e., the sum of alcohol solution and water consumed) remained unchanged. Daily food intake (a variable usually recorded in pharmacological studies on alcohol intake as a signal of animal malaise or nonselectivity of drug action) tended to be higher in the rat groups treated with baclofen or CGP 44532, as compared to the vehicle-treated groups. This is likely to be due to the fact that in the control group, part of the total calorie intake was provided by alcohol. On completion of the treatment, daily alcohol intake progressively increased in the 3 mg/kg baclofen-treated group as well as in the 1 mg/kg CGP 44532-treated group, reaching control values after 10-14 days. Water intake diminished consistently.

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A separate experiment [12] investigated the effects of baclofen on voluntary alcohol intake in alcohol-experienced sP rats, i.e., rats in which the consumption of pharmacologically relevant doses of alcohol was already established before baclofen administration. Alcohol-experienced rats are thought to represent a model of the maintenance or active drinking phase of human alcoholism. In this study, adult male sP rats were singly housed and exposed to alcohol for approximately 2 months before starting with drug treatment. Alcohol (10%, v/v) was offered under the 2-bottle alcohol versus water regimen with unlimited access for 24 h/day. Food was readily available. Alcohol intake averaged 6 g/kg/day across the 2-month period of alcohol exposure, which preceded the start of the experiment. Baclofen was injected intraperitoneally at doses of 0, 2.5, 5 and 10 mg/kg, once a day for a total of 14 consecutive days.

Alcohol intake in vehicle-treated rats averaged between 5.5 and 7 g/kg throughout the 14-day treatment period, whereas those animals treated with baclofen experienced a dose-dependent reduction of up to 40-50% in daily alcohol consumption. However, tolerance to the reducing effect of baclofen on alcohol intake progressively developed on continuing treatment. An increase in daily water intake fully compensated the reduction in alcohol consumption, with the total daily fluid intake remaining virtually unchanged. Food intake was significantly altered only by treatment with 10 mg/kg baclofen. This effect, however, was present only during the first half of the treatment period.

The results of the acquisition and maintenance experiments suggest that stimulation of the GABA_B receptor by baclofen and/or CGP 44532 results in a virtually complete blockade of the disclosure and experience of those effects of alcohol that sustain alcohol drinking behavior, which is otherwise a phenomenon with a rapid onset and stable maintenance in sP rats, as indicated in control rats by the constant daily intake of pharmacologically relevant amounts of alcohol from the very beginning of alcohol exposure.

The above-mentioned experiments, performed with the 2-bottle choice paradigm, focused on the effect of baclofen and CGP 44532 on some consummatory aspects of alcohol ingestive behavior in sP rats. Subsequently, we extended to the appetitive, or motivational, properties of alcohol the investigation on the anti-alcohol effect of GABA_B receptor agonists. Specifically, we evaluated the effect of baclofen on the extinction responding for alcohol, defined as the maximal amount of work that a rat trained to lever-press for alcohol is willing to perform to obtain alcohol [13]. Extinction responding has been proposed to represent an index of the appetitive strength of alcohol [14, 15]: the more the rat works on the lever, the stronger its motivation to gain access to alcohol. Recent work [16] has shown that sP rats trained to lever-press for alcohol displayed a) high values of extinction responding for alcohol, and b) a high and positive correlation between extinction responding and alcohol self-administration of the preceding session. Thus it is confirmed that alcohol possesses strong motivational capacities in sP rats, highlighting the suitability of this rat line for the planned study.

