Acamprosate, but not naltrexone, inhibits conditioned abstinence behaviour associated with repeated ethanol administration and exposure to a plus-maze

Rationale: Drugs that reduce relapse in alcoholics are thought to inhibit either positive reinforcement for drinking (e.g. naltrexone) or negative reinforcement (e.g. acamprosate), and may reduce the impact of conditioned stimuli associated with previous alcohol use. We have developed a model for such conditioning by repeatedly pairing ethanol administration with plus-maze exposure. Substitution of saline for ethanol greatly increased stretched-attend postures and time in the central square, conditioned to the environment. Objective: To test the hypothesis that if this behaviour indicates a negative affective state caused by the expectation of ethanol, it should be inhibited by drugs that reduce negative, but not positive, reinforcement. Methods: The effects of naltrexone and acamprosate on alcohol-conditioned abstinence behaviour were compared. Results: Acute administration of either drug alone produced no significant effects on plus-maze behaviour in naive mice. Naltrexone had no significant effect on the alcohol-conditioned abstinence behaviour, but acamprosate reduced the incidence of stretched-attend postures. Conclusions: The experiments replicated previous findings for alcohol/environment conditioned behaviour, and demonstrated, as predicted, that this was decreased by acamprosate but not by naltrexone. Effects of acamprosate on conditioned negative reinforcement may be the cause of this effect, but more work is required to establish the usefulness of this model in evaluation of anti-relapse drugs. Received: 4 May 1999 / Final version: 21 July 1999     

Features of environment-dependent tolerance to ethanol

Mice given multiple injections of ethanol in a standardized environment develop environment-dependent tolerance to the hypnotic and hypothermic effects of ethanol. These animals also demonstrate environment-dependent cross-tolerance to the hypnotic and hypothermic effects of pentobarbital. Examination of the levels of ethanol in the brain and blood at various times after injection of a test dose of ethanol, as well as the examination of the rate of disappearance of ethanol from the blood, indicated that environment-dependent tolerance could be explained by dispositional factors. On the other hand, mice rendered tolerant to ethanol by a liquid diet technique demonstrate tolerance that is not environment-dependent, and there is no alteration in ethanol levels in different environments for animals fed the liquid diet. When the animals in either paradigm are tested by injecting ethanol directly into the brain, tolerance is observed that is not dependent on the environment. Tolerance produced by these two different paradigms is apparently due to different adaptive strategies used by the animal. Environment-dependent tolerance is partially related to the ability of the animal to change the disposition of ethanol, while environment-independent tolerance may be entirely due to other factors, such as changes in neuronal sensitivity to ethanol.     

The effects of chronic administration of ethanol on startle thresholds in rats

The thresholds for startle responses to electric shock were measured in adult male Wistar strain rats given ethanol daily in doses rising from 3 to 7 g/kg over a 30-day period, and in controls receiving equicaloric doses of sucrose. Tests made 23, 36, or 47 h after ethanol (i.e., during partial or complete ethanol withdrawal) gave threshold values significantly lower than those obtained with sucrose-treated controls. The difference became greater after longer ethanol treatment and larger doses. However, when threshold measurements were made under the acute influence of ethanol in the experimental group, the mean values were virtually equal to those of the sucrose controls. This normalization, by ethanol, of a disturbance produced by absence of ethanol in a chronically treated animal is indicative of physical dependence. Following termination of ethanol treatment there was a gradual return of startle thresholds almost to control values over a relatively short period, indicating that the changes underlying the hyperexcitability are readily reversible.     

Effects of repeated ethanol administration in the plus maze; a simple model for conditioned abstinence behaviour

Classical conditioning is considered to be an important factor in drug dependence. Exposure to an environment in which alcohol has been repeatedly consumed may produce feelings of anxiety and dysphoria in a currently abstinent patient, and may precipitate relapse drinking. The present study modeled this process, by repeatedly exposing mice to an elevated plus maze after ethanol administration. When ethanol injections and maze exposure were repeated for 9 days, and the ethanol injections replaced by saline on the tenth day, mice consistently exhibited a characteristic behaviour pattern, with increased stretched attend postures and proportion of time spent in the central square. This behaviour was different from that previously seen during withdrawal from ethanol, and was not observed when repeated injections of ethanol were given either without, or after, maze exposure. Thus the characteristic behaviour pattern appeared to be contingent on the animals being repeatedly exposed to the maze environment while under the influence of ethanol. In particular, the reduction in stretched attend postures produced by acute ethanol, the tolerance to this behaviour seen after repeated ethanol and the increase in it after replacement of ethanol by saline, are consistent with the pattern predicted for a behavioural response to the absence of ethanol, conditioned to the environment of the maze. Received: 21 May 1998/Final version: 4 September 1998     

