• Energy Research

Ethanol and nicotine are commonly coabused drugs, and the incidence of codependence is greater than would be expected on the basis of the summed probability of dependence on each drug alone. Previous findings from our laboratory and others suggest that interactive mechanisms at the level of discriminative stimulus (S(D)) effects may contribute to this coabuse phenomenon. Specifically, ethanol overshadows the nicotine S(D) whereas nicotine potentiates the stimulus salience of ethanol when the two drugs are conditioned as a drug mixture. The goal of the current study was to begin to delineate the pharmacological bases of these ethanol-nicotine interactions. Three groups of C57BL/6J mice were trained to discriminate 0.8 mg/kg nicotine + 0.5 g/kg ethanol (0.8 N + 0.5 E), 0.8 N + 1.0 E, or 0.8 N + 2.0 E. An NMDA receptor antagonist (MK-801) and three nACh receptor ligands were tested for their ability to generalize from or antagonize, respectively, the drug mixtures. MK-801 fully generalized from the 0.8 N + 1.0 E and 0.8 N + 2.0 E mixtures and partially generalized from 0.8 N + 0.5 E. In contrast, nACh receptor ligands had minimal influence in blocking the perception of 0.8 N + 1.0 E and 0.8 N + 2.0 E mixtures, and only mecamylamine partially blocked 0.8 N+0.5 E. Reduced and enhanced contributions of nACh and NMDA receptors, respectively, in the discrimination of ethanol-nicotine mixtures may contribute to the overshadowing and potentiation phenomena observed previously.

Drinking to intoxication or binge drinking is a hallmark characteristic of alcohol abuse. Although hard to model in rodents, the scheduled high alcohol consumption (SHAC) procedure generates high, stable ethanol intake and blood ethanol concentrations in mice to levels consistent with definitions of binge drinking. The purpose of the present studies was to determine the effects of pharmacological manipulation of the opioidergic, glutamatergic, and γ-aminobutyric acid (GABA)ergic systems on binge drinking with the SHAC procedure. Parallel manipulations were conducted in mice trained in operant self-administration of either sucrose or ethanol. For the SHAC procedure, genetically heterogeneous Withdrawal Seizure Control mice were given varying periods of fluid access, with a 30-min ethanol session every third day (total of seven). Mice were pretreated intraperitoneally with naltrexone (0, 0.6, or 1.25 mg/kg), baclofen (0, 2.5, or 5.0 mg/kg), or 2-methyl-6-(phenylethynyl)-pyridine (MPEP; 0, 3.0, or 10.0 mg/kg) before each ethanol session. For the operant self-administration procedure, separate groups of C57BL/6 mice were trained to complete a single response requirement (16 presses on the active lever) to gain 30 min of access to an ethanol or a sucrose solution. Mice received pretreatments of the same doses of naltrexone, MPEP, or baclofen before the self-administration sessions, with saline injections on intervening days. Naltrexone produced a dose-dependent decrease in binge drinking, and the highest dose also significantly decreased operant self-administration of ethanol and sucrose. Both doses of baclofen significantly decreased binge alcohol consumption, but the higher dose also tended to decrease water intake. The highest dose of baclofen also significantly decreased operant self-administration of sucrose. MPEP (10 mg/kg) significantly decreased binge alcohol consumption and sucrose self-administration. These results indicate that manipulation of the opioidergic, glutamatergic, and GABAergic systems significantly decreased binge drinking.

Although systemic allopregnanolone (ALLO; a positive modulator of GABA(A) receptors) has been shown to enhance ethanol-reinforced responding and to modulate drinking patterns in rodents, the effects of centrally administered ALLO on ethanol intake are not known. The current work examined the effects of intracranial ALLO on operant ethanol self-administration in food- and water-satiated mice, with a procedure designed to estimate ALLO's influence on appetitive versus consummatory processes. Male C57BL/6J (B6) mice were trained to press an ethanol-appropriate lever by being reinforced with 30-min of continuous access to a 10% ethanol solution. Following surgical implantation of a guide cannula aimed at the lateral ventricle and subsequent habituation to vehicle infusions, ALLO (50-400 ng; ICV) was delivered immediately prior to session start. ALLO doses of 100 and 400 ng were further evaluated for their effects on locomotor behavior within activity chambers. ALLO selectively modulated ethanol intake patterns associated with the onset and maintenance of self-administration, while leaving appetitive (i.e., ethanol seeking) measures unaltered. The effects of ALLO on drinking patterns were dissociable from changes in locomotor behavior, as evidenced by the absence of ALLO's influence on response frequency and horizontal distance traveled. These findings support the premise that manipulations in brain ALLO levels may influence the regulatory processes governing ethanol consumption.

The neurosteroid allopregnanolone (ALLO) is a positive modulator of GABA(A) receptors that exhibits a psychopharmacological profile similar to ethanol (i.e., anxiolytic, sedative-hypnotic). Based on research suggesting that manipulation of ALLO levels altered ethanol self-administration in male rodents, the current studies determined whether exogenous ALLO administration or the inhibition of its synthesis in vivo modulated ethanol intake patterns in female C57BL/6J mice. Lickometer circuits collected temporal lick records of ethanol (10%, v/v) and water consumption during daily 2h limited access sessions. Following the establishment of stable ethanol intake, studies examined the effect of an acute ALLO challenge (3.2-24.0 mg/kg) or a 7-day blockade of ALLO production with finasteride (FIN; 50 or 100 mg/kg) on ethanol intake in a within-subjects design. In contrast to results in male mice, ethanol dose (g/kg), ethanol preference and most of the bout parameters were unaltered by ALLO pretreatment in female mice. Ethanol intake in females also was recalcitrant to 7-day treatment with 50 mg/kg FIN, whereas 100 mg/kg FIN significantly reduced the ethanol dose consumed by 35%. The FIN-attenuated ethanol intake was attributable to a significant decrease in bout frequency (up to 45%), with lick patterns indicating reduced maintenance of consumption throughout the 2-h session. FIN also produced a dose-dependent decrease in brain ALLO levels. In conjunction with data in male mice, the present findings indicate that there are sex differences in the physiological regulation of ethanol intake patterns by GABAergic neurosteroids.

