CB1 receptors regulate alcohol-seeking behavior and alcohol self-administration of alcohol-preferring (P) rats

Rationale

The endogenous cannabinoid (CB) system mediates a number of behaviors associated with drug-seeking and drug self-administration. In this study the effects of CB1 receptor manipulations on operant ethanol (EtOH) responding during EtOH-seeking, EtOH-relapse as well as on-going EtOH self-administration were determined.

Methods

Alcohol-preferring (P) rats were trained in 2-lever operant chambers to self-administer 15% EtOH (v/v) and water on a concurrent fixed-ratio 5-fixed-ratio 1 (FR5-FR1) schedule of reinforcement in daily 1-h sessions. After 10 weeks, rats underwent 7 extinction sessions, followed by 2 weeks in their home cages without access to EtOH or operant chambers. Rats were then returned to the operant chambers for testing of EtOH-seeking behavior (no EtOH present) for 4 sessions. After a week in their home cages following the EtOH-seeking test, rats were returned to the operant chambers with access to EtOH and water (relapse). Rats were then maintained in the operant chambers for daily 1-h sessions with access to 15% EtOH and water for several weeks.

Results

The CB1 receptor antagonist (SR141716A), at doses of 1 and 2 mg/kg, i.p. reduced EtOH-seeking and transiently reduced EtOH self-administration during relapse and maintenance. Conversely, treatment with the CB1 receptor agonist CP 55, 940, at doses of 1 and 10 μg/kg i.p., increased EtOH-seeking and EtOH self-administration during relapse.

Conclusions

The results of this study demonstrate that activation of CB1 receptors are involved in regulating EtOH-seeking as well as the reinforcing effects of EtOH under relapse and on-going self-administration conditions.     

Reinstatement of alcohol-seeking by priming injections of alcohol and exposure to stress in rats

 Previous studies using a reinstatement procedure have found that acute reexposure to the self-administered drug and exposure to footshock stress reinstate heroin and cocaine seeking after prolonged drug-free periods. Here we tested whether these findings generalize to alcohol-taking behavior. Male rats were initially allowed to consume alcohol in a two-bottle choice procedure (water versus alcohol) for 30 min/day for 36 days. Rats were then trained for 60 min/day in operant chambers to press a lever for the drug (0.13 ml of 12% w/v of an alcohol solution) for up to 55 days. After stable drug-taking on a fixed-ratio-3 schedule of reinforcement was obtained, lever pressing for alcohol was extinguished by terminating drug delivery for 4–9 days. Reinstatement of drug seeking was then determined after non-contingent priming injections of alcohol (0.24 and 0.48 g/kg; given IP and orally) or exposure to intermittent footshock stress (5 and 15 min; 0.8 mA). Priming injections of alcohol produced a modest dose-dependent reinstatement of drug seeking, whereas footshock stress potently reinstated extinguished alcohol seeking. In contrast, similar parameters of footshock failed to reinstate extinguished sucrose-taking behavior in rats previously trained to lever press for sucrose pellets. These findings extend previous reports on reinstatement of cocaine and heroin seeking by a footshock stressor and by priming drug injections. It also appears that the reinstatement procedure provides an appropriate methodology to study relapse to alcohol-taking behavior in the drug-free state. Received: 9 April 1997 / Final version: 1 August 1997     

Increased accumbal dopamine during daily alcohol consumption and subsequent aggressive behavior in rats