Naltrexone effects on ethanol consumption and response to ethanol conditioned cues in C57BL/6 mice

RATIONALE: The conditions under which naltrexone reduces ethanol consumption and its effect on behavior controlled by ethanol conditioned stimuli remain unclear. OBJECTIVES: The objectives were to determine the effects of naltrexone on ethanol consumption by C57BL/6 (B6) mice when injected subcutaneously (expt 1) or delivered by osmotic minipump (expt 2), and on ethanol conditioned cues (expt 3). METHODS: Naltrexone effects on ethanol consumption and preference were measured in a continuous access two-bottle choice paradigm in groups of mice implanted with osmotic minipumps delivering 0-3.0 mg/kg per day or injected subcutaneously with 0-6.0 mg/kg doses. Naltrexone's (0-3.0 mg/kg) effect on ethanol-conditioned cues was indexed by its effect on the expression of ethanol place conditioning (expt 3). RESULTS: Naltrexone produced a transient reduction in ethanol consumption and a consistent reduction in preference when injected; however, it had no effect on ethanol consumption or preference when delivered continuously by osmotic minipump. Naltrexone attenuated the expression of ethanol place conditioning in a U-shaped dose-response function. CONCLUSIONS: The transient reduction in ethanol consumption produced by injected naltrexone and the absence of an effect when continuously delivered confirms a report that maintaining naltrexone at steady state levels may antagonize its attenuation of ethanol consumption. The reduced expression of ethanol place conditioning in naltrexone-injected mice suggests that the drug can attenuate the reinforcing effects of ethanol conditioned stimuli as was recently reported for lever responding maintained by ethanol conditioned stimuli in rats. These effects were observed at naltrexone doses with no readily apparent adverse side-effects, supporting its usefulness for treating alcoholism.     

Dissociation of tolerance and physical dependence after ethanol/chlordiazepoxide intake

The previously-observed attenuation of withdrawal reactions in mice (group B) fed an ethanol diet containing chlordiazepoxide (CDP) was not due to a difference in the rate of disappearance of blood ethanol levels during chronic diet treatment in group B compared to mice which received only the ethanol diet (group A). Injection of group A mice with CDP or N-demethyl CDP (10 mg/kg) at the time of diet withdrawal did not result in any significant attenuation of withdrawal scores. Injection of the lactam metabolite of CDP (LCDP; 10 mg/kg) resulted in significantly attenuated withdrawal scores at 4 and 6 hr only, but the pattern of withdrawal scores were different from that for group B mice. Moreover, blood LCDP level, in mice injected with LCDP, at 4 hr was at least five times higher than that attained in group B mice (from diet containing CDP). These results support our previous conclusion that the presence of major metabolites of CDP during withdrawal could only account for a minor contribution to the protective effect. Mice in A and B did not differ in the degree of functional tolerance which developed as a result of ethanol intake. Thus, there was an apparent dissociation between tolerance and physical dependence in the mice which had consumed the CDP/ethanol diet. The magnitude of decrease of GABA levels in the cerebellum and cerebral cortex at 4 hr after withdrawal also did not differ between the two groups, suggesting the reduction in GABA levels could not be correlated with the intensity of withdrawal signs.     

State-dependent effects of ethanol on active and passive avoidance learning

The present set of experiments examined the importance of the response initiation-inhibition parameter of certain avoidance conditioning tasks in the production of state-dependent dissociative effects with ethanol. On those tasks involving some degree of response inhibition (passive avoidance and two-way active avoidance), animals receiving ethanol during training were more impaired in their testing performance than those receiving saline during training (anterograde amnestic effect), and animals injected with ethanol during training and saline during testing displayed dissociation of their acquired avoidance behaviors during testing (asymmetrical dissociation effect). On the task involving a response initiation element with little contamination by response suppression factors (one-way active avoidance), dissociation of avoidance behavior during testing was found both for the animals trained under ethanol and tested under saline and for the animals trained under saline and tested under ethanol (symmetrical dissociation effect). The results were discussed in terms of possible joint effects of symmetrical state-dependency and other behavioral properties of the drug. However, an alternative interpretation could not be ruled out, namely that the mechanisms involved in the impairment found on the testing day for the drug-placebo and placebo-drug groups may be different. It was suggested that the drug-placebo group may represent the more general example of state-dependent dissociation effects, whereas the production of state-dependent dissociation effects in the placebo-drug group may depend upon the type of behavior conditioned and/or the strength of such conditioning.     

