Binge-like Ethanol Consumption Research Articles

A paradigm for ethanol consumption in head-fixed mice during prefrontal cortical two-photon calcium imaging

The prefrontal cortex (PFC) is a hub for cognitive behaviors and is a key target for neuroadaptations in alcohol use disorders. Recent advances in genetically encoded sensors and functional microscopy allow multimodal in vivo PFC activity recordings at subcellular and cellular scales. While these methods could enable a deeper understanding of the relationship between alcohol and PFC function/dysfunction, they typically require animals to be head-fixed. Here, we present a method in mice for binge-like ethanol consumption during head-fixation. Male and female mice were first acclimated to ethanol by providing home cage access to 20% ethanol (v/v) for 4 or 8 days. After home cage drinking, mice consumed ethanol from a lick spout during head-fixation. We used two-photon calcium imaging during the head-fixed drinking paradigm to record from a large population of PFC neurons (>1000) to explore how acute ethanol affects their activity. Drinking exerted temporally heterogeneous effects on PFC activity at single neuron and population levels. Intoxication modulated the tonic activity of some neurons while others showed phasic responses around ethanol receipt. Population level activity did not show tonic or phasic modulation but tracked ethanol consumption over the minute-timescale. Network level interactions assessed through between-neuron pairwise correlations were largely resilient to intoxication at the population level while neurons with increased tonic activity showed higher synchrony by the end of the drinking period. By establishing a method for binge-like drinking in head-fixed mice, we lay the groundwork for leveraging advanced microscopy technologies to study alcohol-induced neuroadaptations in PFC and other brain circuits.This article is part of the Special Issue on "PFC circuit function in psychiatric disease and relevant models".

Basolateral amygdala neuropeptide Y system modulates binge ethanol consumption.

Neuropeptide Y (NPY) signaling regulation of corticolimbic communication is known to modulate binge-like ethanol consumption in rodents. In this work we sought to assess the impact of intra-BLA NPY system modulation on binge-like ethanol intake and to assess the role of the NPY1R+ projection from the BLA to the mPFC in this behavior. We used "drinking-in-the-dark" (DID) procedures in C57BL6J mice to address these questions. First, the impact of intra-BLA administration of NPY on binge-like ethanol intake was assessed. Next, the impact of repeated cycles of DID intake on NPY1R expression in the BLA was assessed with use of immunohistochemistry (IHC). Finally, chemogenetic inhibition of BLA→mPFC NPY1R+ projections was assessed to determine if limbic communication with the mPFC was specifically involved in binge-like ethanol intake. Importantly, as both the BLA and NPY system are sexually dimorphic, both sexes were assessed in these studies. Intra-BLA NPY dose-dependently decreased binge-like ethanol intake in males only. Repeated DID reduced NPY1R expression in the BLA of both sexes. Silencing of BLA→mPFC NPY1R+ neurons significantly reduced binge-like ethanol intake in both sexes in a dose-dependent manner. We provide novel evidence that (1) intra-BLA NPY reduces binge-like ethanol intake in males; (2) binge-like ethanol intake reduces NPY1R levels in the BLA; and (3) chemogenetic inhibition of BLA→mPFC NPY1R+ neurons blunts binge-like drinking in male and female mice. These observations provide the first direct evidence that NPY signaling in the BLA, and specifically BLA communication with the mPFC, modulates binge-like ethanol consumption.

Somatostatin signaling modulates binge drinking behavior via the central nucleus of the amygdala

Somatostatin (SST) is a neuropeptide widely expressed in the central nervous system with dense expression in limbic regions such as the extended amygdala. It has recently gained attention for playing a role in modulating alcohol use disorders and co-morbid neuropsychiatric disorders. However, the role of SST in the central nucleus of the amygdala (CeA), a key region for neuropeptide regulation of alcohol and anxiety related behaviors, in alcohol consumption has not been assessed. In this work we perform an initial examination of the interaction between the CeA SST system and binge ethanol intake. Binge intake is a dangerous pattern of excessive ethanol consumption associated with health complications and the transition into alcohol dependence. We use the Drinking in the Dark (DID) model of binge intake in C57BL/6J male and female mice to examine: 1) the impact of 3 DID cycles on CeA SST expression; 2) the effect of intra-CeA SST injection on binge-like ethanol consumption; and 3) if the SST receptor 2 or 4 (SST2R or SST4R) mediate any effect on consumption. Our results show binge-like ethanol intake decreases SST expression in the CeA, but not neighboring basolateral amygdala. We further found intra-SST CeA administration reduces binge ethanol intake. This decrease was replicated by the administration of an SST4R agonist. These effects were not sex-dependent. Overall, this work lends further support for SST playing a role in alcohol related behaviors and as a potential therapeutic target.