Background Alcohol drinking may lead to increased aggression in certain individuals, and both fighting and drinking increase levels of dopamine and serotonin in mesocorticolimbic structures. Assessing the dynamic changes in these neurotransmitters during the course of drinking and fighting has remained challenging. Objective The objective of the study was to learn about ongoing monoaminergic activity in the nucleus accumbens of rats that engaged in aggressive behavior after having consumed low doses of alcohol. Materials and methods After male members of breeding pairs of Long–Evans rats displayed reliable aggression toward an intruder into their home cage, they were trained to consume a 10% alcohol solution, leading to blood alcohol levels of 20–80 mg/dl. Subsequently, the effect of daily alcohol self-administration on aggression was determined in biweekly confrontations with an intruder. Finally, rats were implanted with a microdialysis probe aimed at the n. accumbens for sample collection before, during, and after a 10-min alcohol drinking session followed by a 10-min aggressive confrontation. Results Accumbal dopamine, but not serotonin, levels tended to increase in anticipation of the daily alcohol session, reaching significance immediately after the alcohol session and remaining significantly elevated (by 40%) during and after the subsequent confrontation. No such changes were seen in residents that confronted an intruder without preceding alcohol consumption. Animals that had a history of becoming more aggressive after consumption of low levels of alcohol showed similar changes in dopamine levels as did animals that had no such history. Conclusions The rise in accumbal dopamine confirms previous findings and seems to reflect the anticipation of alcohol consumption; it persisted during the aggressive confrontation regardless of the level of aggression. The daily alcohol drinking for several months may have facilitated dopamine release and masked any further changes associated with the aggressive encounter.     

Long-term alcohol consumption alters dorsal striatal dopamine release and regulation by D2 dopamine receptors in rhesus macaques

The dorsal striatum (DS) is implicated in behavioral and neural processes including action control and reinforcement. Alcohol alters these processes in rodents, and it is believed that the development of alcohol use disorder involves changes in DS dopamine signaling. In nonhuman primates, the DS can be divided into caudate and putamen subregions. As part of a collaborative effort examining the effects of long-term alcohol self-administration in rhesus macaques, we examined DS dopamine signaling using fast-scan cyclic voltammetry. We found that chronic alcohol self-administration resulted in several dopamine system adaptations. Most notably, dopamine release was altered in a sex- and region-dependent manner. Following long-term alcohol consumption, male macaques, regardless of abstinence status, had reduced dopamine release in putamen, while only male macaques in abstinence had reduced dopamine release in caudate. In contrast, female macaques had enhanced dopamine release in the caudate, but not putamen. Dopamine uptake was also enhanced in females, but not males (regardless of abstinence state). We also found that dopamine D2/3 autoreceptor function was reduced in male, but not female, alcohol drinkers relative to control groups. Finally, we found that blockade of nicotinic acetylcholine receptors inhibited evoked dopamine release in nonhuman primates. Altogether, our findings demonstrate that long-term alcohol consumption can sex-dependently alter dopamine release, as well as its feedback control mechanisms in both DS subregions.Subject terms: Addiction, Reward     

Sex differences in the motor inhibitory and stimulatory role of dopamine D1 receptors in rats

We investigated sex differences in the motor responses to the full and selective dopamine D1-like receptor agonist, (+/-)-6-chloro-7,8-dihydroxyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrobromide (SKF-81297; 0.3, 3, and 10 mg/kg, s.c.), in non-habituated adult rats. In general, SKF-81297 produced a biphasic effect on motor activity (including locomotion, rearing and exploratory activity) which consisted of an initial short inhibition followed by a long-lasting stimulation. These effects were dose- and sex-dependent. The inhibitory phase was more pronounced in males than females while the opposite was true for the stimulatory phase. Importantly, the motor inhibitory effects of SKF-81297 were not due to an increase in stereotypy (e.g., grooming activity). These biphasic effects on several motor parameters suggest the presence of two distinct dopamine D1 receptor populations which have opposite effects on motor activity and which are, in part, sexually dimorphic.     