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In the baclofen study [13], adult male sP rats were initially trained to lever-press for oral alcohol (15%, v/v) in daily 30-min sessions under a fixed ratio 4 (FR4) schedule (i.e., every 4 consecutive presses of the lever resulted in the presentation of a 0.1 ml drop of alcohol solution). After approximately 20 sessions, all rats displayed a robust and stable lever-pressing behavior, which resulted in a mean alcohol intake of 0.6 g/kg/session, with blood alcohol levels in the range of 40-50 mg%. A separate group of sP rats, trained to lever-press for 3% (w/v) sucrose under identical conditions, was included in the study to assess the specificity of the baclofen action on extinction responding for alcohol. Extinction responding for alcohol or sucrose was defined as the maximal number of lever presses attained by each rat in the absence of alcohol or sucrose reinforcement. More specifically, during extinction sessions, rats were exposed to the operant chamber for 30 min with lever-pressing not resulting in any alcohol or sucrose presentation. On test sessions, baclofen was injected intraperitoneally at doses of 0, 1, 2 and 3 mg/kg. Each dose of baclofen was tested in every single rat of both groups under a latin-square design.

Extinction responding for alcohol and sucrose in saline-treated rats averaged 54.8 + 8.4 and 55.6 + 13.2 (mean + SEM), respectively, suggesting that 15% alcohol and 3% sucrose had comparable motivational properties in sP rats. Pretreatment with baclofen resulted in a dose-dependent suppression of extinction responding for alcohol: lever pressing of the rat groups treated with 1, 2 and 3 mg/kg baclofen was 64, 88 and 98% lower, respectively, than that observed in saline-treated rats. Seven out of eight rats in the 3 mg/kg baclofengroup, in fact, completely avoided pressing the lever. In the sucrose experiment, only the highest dose of baclofen tested significantly affected extinction responding for sucrose; extinction values in the rat groups treated with 1, 2 and 3 mg/kg baclofen were 23, 46 and 99% lower, respectively, than those observed in saline-treated rats. Taken together, these results suggest that baclofen was more potent in reducing the motivation for alcohol than for sucrose.

A separate experiment found that the above doses of baclofen did not affect spontaneous motor activity in sP rats, when tested in an open-field arena, suggesting that the suppressing effects of baclofen on extinction responding were indeed secondary to its ability in reducing the appetitive strength of alcohol and, furthermore, not due to muscle-relaxant or sedative properties of the drug.

These results suggest the involvement of the GABA_B receptor in the neural system mediating alcohol reinforcement in sP rats, and that the reducing effects of baclofen on alcohol intake in sP rats may be secondary to its ability in suppressing the motivational properties of alcohol.

In conclusion, there appear to be some discrepancies of present-day data in the literature (indicating mixed effects of baclofen on alcohol intake and self-administration [2, 3, 4, 5, 6, 7, 8, 9 and 10]) and those collected in sP rats (which consistently depict a reducing effect of baclofen and CGP 44532 on alcohol preference, consumption and motivational properties). Differences in the baclofen dose-range, route of baclofen administration, and the procedure of alcohol exposure may account, at least in part, for these discrepancies. However, the strain of rats used in these

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studies may be - to our understanding - an important factor in explaining these differences. Rats of the sP lines apparently possess a genetically determined sensitivity to the reducing effect of GABA_B receptor agonists on alcohol intake and reinforcement, which may not be present in other rat strains. This peculiar sensitivity makes sP rats a proper animal model for investigations concerning the role of the GABA_B receptor in the control of alcohol-ingestive behavior. Furthermore, the inconsistencies among the data generated with sP and other rat strains are not completely surprising, as a wide heterogeneity has historically been found among the different lines of alcohol-preferring rats with regard to the efficacy of certain drugs in reducing alcohol intake and self-administration. Thus, this replicates, to some extent, the different efficacies of some pharmacotherapies among the different types of alcoholics (e.g. [17, 18 and 19]).

Effect of GABA_B receptor agonists on relapse-like behaviors in rats

With a deeper focus on the scope of the present book, we report here the results of a series of experiments designed to investigate the effects of baclofen and CGP 44532 on the so-called alcohol deprivation effect (ADE), i.e., the transient increase in alcohol intake which occurs in several animal species following a period of abstinence from alcohol. This phenomenon has been proposed to model the loss of control over alcohol and the episodes of alcohol relapse seen in human alcoholics (see [20, 21]). Rats of the sP line appear to constitute a proper animal model for pharmacological investigations of ADE, as they have been found to display a pronounced ADE during the first hour of re-access to alcohol following its deprivation [22, 23]. Furthermore, ADE in sP rats has been found to be reduced with naltrexone (this laboratory, unpublished observations), a drug reported to possess some efficacy in reducing the likelihood of relapse in alcoholics (see [24]), providing evidence for the predictive value of this animal model with regard to human pathology.