Selective PDE4B and PDE4D inhibitors produce distinct behavioral responses to ethanol and GABAergic drugs in mice

Apremilast is a phosphodiesterase (PDE) type 4 inhibitor that is nonselective at subtypes PDE4A-D. It modulates ethanol and GABAergic responses via protein kinase A (PKA) phosphorylation of specific GABAA receptor subunits and has opposite effects on ethanol-induced ataxia in wild-type and GABAA β3-S408/409A knock-in mice. We hypothesized that these different effects are due to preferential actions at different PDE4 subtypes. To test this hypothesis, we compared effects of selective PDE4 inhibitors on responses to ethanol and GABAergic drugs in male and female C57BL/6J mice. The PDE4B inhibitor A33 accelerated recovery from ataxia induced by ethanol and diazepam but did not alter ataxia induced by propofol. The PDE4D inhibitor D159687 accelerated recovery from diazepam-induced ataxia but prolonged recovery from ethanol- and propofol-induced ataxia. A33 shortened, while D159687 prolonged, the sedative-hypnotic effects of ethanol. Both drugs shortened diazepam's sedative-hypnotic effects. The modulatory effects of A33 and D159687 were completely prevented by the PKA inhibitor H89. Only D159687 prevented development of acute functional tolerance to ethanol-induced ataxia. D159687 transiently reduced two-bottle choice drinking in male and female mice that had consumed ethanol for 3 weeks and transiently reduced two-bottle choice, every-other-day drinking in male mice. A33 did not alter ethanol drinking in either procedure. Neither drug altered binge-like ethanol consumption or blood ethanol clearance. Thus, D159687 produced behavioral effects similar to apremilast, although it produced a more transient and smaller reduction in drinking. These results indicate that PDE4D inhibition contributes to apremilast's ability to reduce ethanol drinking, whereas PDE4B inhibition is not involved.

Chemogenetic inhibition of corticotropin-releasing factor neurons in the central amygdala alters binge-like ethanol consumption in male mice.

Repetitive bouts of binge drinking can lead to neuroplastic events that alter ethanol's pharmacologic effects and perpetuate excessive consumption. The corticotropin-releasing factor (CRF) system is an example of ethanol-induced neuroadaptations that drive excessive ethanol consumption. Our laboratory has previously shown that CRF antagonist, when infused into the central amygdala (CeA), reduces binge-like ethanol consumption. The present study extends this research by assessing the effects of silencing CRF-producing neurons in CeA on binge-like ethanol drinking stemming from "Drinking in the Dark" (DID) procedures. CRF-ires-Cre mice underwent surgery to infuse Gi/o-coupled Designer Receptors Exclusively Activated by Designer Drugs (DREADD) virus or a control virus into either the CeA or basolateral amygdala (BLA). Gi/o-DREADD-induced CRF-neuronal inhibition in the CeA resulted in a 33% decrease in binge-like ethanol consumption. However, no effect on ethanol consumption was seen after DREADD manipulation in the BLA. Moreover, CeA CRF-neuronal inhibition had no effect on sucrose consumption. The effects of silencing CRF neurons in the CeA on ethanol consumption are not secondary to changes in motor function or anxiety-like behaviors as assessed in the open-field test (OFT). Finally, the DREADD construct's functional ability to inhibit CRF-neuronal activity was demonstrated by reduced ethanol-induced c-Fos following DREADD activation. Together, these data suggest that the CRF neurons in the CeA play an important role in binge ethanol consumption and that inhibition of the CRF-signaling pathway remains a viable target for manipulating binge-like ethanol consumption. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Impact of Adolescent Social Isolation on Adult, Binge-Like Ethanol Consumption and Plasma Corticosterone in High-Alcohol-Preferring Mice