Manipulation of endogenous levels of the GABAergic neurosteroid allopregnanolone alters sensitivity to some effects of ethanol. Chronic ethanol withdrawal decreases activity and expression of 5α-reductase-1, an important enzyme in allopregnanolone biosynthesis encoded by the 5α-reductase-1 gene (Srd5a1). The present studies examined the impact of Srd5a1 deletion in male and female mice on several acute effects of ethanol and on chronic ethanol withdrawal severity. Genotype and sex did not differentially alter ethanol-induced hypothermia, ataxia, hypnosis, or metabolism, but ethanol withdrawal was significantly lower in female versus male mice. On the elevated plus maze, deletion of the Srd5a1 gene significantly decreased ethanol's effect on total entries versus wildtype (WT) mice and significantly decreased ethanol's anxiolytic effect in female knockout (KO) versus WT mice. The limited sex differences in the ability of Srd5a1 genotype to modulate select ethanol effects may reflect an interaction between developmental compensations to deletion of the Srd5a1 gene with sex hormones and levels of endogenous neurosteroids.

Recent evidence suggests that GABA(A) receptor ligands may regulate ethanol intake via effects at both synaptic and extrasynaptic receptors. For example, the endogenous neurosteroid, allopregnanolone (ALLO) has a similar pharmacological profile as ethanol, and it alters ethanol intake in rodent models. Additionally, recent evidence suggests that δ-subunit-containing extrasynaptic GABA(A) receptors may confer high sensitivity to both ethanol and neurosteroids. The purpose of the present study was to determine the effects of ganaxolone (GAN; an ALLO analog) and gaboxadol (THIP; a GABA(A) receptor agonist with selectivity for the extrasynaptic δ-subunit) on ethanol intake, drinking patterns, and bout characteristics in operant and limited-access self-administration procedures. In separate studies, the effects of GAN (0-10 mg/kg) and THIP (2-16 mg/kg) were tested in C57BL/6J male mice provided with 2-h access to a two-bottle choice of water or 10% ethanol or trained to respond for 30 min of access to 10% ethanol. GAN had no overall significant effect on operant ethanol self-administration, but tended to decrease the latency to consume the first bout. In the limited-access procedure, GAN dose-dependently decreased ethanol intake. THIP dose-dependently decreased ethanol intake in both paradigms, altering both the consummatory and appetitive processes of operant self-administration as well as shifting the drinking patterns in both procedures. These results add to literature suggesting time-dependent effects of neurosteroids to promote the onset, and to subsequently decrease, ethanol drinking behavior, and they support a role for extrasynaptic GABA(A) receptor activation in ethanol reinforcement.

The neuroactive steroid allopregnanolone (ALLO) is a positive modulator of GABAA receptors, and manipulation of neuroactive steroid levels via injection of ALLO or the 5α-reductase inhibitor finasteride alters ethanol self-administration patterns in male, but not female, mice. The Srd5a1 gene encodes the enzyme 5α-reductase-1, which is required for the synthesis of ALLO. The current studies investigated the influence of Srd5a1 deletion on voluntary ethanol consumption in male and female wildtype (WT) and knockout (KO) mice. Under a continuous access condition, 6 and 10 % ethanol intake was significantly greater in KO versus WT females, but significantly lower in KO versus WT males. In 2-h limited access sessions, Srd5a1 deletion retarded acquisition of 10 % ethanol intake in female mice, but facilitated it in males, versus respective WT mice. The present findings demonstrate that the Srd5a1 gene modulates ethanol consumption in a sex-dependent manner that is also contingent upon ethanol access condition and concentration.

Schedule-induced polydipsia (SIP) is generated by subjecting a highly motivated animal to a sub-optimal rate of food reinforcement while also providing access to a fluid. SIP is one of several adjunctive (or displacement) behaviors that are expressed in an exaggerated form that is deemed 'excessive.' This feature makes SIP an attractive model for studying an excessive ethanol drinking phenotype in rodents. Multiple experimental variables are crucial for the full manifestation of adjunctive drinking, including the degree of food deprivation, the inter-pellet interval selected, and the size of the food reward offered. Although these variables were extensively studied and optimized for water polydipsia in rats, a similarly customized approach to ethanol SIP and application of the procedure in mice have largely been curtailed in favor of the default variable values historically used for water SIP in rats. Further, ethanol SIP also requires careful consideration of variables such as taste and ethanol concentration. Investigation of the stress axis and neurochemical systems such as dopamine and serotonin in mediating adjunctive drinking stemmed from two leading hypotheses regarding the underlying mechanisms of SIP generation: 1) SIP as a coping strategy to mitigate stress associated with the aversive environmental condition, and 2) SIP as a displacement of reward in a highly motivated animal. Ethanol SIP is a powerful model of excessive intake because it can generate an ethanol-dependent state and sustain frequent and intoxicating levels of blood ethanol with voluntary oral consumption. The required food deprivation and the loss of the excessive drinking phenotype following removal of the generator schedule are the two main limitations of the model. Future utility of ethanol SIP will be enhanced by more fully dissecting the underlying hormonal and neurochemical mechanisms and optimizing experimental variables for ethanol SIP on a per species and strain basis.