Apomorphine-induced behavioural sensitization in rats: individual differences, role of dopamine and NMDA receptors

Apomorphine-induced behavioural sensitization was studied in male Wistar rats. The acute administration of apomorphine (0.5 mg/kg s.c.), a dopamine agonist, did not affect the locomotor activity of rats, but it caused stereotyped behaviour characterized by repeated gnawing, licking and sniffing. A significant increase in the locomotor activity became evident after repeated treatments with apomorphine (0.5 mg/kg twice daily for 14 days). However, there were marked individual differences in the sensitization of rats to apomorphine. One third of animals did not react with increased locomotor activity even after the 2-week administration of apomorphine, whereas the other one third needed only a few injections to display increased behavioural response to apomorphine. The behavioural response of the remaining one third of rats was between weak and strong responders. Simultaneously, the stereotyped behaviour occurred earlier and its intensity tended to be lower after repeated treatment with apomorphine. Nevertheless, the established changes of stereotyped behaviour did not correlate with the increase of locomotor activity. The administration of amphetamine (2.5 mg/kg, s.c.), an indirect dopamine agonist, but not a non-competitive NMDA antagonist dizocilpine (0.25 mg/kg i.p.), tended to cause a similar response profile with apomorphine in sensitized rats. The ED₅₀ values of the dopamine antagonists blocking apomorphine-induced increase in the locomotor activity were the following: 0.09 mg/kg for raclopride (dopamine D₂ antagonist), 0.023 mg/kg for SCH 23390 (dopamine D₁ antagonist), 6.42 mg/kg for clozapine (dopamine D₄ antagonist). This supports the involvement of D₁ and D₂ receptors in the expression of apomorphine-induced behavioural sensitization. The concomitant administration of dizocilpine (0.5 mg/kg), SCH 23390 (0.05 mg/kg), raclopride (0.1 mg/kg) and clozapine (20 mg/kg) with apomorphine (0.5 mg/kg twice daily for 2 weeks) antagonized the development of behavioural sensitization to apomorphine. Accordingly, at least three different molecular targets, namely dopamine D₁ and D₂, and NMDA receptors, are involved in the development of apomorphine-induced behavioural sensitization.     

Combined antagonism of glutamate mGlu5 and adenosine A2A receptors interact to regulate alcohol-seeking in rats

Adenosine and glutamate have been implicated as mediators involved in the self-administration of alcohol. In the present study we sought to determine whether adenosine receptors could interact with metabotropic glutamate receptors to regulate operant responding for alcohol and also the integration of the salience of alcohol-paired cues. Alcohol-preferring (iP) rats were trained to self-administer alcohol under operant conditions. The availability of alcohol was paired with an olfactory cue plus a stimulus light. Rats were examined under fixed ratio responding and also following extinction under a cue-induced reinstatement paradigm. Administration of the selective adenosine A2A receptor antagonist, SCH 58261, reduced fixed ratio responding for alcohol in iP rats in a dose-related manner. Furthermore, the combination of a subthreshold dose of SCH 58261 with a subthreshold dose of the mGlu5 receptor antagonist MTEP also reduced alcohol self-administration and increased the latency to the first reinforced response, suggesting a pre-ingestive effect. Moreover, this combination of SCH 58261 and MTEP also prevented the conditioned reinstatement of alcohol-seeking elicited by the re-presentation of cues previously paired with alcohol availability. In contrast, combinations of the selective adenosine A1 receptor antagonist, DPCPX, with either SCH 58261 or MTEP had no effect on alcohol responding. Collectively, these data suggest a functional interaction between adenosine A2A and mGlu5 receptors in relation to alcohol-seeking and the integration of the drug-related cues.     

Lateralization of neocortical dopamine receptors and dopamine level in normal Wistar rats

A lateral asymmetry of dopamine receptors (D-1, D-2) and dopamine (DA) level was found in the left and right neocortices of male Wistar rats. The specific D-1 receptor binding (3H-SCH 23390) was by 24% (p less than 0.05), the specific D-2 receptor binding (3H-spiperone)----by 31% (0.1 greater than p greater than 0.05) and the DA level ---- by 80% higher (p less than 0.001) in the right than in the left neocortex. In the other examined brain regions (striatum and limbic system) no lateralization in D-1 or D-2 receptors was observed. The physiological significance of the neocortical lateralization (DA receptors and level) is unknown, yet it is important for experimental and methodological reasons.     