Adolescent stress exposure increases the likelihood of alcohol misuse and alcohol use disorder (AUD) in adulthood; however, it is not clear how genetic and environmental factors interact to increase risk. This study examined how adolescent social isolation affects adult binge-like ethanol drinking and levels of the stress hormone corticosterone in male and female mice with a genetic predisposition toward high alcohol preference (HAP). Twenty-eight HAP mice were separated into group-housed (GH) and socially isolated (SI) conditions (n = 13, 13). Binge drinking was assessed using a drinking in the dark (DID) procedure. Blood samples were taken before DID and after the 4th (last) day of DID. Overall, adolescent social isolation increased adult binge drinking in a sex-and time-dependent manner. Analysis of Hour 1 intake across the 4 days indicated that SI males drank significantly more than GH males, and this was not the case in females. There was no significant effect of housing for Hour 2. On Day 4, after 2 hours of consumption, social isolation increased drinking regardless of sex. Plasma corticosterone (CORT) levels decreased following drinking, but there were no sex or housing group differences. There were correlations between CORT levels and drinking, but only for SI females. These findings demonstrate that adolescent social isolation promotes binge-like drinking in both male and female adult mice with a genetic predisposition for high alcohol preference; however, this relationship is time-dependent, and males may be more sensitive than females to social isolation stress. Additionally, corticosterone levels change with regard to binge-drinking and sex.

Repetitive binge-like consumption based on the Drinking-in-the-Dark model alters the microglial population in the mouse hippocampus.

Alcoholism causes various maladaptations in the central nervous system, including the neuroimmune system. Studies of alcohol-induced dysregulation of the neuroimmune system generally focus on alcohol dependence and brain damage, but our previous research indicates that repetitive binge-like consumption perturbs cytokines independent of cell death. This paper extends that research by examining the impact of binge-like consumption on microglia in the hippocampus and the amygdala. Microglia were assessed using immunohistochemistry following binge-like ethanol consumption based on Drinking-in-the-Dark model. Immunohistochemistry results showed that binge-like ethanol consumption caused an increase in Iba-1 immunoreactivity and the number of Iba-1+ cells after one Drinking-in-the-Dark cycle. However, after three Drinking-in-the-Darkcycles, the number of microglia decreased in the hippocampus. We showed that in the dentate gyrus, the average immunoreactivity/cell was increased following ethanol exposure despite the decrease in number after three cycles. Likewise, Ox-42, an indicator of microglia activation, was upregulated after ethanol consumption. No significant effects on microglia number or immunoreactivity (Iba-1 nor Ox-42) were observed in the amygdala. Finally, ethanol caused an increase in the expression of the microglial gene Aif-1 during intoxication and ten days into abstinence, suggesting persistence of ethanol-induced upregulation of microglial genes. Altogether, these findings indicate that repetitive binge-like ethanol is sufficient to elicit changes in microglial reactivity. This altered neuroimmune state may contribute to the development of alcohol use disorders.

Adolescent Ethanol Exposure Alters Cholinergic Function and Apical Dendritic Branching Within the Orbital Frontal Cortex

During adolescence, heavy binge-like ethanol consumption can lead to frontocortical structural and functional impairments. These impairments are likely driven by adolescence being a critical time point for maturation of brain regions associated with higher-order cognitive functioning. Rodent models of heavy binge-like ethanol exposure show consistent disruptions to the typical development of the prefrontal cortex (PFC). All deep cortical layers receive cholinergic projections that originate from the Nucleus basalis of Meynert (NbM) complex. These cholinergic projections are highly involved in learning, memory, and attention. Adolescent intermittent ethanol exposure (AIE) induces cholinergic dysfunction as a result of an epigenetic suppression of the genes that drive the cholinergic phenotype. The current study used a model of AIE to assess structural and functional changes to the frontal cortex and NbM following binge-like ethanol exposure in adolescence. Western blot analysis revealed long-term disruptions of the cholinergic circuit following AIE: choline acetyltransferase (ChAT) was suppressed in the NbM and vesicular acetylcholine transporter (VAChT) was suppressed in the orbitofrontal cortex (OFC). In vivo microdialysis for acetylcholine efflux during a spatial memory task determined changes in cholinergic modulation within the PFC following AIE. However, AIE spared performance on the spatial memory task and on an operant reversal task. In a second study, Golgi-Cox staining determined that AIE increased apical dendritic complexity in the OFC, with sex influencing whether the increase in branching occurred near or away from the soma. Spine density or maturity was not affected, likely compensating for a disruption in neurotransmitter function following AIE.