Role of Pavlovian conditioning in the development of tolerance and cross-tolerance to the hypothermic effect of ethanol and hydralazine

The role of Pavlovian conditioning in the development of tolerance to the hypothermic effect of ethanol and of cross-tolerance to hydralazine was investigated. In the first study, two groups of rats were treated on alternate days with ethanol (2 or 4 g/kg, respectively, IP) in a novel and distinctive environment (DR). On the non-alcohol days, they received saline in the home room (HR). A control group received saline in both environments. Tolerance to the hypothermic effect of ethanol in the DR was demonstrable in both the 2 and 4 g/kg treatment groups. Tolerance in the HR, however, was observed only in the 4 g/kg treated group. Cross-tolerance to the hypothermic effect of hydralazine was observed for both ethanol-treated groups in the DR but not in the HR. In the second study, ethanol treatment was carried out by daily intubation with 6 g/kg ethanol in the home cage. Tolerance to ethanol-induced hypothermia was demonstrated either in the home cage or in a novel environment. This treatment, however, failed to confer cross-tolerance to the hypothermic effect of hydralazine. These findings suggest that conditioning plays a predominant role in the tolerance produced by low but not by high treatment dosage. The data also suggest that conditioning might be a separate component in tolerance development, which is of special importance in tolerance to behavioral effects in the whole animal rather than to cellular or molecular effects.     

Tolerance to hypothermia induced by ethanol depends on specific drug effects

Two experiments were conducted to test the hypothesis that tolerance to the hypothermic effect of ethanol fails to develop if rats are denied the unconditional stimulus represented by hypothermia. In both experiments, rats were injected with either ethanol (1.9 or 2.5 g/kg) or saline and given microwave hyperthermia (MHT) to offset the hypothermic effect of the drug or sham-MHT. In one experiment, rats no longer demonstrated a hyperthermic response to a saline challenge after hypothermia was offset during 5 MHT treatment sessions. In a second experiment, rats prevented from becoming hypothermic did not develop tolerance to the hypothermic effect of ethanol due to MHT treatment, but did become tolerant to the ataxic effects of ethanol, which were unaffected by MHT. Results suggest that rats must experience the specific consequences of a drug to become tolerant to that effect.     

A simple drinking test for measuring the effects of ethanol on the central nervous system

An objective behavioral test is described for determining different degrees of alcohol intoxication in rats and other small laboratory animals. It is based on a measure of the length of time taken for a water-deprived rat to drink (standing upright) following the administration of a challenge dose of ethanol. The differential effects on drinking impairment of various doses of ethanol IP were examined in either the same (group A) or different (group B) animals. The test was shown to be sensitive to dose differences as small as 0.25 g/kg and to be applicable over a wide range of doses (1–4 g/kg). Lower (0.5 g/kg) or higher doses (4.5 and 5.0 g/kg) were, respectively, either ineffective or generated relatively higher latency scores than those obtained with the intermediate range of doses. The dose-response curve thus showed two rectilinear segments; regression coefficients were 154 and 183 min/g/kg^-1 in the dose range up to 2.5 and up to 4.0 g/kg for groups A and B, respectively, 431 min/g/kg^-1 for doses greater than 4.0 g/kg (group B). Blood alcohol levels measured at the onset of drinking did not differ significantly as a function of dose, between 1.0 and 4.0 g/kg (mean 0.864±0.070 mg/ml), but were higher compared to those observed with doses of 4.5 and 5.0 g/kg (0.281±0.180 mg/ml). The applicability of the drinking test to measures of initial and acquired tolerance toward drug effects is discussed.     

Behavioral stimulation induced by ethanol withdrawal

A model for the study of an ethanol withdrawal syndrome on operant behavior is described. Rats maintained of 16% w/v solutions of ethanol for several months were trained to perform on a DRL-15 schedule. On withdrawal of ethanol the interresponse times were significantly shortened concomitant with an increase in the total number of responses.     