Severity of alcohol withdrawal symptoms depends on developmental stage of Long-Evans rats

To investigate alcohol dependency and the potential role of age of initial alcohol consumption, Long-Evans (LE) rats were fed an ethanol-containing liquid diet starting at postnatal (P) ages (days): P23-27 (juvenile), P35-45 (adolescent) or P65-97 (young adult). Severity of subsequent withdrawal symptoms was dependent on age when consumption began and on duration of alcohol consumption. Frequency of withdrawal seizures was highest for rats starting consumption as juveniles, intermediate for adolescents and lowest for adults. Normalized to body weight, alcohol consumption was significantly higher for adolescent and juvenile rats than for adults. Sprague-Dawley rats that began alcohol consumption as adolescents (P35) had a level of alcohol consumption identical to that of the adolescent LE rats but showed much lower frequency of withdrawal seizures when tested after 2, 3 and 5 weeks of alcohol consumption. Based on several indicators, the capacity of the juveniles to metabolize ethanol is equal to or exceeds that of adults. Recoveries from a single dose of ethanol (2.5 g ethanol/kg body weight) were faster for juvenile LE rats than adults. The rate of decline in blood ethanol concentration was identical for juvenile and adult rats while the corrected ethanol elimination rate was higher for juveniles. The primary isozyme of alcohol dehydrogenase (ADH) in rat liver, ADH-3, had a similar Km and higher activity in liver preparations from juveniles. In conclusion, LE rats beginning alcohol consumption as juveniles or adolescents develop a severe alcohol withdrawal syndrome that may not be attributed entirely to higher levels of consumption and was not explained by any obvious deficiencies in metabolism.     

Sex differences in the contribution of nicotine and nonpharmacological stimuli to nicotine self-administration in rats

Rationale Sex differences have been reported for the impact of nicotine and nonpharmacological cues on smoking. While nonpharmacological environmental stimuli have also been shown to influence nicotine self-administration in rats, there have been no attempts to examine the impact of sex differences in the contributions of nicotine and nondrug stimuli to this behavior.Objectives This experiment investigated sex differences in operant responding for nicotine in rats when drug infusions were delivered either in the absence of, or in combination with, a nonpharmacological stimulus.Methods Initially, male and female rats acquired self-administration for nicotine alone across a range of doses (0.03, 0.06, and 0.15 mg kg^–1 inf^–1, freebase). After stable acquisition, nicotine infusions were combined with a weakly reinforcing, compound visual stimulus.Results While there was no overall effect of dose on active lever responding for nicotine in the absence of the visual stimulus, female rats responded more on the reinforced lever than males at 0.06 and 0.15 mg kg^–1 inf^–1 on an FR5 schedule. However, they also showed increased responding on the nonreinforced lever compared to males at the same doses. Combining nicotine infusions with the visual stimulus doubled responding compared to nicotine alone at 0.03 and 0.06, but not at 0.15 mg kg^–1 inf^–1: this effect was significantly greater for female rats.Conclusions These data highlight the prominent contribution of nonpharmacological stimuli to nicotine-reinforced behavior across a range of doses in both male and female rats. They also reveal sex differences in operant responding for nicotine under conditions where a nonpharmacological stimulus is either absent, or combined with drug delivery.     

Increase in the hypotensive effect of bromocriptine induced by spinal transection in rats: contribution of spinal dopamine receptors.