Lateral hypothalamus-projecting noradrenergic locus coeruleus pathway modulates binge-like ethanol drinking in male and female TH-ires-cre mice

A growing body of literature implicates noradrenergic (NE) signaling in the modulation of ethanol consumption. However, relatively few studies have detailed specific brain pathways that mediate NE-associated binge-like ethanol consumption. To begin to fill this gap in the literature, male and female C57BL6/J and TH-ires-cre mice underwent pharmacological and chemogenetic testing, respectively, in combination with “drinking in the dark” procedures to model binge-like consumption of ethanol or sucrose solutions. First, we showed that intraperitoneal administration of the NE reuptake inhibitor, reboxetine, blunted binge-like ethanol intake in C57BL6/J mice. Chemogenetic activation of locus coeruleus (LC) tyrosine hydroxylase (TH)-expressing neurons blunted binge-like ethanol intake regardless of sex. Chemogenetic activation of LC projections to the lateral hypothalamus (LH), a region implicated in ethanol consumption, blunted binge-like ethanol drinking without altering sucrose intake in ethanol-experienced or ethanol-naïve mice. In C57BL/6 J mice, LH-targeted microinfusion of an α1-adrenergic receptor (AR) agonist blunted binge-like ethanol intake across both sexes, while LH infusion of a β-AR agonist blunted binge-like ethanol intake in females exclusively. Finally, in mice with high baseline ethanol intake both an α1- AR agonist and an α-2 AR antagonist blunted binge-like ethanol intake. The present results provide novel evidence that increased NE tone in a circuit arising from the LC and projecting to the LH reduces binge-like ethanol drinking in mice, and may represent a novel approach to treating binge or heavy drinking prior to the development of dependence.This article is part of the special Issue on “Neurocircuitry Modulating Drug and Alcohol Abuse”.

Effect of chronic binge-like ethanol consumption on subsequent cocaine reinforcement in rhesus monkeys

BackgroundAlthough most individuals with cocaine use disorder also abuse alcohol, little is known about the behavioral and pharmacological mechanisms that promote co-abuse. For example, it is unclear whether prior experience with alcohol renders individuals more sensitive to cocaine when it is subsequently experienced. MethodsThis study examined the effects of chronic ethanol consumption on subsequent cocaine reinforcement in rhesus monkeys. Six monkeys consumed 2.0 g/kg ethanol in a binge-drinking paradigm and 6 monkeys drank a non-alcoholic solution 5 days per week. After 9 months, each monkey’s sensitivity to acquiring cocaine self-administration was determined. Monkeys performed an operant response resulting in food pellet delivery under a fixed-ratio 30 schedule of reinforcement. Saline, then ascending doses of cocaine, were substituted for food pellets until a cocaine dose was reached at which the number of cocaine injections delivered differed significantly from saline injections delivered. Following acquisition, a complete cocaine dose-effect curve was generated to determine whether ethanol consumption altered the reinforcing potency of cocaine determined by calculating the ED50 of the ascending limb of each subject’s curve. ResultsAlthough individual variability was observed, the cocaine dose which initially served as a reinforcer did not differ between ethanol-drinking and control groups and, within the ethanol-drinking group, was not related to the amount of ethanol consumed. Moreover, the reinforcing potency of cocaine did not differ between groups. ConclusionTaken together, the data suggest that a history of binge-like alcohol consumption does not affect sensitivity to cocaine when it is subsequently first experienced.

Binge-like ethanol drinking activates anaplastic lymphoma kinase signaling and increases the expression of STAT3 target genes in the mouse hippocampus and prefrontal cortex.