In experiments carried out to test the effect of baclofen [25] and CGP 44532 on ADE, individually housed adult male sP rats were initially offered alcohol (10%, v/v) and water under the standard 2-bottle choice with unlimited access for 8 consecutive weeks. Subsequently, rats were divided into 2 groups (matched for alcohol intake over the last 7 days): one group was deprived of alcohol for 14 consecutive days, during which water was the sole fluid available (alcohol-deprived rats); the second group continued to have unlimited access to alcohol and water (alcohol-nondeprived rats), consuming an average of approximately 6 g/kg/day alcohol. At the end of the deprivation phase, 30 min before lights off, rats of both groups (alcohol-deprived and -nondeprived) were further divided into 4 subgroups (n = 7-8 in both experiments) and acutely injected with 0, 1, 1.7 and 3 mg/kg baclofen, or with 0, 0.03, 0.1 and 0.17 mg/kg CGP 44532. Both drugs were injected intraperitoneally. Alcohol was re-presented at lights off and its consumption was recorded 60 min later (previous studies indicated that 60 min after alcohol representation

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is the time interval during which ADE is maximal in sP rats [22, 23]). Standard rat chow was available throughout the studies.

Figure 1. Suppressing effect of the GABA_B receptor agonists, baclofen (top) and CGP 44532 (bottom), on the alcohol deprivation effect (ADE) in sP rats given alcohol under the 2-bottle choice regimen. Each bar is the mean + SEM of n = 7-8. *: P < 0.05 with respect to saline-treated alcohol-nondeprived rats; +: P < 0.05 with respect to saline-treated alcohol-deprived rats (Newman-Keuls test). Top panel reprinted from Drug Alcohol Dep 70: 105-108, 2003, with permission from Elsevier.

In both experiments, alcohol intake was 50-80% higher in saline-treated alcohol-deprived rats than in saline-treated alcohol-nondeprived rats (Fig. 1); this increase in alcohol intake was indicative of the development of a marked ADE. This increase in alcohol intake was eliminated by baclofen and CGP 44532. Indeed, all doses of baclofen (Fig. 1, top panel) and the two highest doses of CGP 44532 (Fig. 1, bottom panel) resulted in a virtually complete suppression of the extra intake of alcohol produced by alcohol deprivation. Importantly, no dose of baclofen or CGP 44532 affected water and food intake, tending to exclude that the action of baclofen and CGP 44532 on ADE was due to their muscle-relaxant and sedative effects. Accordingly, complementary experiments on motor activity found that the doses of baclofen and CGP 44532 that suppressed ADE neither affected the time spent moving, nor the distance

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traveled, nor the number of rearings (measures of horizontal and vertical motor activities in rodents) in alcohol-consuming sP rats tested in an open-field arena.

As demonstrated for several other alcohol-related behavioral responses, multiple receptor systems are likely to be involved in the expression of ADE. Indeed, drugs acting at the glutamate [26, 27, 28, 29, 30 and 31], opioid [32, 33 and 34], dopamine [35], GABAA/benzodiazepine [36], and cannabinoid [37] receptors have also been reported to modulate ADE in rats and mice. The results of the present study include the GABA_B receptor in the neural substrate mediating ADE.

Finally, because of the predictive validity of ADE as an experimental model of alcohol relapse, the results of the present study suggest that baclofen may possess some efficacy in preventing relapse in human alcoholics. The results of preliminary, clinical surveys, reviewed by Addolorato and colleagues [1], apparently support this hypothesis, suggesting that baclofen may constitute a novel medication for alcoholism.

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