Intravenous (i.v.) administration of bromocriptine (150 micrograms/kg) in conscious normotensive rats with chronic spinal cord transection (at T5-T7), pretreated or not with i.v. propranolol (0.5 mg/kg), induced significant decreases in mean arterial blood pressure (MAP) which were greater and longer lasting than those in intact rats (-15 to -20 as compared with -10 mm Hg) for 8 days after transection. To assess the spinal and/or peripheral origin of this phenomenon, rats were also pretreated with either i.v. (0.3 mg/kg) or intrathecal (i.t.; 93 nmol/rat; at T9-T10) administration of domperidone, a selective dopamine (DA)2 receptor antagonist incapable of crossing the blood-brain barrier (BBB) freely. The increase in hypotension induced by spinal section was suppressed by i.t. but not by i.v. domperidone. In intact rats, bromocriptine elicited an increase in heart rate (HR; approximately 50 beats/min more), which was prevented by i.v. propranolol treatment. In spinal cord-transected rats, however, it had a significant bradycardic effect (approximately 50 beats/min less), which was antagonized by i.t.-administered domperidone. These results suggest that enhancement of the hypotensive effects induced by systemic administration of bromocriptine after a complete thoracic spinal transection is fully mediated by spinal DA2 receptors. This finding may help explain the increased orthostatic hypotension induced by DA receptor agonists in Parkinsonian patients with spinal lesions.     

Effects of prenatal alcohol consumption on open-field behaviour and alcohol preference in rats

Throughout gestation pregnant Wistar rats consumed a liquid diet containing sustagen and ethanol. Control mothers were fed on lab chow. Subsequently, the offspring of the ethanol-fed mothers displayed significantly greater activity (ambulation and rearing) in an open-field test. Further, pups whose mothers were exposed to ethanol consumed significantly more ethanol in a preference test, compared to control offspring.     

Taste aversion learning and schedule-induced alcohol consumption in rats

Twelve hungry rats were exposed to the intermittent delivery of food with a 5% (v/v) ethanol solution freely available. All developed high levels of schedule-induced alcohol polydipsia within ten 1-hr sessions. Immediately following the eleventh session half of the animals received an intraperitoneal injection of lithium chloride (experimental group) and the other half received an injection of sodium chloride (control group). Their alcohol intakes did not differ during the twelfth session and so the treatments were repeated following the thirteenth session. During the fourteenth session, the experimental group drank very little alcohol compared to the control group, indicating that they had learned a taste-aversion to the alcohol in only two conditioning trials. These results extend previous work on the role of taste aversion in suppressing alcohol intake by demonstrating that the technique can be used to suppress schedule-induced alcohol polydipsia as well as thirst-motivated alcohol intake.     

Stress and conflict conditions leading to and maintaining voluntary alcohol consumption in rats

Four experiments were conducted to investigate the effects of unavoidable shock, conflict conditions, taste, and food deprivation on the voluntary consumption of alcohol by rats. Experiment 1 showed that when rats were given unavoidable shocks for one hour every day, those living in their home cages consumed greater amounts of a 5% ethanol solution than did rats living in the shock chambers. Experiment 2 revealed that this increased alcohol consumption was maintained and further elevated when these same rats were subjected to conflict, and it did not decrease when the conflict conditions were terminated. When the unavoidable shock conditions were repeated in Experiment 3 with naive rats and the fluid choice consisted of a plain sucrose solution and one containing alcohol, rats in both the shock box and safety cage living conditions consumed very little of the sucrose-plus-alcohol solution. Rats living in the aversive environment even decreased consumption of the plain sucrose solution. Experiment 4 showed that simple food deprivation can also result in an increased intake of an alcohol solution. The tension reduction hypothesis cannot account for these results: they demonstrate that deprivation can influence alcohol consumption, and indicate that an aversive environment can interfere with drinking of any solution. The results also demonstrate both the positive and negative properties that alcohol can have.     