Alcohol use disorder (AUD) has a complex pathogenesis, making it a difficult disorder to treat. Identifying relevant signaling pathways in the brain may be useful for finding new pharmacological targets to treat AUD. The receptor tyrosine kinase anaplastic lymphoma kinase (ALK) activates the transcription factor STAT3 in response to ethanol in cell lines. Here, we show ALK activation and upregulation of known STAT3 target genes (Socs3, Gfap and Tnfrsf1a) in the prefrontal cortex (PFC) and ventral hippocampus (HPC) of mice after 4 days of binge-like ethanol drinking. Mice treated with the STAT3 inhibitor stattic drank less ethanol than vehicle-treated mice, demonstrating the behavioral importance of STAT3. To identify novel ethanol-induced target genes downstream of the ALK and STAT3 pathway, we analyzed the NIH LINCS L1000 database for gene signature overlap between ALK inhibitor (alectinib and NVP-TAE684) and STAT3 inhibitor (niclosamide) treatments on cell lines. These genes were then compared with differentially expressed genes in the PFC of mice after binge-like drinking. We found 95 unique gene candidates, out of which 57 had STAT3 binding motifs in their promoters. We further showed by qPCR that expression of the putative STAT3 genes Nr1h2, Smarcc1, Smarca4 and Gpnmb were increased in either the PFC or HPC after binge-like drinking. Together, these results indicate activation of the ALK-STAT3 signaling pathway in the brain after binge-like ethanol consumption, identify putative novel ethanol-responsive STAT3 target genes, and suggest that STAT3 inhibition may be a potential method to reduce binge drinking in humans.

Binge-Like Ethanol Drinking Increases Otx2, Wnt1, and Mdk Gene Expression in the Ventral Tegmental Area of Adult Mice.

Alcohol use disorder is associated with pathophysiological changes in the dopaminergic system. Orthodenticle homeobox 2 (OTX2) is a transcription factor important for the development of dopaminergic neurons residing in the ventral tegmental area (VTA), a critical region of the brain involved in drug reinforcement. Previous studies have demonstrated that ethanol exposure during embryonic development reduces Otx2 mRNA levels in the central nervous system. We hypothesized that levels of OTX2 would be altered by binge-like ethanol consumption in adult animals. To test this, Otx2 mRNA and protein levels in the mouse VTA were measured by quantitative real-time PCR and western blotting, respectively, after mice drank ethanol for 4 days in a procedure that elicits binge levels of ethanol consumption (drinking in the dark). Expression of known and putative OTX2 transcriptional target genes (Sema3c, Wnt1, and Mdk) were also measured in the VTA after ethanol drinking. Otx2 mRNA and protein levels were elevated in the VTA 24 hours after the fourth drinking session and there was a corresponding increase in the expression of Mdk transcript. Interestingly, Wnt1 transcript was elevated in the VTA immediately after the fourth drinking session but returned to control levels 24 hours later. We next investigated if viral-mediated reduction of Otx2 in the mouse VTA would alter ethanol or sucrose intake. Lentiviral vectors expressing a shRNA targeting Otx2 or a control shRNA were injected into the VTA and mice were tested in the drinking in the dark protocol for ethanol and sucrose drinking. Reducing levels of OTX2 in the VTA did not alter ethanol or sucrose consumption. One limitation is that the extent of OTX2 reduction may not have been sufficient. Although OTX2 in the VTA may not play a role in binge-like drinking in adult mice, OTX2 could contribute to ethanol-induced transcriptional changes in this region.

Effects of Paternal Preconception Vapor Alcohol Exposure Paradigms on Behavioral Responses in Offspring.

We and others previously reported that paternal preconception chronic ethanol exposure leads to molecular, physiological, and behavioral changes in offspring including reduced ethanol consumption and preference relative to controls. The goal of the present study was to further explore the impact of paternal ethanol exposure on a wide variety of basal and drug-induced behavioral responses in first generation offspring. Adult male mice were exposed to chronic intermittent vapor ethanol or control conditions for 5–6 weeks before being mated with ethanol-naïve females to produce ethanol (E)- and control (C)-sired offspring. E-sired male offspring showed stress hyporesponsivity in a stress-induced hyperthermia assay and E-sired female offspring had reduced binge-like ethanol consumption in a drinking in the dark assay compared to C-sired offspring. E-sired offspring also showed altered sensitivity to a sedative/hypnotic dose of the GABAergic drug midazolam, but not ketamine or ethanol, in a loss of the righting response assay. E-sired offspring did not differ from controls in marble burying, novel object location, novel object recognition, social interaction, bottle-brush, novelty suppressed feeding, prepulse inhibition, every-other-day ethanol drinking, or home cage activity assays. This study adds to a growing body of literature suggesting that like in utero alcohol exposure, paternal preconception alcohol exposure can also have effects that persist and impact behavior of offspring.