Acamprosate and naltrexone treatment effects on ethanol and sucrose seeking and intake in ethanol-dependent and nondependent rats

Rationale  Two pharmacotherapies are approved for treating alcohol craving (acamprosate and naltrexone), but both have shown mixed findings in animals and humans. Objectives  The present experiments utilized a “reinforcer blocking” approach (i.e., rats were able to consume ethanol during treatment) to better understand the efficacy of these treatments for ethanol seeking and drinking using ethanol-dependent and nondependent rats. Materials and methods  In “nondependent” experiments, drugs (acamprosate 50, 100, and 200 mg/kg; naltrexone 0.1, 0.3, and 1.0 mg/kg) were administered over 3-week periods prior to operant sessions with a low response requirement to gain access to reinforcers for 20 min. For “dependent” experiments, rats were made dependent in vapor/inhalation chambers. Results  Acamprosate and naltrexone had similar effects on intake in nondependent and dependent rats; neither drug was selective for ethanol over sucrose drinking. In nondependent animals, naltrexone was more efficacious at more doses than acamprosate, and acamprosate’s effects were limited to a dose that also had adverse effects on body weight. Both pharmacotherapies showed more selectivity when examining reinforcer seeking. In nondependent rats, acamprosate and naltrexone had response-attenuating effects in ethanol, but not sucrose, groups. In dependent animals, acamprosate had selective effects limited to a decrease in sucrose seeking. Naltrexone, however, selectively decreased ethanol-seeking in nondependent rats. Conclusions  The naltrexone-induced decreases in seeking suggested a change in incentive motivation which was selective for ethanol in nondependent rats. The “nondependent” paradigm may model early stages of “problem drinking” in humans, and the findings suggest that naltrexone could be a good intervention for this level of alcohol abuse and relapse prevention.     

An extinction cue reduces spontaneous recovery of ataxic ethanol tolerance in rats

Rationale and objectives Ethanol ataxia experiments with rats investigated cue effects on conditioned tolerance. Spontaneous recovery (SR) was assessed 1 day and 18 days after extinction with conditioned stimuli (CSs) paired or unpaired with an ethanol unconditioned stimulus (US). Behavioral tolerance was assessed by not tilting the apparatus during conditioning. Non-associative processes were assessed post-conditioning with or without a buzzer cue. Boutons (1993, Psychol Bull 114:80–99) memory theory was tested using an extinction cue and an associatively neutral cue presented during SR testing.Methods Tolerance was conditioned to a room + strobelight CS by ethanol injections experienced on a tilting floor (standard conditioning). Controls received no ethanol or ethanol, either during the CS without the floor tilting or 11 h post-CS. SR testing occurred 1 day or 18 days after extinction (experiment 1). Conditioning was followed by tolerance and CR tests either with or without a 15-s buzzer cue (experiment 2). In extinction, the CS and cue occurred without ethanol; the cue occurred before 7% or none of the extinction trials. Testing occurred 18 days after extinction with or without that cue (experiment 3), or with an equally familiar (neutral) cue presented before conditioning (experiment 4). Results Tolerance developed without floor tilting. CS–US unpairings prevented tolerance. Tolerance SR occurred 18 days but not 1 day after extinction only after CS–US pairings (experiment 1). Post-conditioning tests showed no unconditioned effects of the cue (experiment 2). Testing with no cue 1 day after extinction with the cue resulted in no tolerance increase. The extinction cue reduced SR (experiments 3 and 4); the neutral cue did not (experiment 4).Conclusions Cues correlated with extinction reduce SR. Non-associative and practice processes, Boutons (1993, Psychol Bull 114:80–99) memory theory, alternative interpretations, and clinical implications are discussed.This research was supported by funding from Denison University.     

Behavioral augmentation of tolerance to ethanol in the rat

Three groups of rats were trained on a circular maze task, then were tested under the influence of ethanol. Thereafter, all three groups received ethanol daily, on different schedules. One group (psychological) received ethanol (1.2 g/kg i.p.) just before each treatment session; another group (physiological) received the same dose immediately after the session; and the control group received only saline. All three groups were tested under ethanol every fourth day. The psychological group showed significant tolerance by the second test day, and maximal tolerance by the fourth. The physiological group reached the same maximum tolerance by the sixth test day, while the controls showed no increase in tolerance. Addition of daily gavage with ethanol (6 g/kg) did not modify the level of tolerance in the psychological or physiological groups, but raised the controls to the same level as the others. None of the changes in tolerance were attributable to increased rate of ethanol elimination. It is concluded that production of tolerance by these various techniques is distinguishable only with respect to rate.     

Simple method for producing an alcohol withdrawal syndrome in rats.

Rats received intragastric intubations of ethanol at 8 hr intervals for 1, 7, 15 or 30 days. The dosage for each animal was one which produced observable signs of intoxication 1 hr after the intubation. All of the rats in the experimental groups developed a tolerance to ethanol as indicated by the increasing dose required to induce intoxication, but the degree of tolerance was related to the duration of the ethanol administration. During the withdrawal period the incidence of hyperreactivity, convulsive symptoms, and the susceptibility to audiogenic seizures was determined for all 4 groups. Although every experimental animal displayed withdrawal symptoms, the incidence of these symptoms was found to be an increasing, negatively accelerated function of the duration of ethanol exposure. For situations where voluntary consumption of alcohol is not necessary this method is a simple, controlled, reliable, way of inducing ethanol tolerance and physical dependence in rats.     