Individual differences in acute alcohol impairment of inhibitory control predict ad libitum alcohol consumption

Rationale  Research has begun to examine how acute cognitive impairment from alcohol could contribute to alcohol abuse. Specifically, alcohol-induced impairment of inhibitory control could compromise the drinker’s ability to stop the self-administration of alcohol, increasing the risk of binge drinking. Objective  The present study was designed to test this hypothesis by examining the relation between acute alcohol impairment of inhibitory control and alcohol consumption during a single drinking episode. Materials and methods  Twenty-six healthy adults performed a cued go/no-go task that measured inhibitory control. The study tested the degree to which their inhibitory control was impaired by a moderate dose of alcohol (0.65 g/kg) versus a placebo and the extent to which individual differences in this impairment predicted levels of alcohol consumption as assessed by ad lib drinking in the laboratory. Results  In accord with the hypothesis, greater impairment of inhibitory control from alcohol was associated with increased ad lib consumption. Conclusion  Acute impairment of inhibitory control might be an important cognitive effect that contributes to abuse in addition to the positive rewarding effects of the drug.     

Lithium effects on adjunctive alcohol consumption in rats

Rats receiving chronic administration of lithium chloride (20 mEq/l) in their drinking water were tested for acquisition of adjunctive alcohol (10% v/v) consumption. Contrary to expectations, subjects receiving lithium acquired the adjunctive drinking more rapidly, and under less optimal conditions, than did control subjects. The high death rate in subjects receiving lithium while undergoing a concurrent alcohol withdrawal suggests that particular caution must be observed when lithium is used in the treatment of alcoholics.     

The failure of p-chlorophenylalanine to affect voluntary alcohol consumption in rats.

1 The effects of p-chlorophenylalanine (PCPA) administered orally and intraperitoneally on rat brain 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) content were compared. The depletion of brain 5-HT and 5-HIAA following PCPA (316 mg/kg) injected intraperitoneally every third day was not significantly different from that following the administration of PCPA (316 mg/kg) by stomach tube on eight consecutive days. 2 Rats tested for alcohol preference before, during and after treatment with PCPA on two intraperitoneal dose regimens (either 316 mg/kg then 100 mg/kg four days later or 316 mg/kg three times at intervals of three days) showed no reduction in voluntary alcohol consumption. 3 The results indicated that depletion of brain 5-HT and 5-HIAA is not responsible for the reduction of voluntary alcohol intake which has been reported to follow chronic oral administration of PCPA to the rat (Myers & Veale, 1968). The possibility of a learned aversion to alcohol due to an association with PCPA administration is discussed.     

Contribution of spinal dopamine receptors to the hypotensive action of bromocriptine in rats.

Bromocriptine, a dopamine (DA) receptor agonist, has been reported to have hypotensive effects in anesthetized and conscious normotensive rats but its mechanism of action is still not fully understood. Therefore, we studied the changes in mean arterial blood pressure (MAP) and heart rate (HR) elicited by an intravenous (i.v.) administration of bromocriptine (150 micrograms/kg), in either pentobarbital-anesthetized or conscious normotensive rats, pretreated with either i.v. (0.3 mg/kg) or intrathecal (i.t.) (93 nmol) domperidone, a DA receptor antagonist that does not cross the blood-brain barrier. In these preparations, i.v. administration of bromocriptine elicited dose-dependent decreases in MAP and rises in HR. The hypotensive effect was antagonized partially by i.t. and fully by i.v. domperidone. However, the latter compound did not modify the tachycardia, which could be blocked by propranolol (0.5 mg/kg i.v.). In rats pretreated with the latter beta-adrenoceptor antagonist, bromocriptine produced only a decrease in blood pressure that was inhibited by i.v. and i.t. domperidone. These results suggest that, in anesthetized and conscious normotensive rats, the hypotension induced by systemic administration of bromocriptine is fully mediated by DA2 dopamine receptors, which are located partly within the spinal cord and partly in the peripheral circulation.     

Dietary caffeine and alcohol consumption by rats.

The alcohol consumption of malnourished rats increased slowly but substantially during a 4-week period in which caffeine was added to their marginally adequate diet. Consumption fell to precaffeine levels as soon as caffeine was withdrawn.