Antibiotic-induced disruption of commensal microbiome linked to increases in binge-like ethanol consumption behavior

Research focusing on the gut-brain axis is growing, but the interplay of ethanol (alcohol molecule), the gut microbiome, the brain and behavior is poorly understood. In the current study, we remodeled the gut microbiota by providing adult male C57BL/6J mice with a non-absorbable antibiotic cocktail (ABX) in the drinking water and tested ethanol consumption behavior in a binge-like “Drinking in the Dark” model. Notably, 2 weeks of ABX pre-treatment significantly increased ethanol consumption during the 6 weeks of ethanol exposure in the DID paradigm. ABX treatment also appeared to prevent anxiety-like behavior during ethanol withdrawal period. ABX-treated mice expressed reduced bacterial diversity and modified microbiota compositions within cecal samples. There were drastically reduced levels of commensal Firmicutes and increases in the Bacteroidetes and Verrucomicrobia populations. Importantly, the relative abundance of Firmicutes inversely correlated to ethanol intake levels regardless of antibiotic treatment, whereas Bacteroidetes and Verrucomicrobia populations negatively correlated to ethanol intake levels. This is the first report demonstrating that ABX-induced disruption of the gut commensal microbiota leads to increased ethanol consumption in mice. This work reveals an important relationship between the gut microbiota and ethanol consumption behavior and supports the use of microbial-targeted approaches to study gut-brain interactions during alcohol use disorder.

Prolonged caffeine intake decreases alveolar bone damage induced by binge-like ethanol consumption in adolescent female rats

Ethanol consumption has been reported to negatively impact on periodontal disease. In particular, oral cavity disorders occur upon ethanol exposure during adolescence, a life period associated with particular patterns of short and intense (‘binge-like’) ethanol consumption that is most deleterious to oral health. The hazardous central effects of ethanol have been linked to the overfunction of adenosine receptors, which are antagonized by caffeine, a bioactive substance present in numerous natural nutrients, which can also modify bone metabolism. The aim of this study was to investigate the effects of caffeine on alveolar bone damage induced by an ethanol binge drinking paradigm during adolescence. Female Wistar rats (35 days old; n = 30) were allocated to six groups: control (vehicle), ethanol (3 g/kg/day; 3 days On-4 days Off challenge), caffeine (10 mg/kg/day), caffeine plus ethanol, SCH58261 (0.1 mg/kg/day, an antagonist of A2A receptors), and SCH58261 plus ethanol. Bone micromorphology and vertical bone loss were analyzed by computed microtomography. Our data showed that ethanol binge drinking reduced alveolar bone quality, with repercussion on alveolar bone size. This ethanol-induced alveolar bone deterioration was abrogated upon treatment with caffeine, but not with SCH58261. This shows that caffeine prevented the periodontal disorder caused by ethanol binge drinking during adolescence, an effect that was not mediated by adenosine A2A receptor blockade.

One-off liver directed AAV gene therapy achieves long term uptake of acid beta-glucocerebrosidase by macrophages of affected tissues in Gaucher disease

Binge alcohol drinking is a health burden in the United States, which has an alarming economic impact. Unfortunately, medications available for alcohol abuse have low efficacy or adverse side effects, creating a need to evaluate novel therapies. Growing research suggests that 18-Methoxycoronaridine (18-MC), an α3β4 nicotinic acetylcholine receptor (nAChR) antagonist, may be effective at reducing ethanol consumption. However, its effects on binge-like ethanol consumption and other ethanol behaviors have not been examined. The present study examined the effect of α3β4 nAChRs antagonism on basal locomotor activity in male and female C57BL/6J mice. Next we tested the effect of 18-MC on binge-like ethanol consumption, ethanol-induced sedation, and ethanol metabolism. Finally, we tested the effect of α3β4 nAChRs on saccharin consumption to ensure effects were specific for ethanol. We observed that 18-MC decreased binge-like ethanol consumption without altering saccharin consumption, the sedative effects of ethanol, or ethanol metabolism. High doses of 18-MC caused locomotor sedation in C57BL/6J mice, but the effects were brief and likely did not contribute to differences in ethanol consumption. Our results support the involvement of the α3β4 nAChRs in binge-like ethanol intake, and further work should explore the use of 18-MC for treatment of alcohol use disorders.