Changing environmental cues reduces tolerance to nicotine-induced anorexia

Male rats on a 22-h food deprivation schedule were injected daily with a low dose of nicotine and allowed to drink sweetened milk for 10 min in a test cage in the colony room. Nicotine initially suppressed milk intake but complete tolerance developed within 10 days so that the amount of intake did not differ from saline controls. The role of temporal cues was tested on the next day by changing the timing of cues, and omitting others that normally preceded nicotine injection while keeping constant the physical environment within which injection and testing took place and the drug-test interval. Changing the timing of injection significantly suppressed milk intake. These results show that tolerance to the anorectic effects of a low dose of nicotine is partially dependent on the presence and timing of cues associated with tolerance acquisition.     

Interactive effects of ethanol and nicotine on learning in C57BL/6J mice depend on both dose and duration of treatment

Objective and rationale Alcohol and nicotine are commonly co-abused; one possible explanation for co-abuse is that each drug ameliorates the aversive effects of the other. Both drugs have dose-dependent effects on learning and memory. Thus, this study examined the interactive effects of acute ethanol and acute, chronic, or withdrawal from chronic nicotine on fear conditioning in C57BL/6J mice. Materials and methods Conditioning consisted of auditory conditioned stimulus-foot-shock unconditioned stimulus pairings. For acute studies, saline or ethanol, then saline or nicotine was administered before training, and saline or nicotine was also administered before testing. For chronic and withdrawal studies, saline or nicotine was administered chronically, and ethanol or saline was administered before training. Results Acute nicotine (0.09 mg/kg) reversed ethanol-induced deficits (1.0 and 1.5 g/kg) in contextual and cued fear conditioning, whereas a low dose of ethanol (0.25 g/kg) reversed nicotine (6.3 mg kg⁻¹ day⁻¹) withdrawal-induced deficits in contextual conditioning. Tolerance developed for the effects of nicotine on ethanol-induced deficits in conditioning and cross-tolerance between chronic nicotine and acute ethanol was seen for the enhancing effects of ethanol on conditioning. Conclusions The complex and sometimes polar actions of ethanol and nicotine on behavior may contribute to co-abuse of these drugs. Specifically, smoking may initially reduce the aversive effects of ethanol, but tolerance develops for this effect. In addition, low doses of alcohol may lessen nicotine withdrawal symptoms.     

Does tolerance develop to the activating,as well as the depressant,effects of ethanol?

Genetically determined differences were demonstrated in the response of mice to low doses of ethanol. Ethanol (1.35 g/kg) produced an increase in locomotion in DBA/2 and BALB/c mice, but did not alter the locomotor activity of C57B1/6 mice. Chronic administration of ethanol produced tolerance to the sedative/hypnotic effects of high doses of ethanol in DBA/2 and BALB/c mice, but the equivalent chronic ethanol administration paradigm produced no tolerance to the activating effects of ethanol in these animals. C57B1/6 mice became tolerant to the hypnotic effects of ethanol, but no change in the behavior of these mice, given a low dose of ethanol, was noted after the mice were withdrawn from chronic feeding with ethanol-containing diets. The results indicate the presence of different mechanisms for tolerance development to the activating and depresant effects of ethanol, and indicate that strain-dependent differences in the activating effects of ethanol are not determined by an animal's greater sensitivity to the sedating effects of this drug.     

The effects of repeated doses of ethanol on exploration and its habituation

The effects of ethanol (0.8–2.4 g/kg) on exploratory behavior and its habituation were investigated by testing DBA/2 mice in a holeboard apparatus. Mice familiar with the holeboard had lower levels of exploration than animals with no previous experience in the apparatus. If the animals received ethanol during their first exposure to the holeboard they did not show the same degree of habituation. Ethanol (0.8 g/kg) increased the exploration of animals naive to the holeboard, but failed significantly to increase the exploration of animals familiar with the apparatus. An increase in locomotor activity was observed following treatment with ethanol (2.4 g/kg). This was potentiated if animals received a single treatment with ethanol 3 days earlier, but only if they were tested in the apparatus following the initial treatment. These results have important implications for the design of experiments investigating tolerance and sensitization to ethanol's effects on locomotor activity and exploration.