Corticotropin Releasing Factor Type 1 and 2 Receptor Signaling in the Medial Prefrontal Cortex Modulates Binge-Like Ethanol Consumption in C57BL/6J Mice.

Corticotropin releasing factor (CRF) signaling via limbic CRF1 and 2 receptors (CRF1R and CRF2R, respectively) is known to modulate binge-like ethanol consumption in rodents. Though CRF signaling in the medial prefrontal cortex (mPFC) has been shown to modulate anxiety-like behavior and ethanol seeking, its role in binge ethanol intake is unknown. Here, we used “drinking-in-the-dark” (DID) procedures in male and female C57BL/6J mice to address this gap in the literature. First, the role of CRF1R and CRF2R signaling in the mPFC on ethanol consumption was evaluated through site-directed pharmacology. Next, we evaluated if CRF1R antagonist reduction of binge-intake was modulated in part through CRF2R activation by co-administration of a CRF1R and CRF2R antagonist. Intra-mPFC inhibition of CRF1R and activation of CRF2R resulted in decreased binge-like ethanol intake. Further, the inhibitory effect of the CRF1R antagonist was attenuated by co-administration of a CRF2R antagonist. We provide novel evidence that (1) inhibition of CRF1R or activation of CRF2R in the mPFC reduces binge-like ethanol intake; and (2) the effect of CRF1R antagonism may be mediated via enhanced CRF2R activation. These observations provide the first direct behavioral pharmacological evidence that CRF receptor activity in the mPFC modulates binge-like ethanol consumption.

Emotional Reactivity to Incentive Downshift in Adult Rats Exposed to Binge-Like Ethanol Exposure During Adolescence.

Alcohol use in adolescents is often characterized by binge-like ethanol consumption pattern, which is associated with long-term health consequences and even with important harms to his developing brain. Among this, ethanol exposure induces long-lasting alterations in anxiety-related neurobiological systems such as corticotropin releasing factor (CRF) or melanocortin system (MC). Recently, it has been demonstrated that adult rats exposed to adolescent intermittent ethanol (AIE) exposure exhibited anxiogenic-like behavior. Given that it has been demonstrated that negative affective state is relevant to development of addictive behavior, it is tempting to suggest that increased risk of adult abusive alcohol use exhibited in rats exposed to ethanol during adolescence may be related with differences in anxiety-related behavior. We conducted a study investigating the emotional reactivity after a reward devaluation (12-to-1 pellet or 32-to-4% sucrose downshift) in adult rats exposed to binge-like ethanol exposure during adolescence. For this aim, adolescent Sprague-Dawley rats were treated with ethanol (2.5 g/kg ip; AIE) or saline (AIS) for 2 consecutive days at 48-h intervals over a 14-day period (PND30-PND43). Following 25 free-ethanol days, adult rats were trained in consummatory and instrumental successive negative contrast task (cSNC and iSNC). Our data shows that both AIE and AIS groups exhibited suppression of the consummatory and instrumental behavior after reward devaluation relative to unshifthed control. Also, adult rats exposed to alcohol during adolescence exhibited a particularly strong negative affective state (lower sucrose consumption) with regards to the AIS group in the cSNC. This data demonstrated that adolescent binge-like ethanol exposure might trigger a greater emotional reactivity following incentive downshift, which might be linked to higher vulnerability to substance use disorder.

Ketamine normalizes binge drinking-induced defects in glutamatergic synaptic transmission and ethanol drinking behavior in female but not male mice

Ketamine is a fast acting experimental antidepressant with significant therapeutic potential for emotional disorders such as major depressive disorder and alcohol use disorders. Of particular interest is binge alcohol use, which during intermittent withdrawal from drinking involves depressive-like symptoms reminiscent of major depressive disorder. Binge drinking has been successfully modeled in mice with the Drinking in the Dark (DID) paradigm, which involves daily access to 20% ethanol, for a limited duration and selectively during the dark phase of the circadian light cycle. Here we demonstrate that DID exposure reduces the cell surface expression of NMDA- and AMPA-type glutamate receptors in the prelimbic cortex (PLC) of female but not male mice, along with reduced activity of the mammalian target of rapamycin (mTOR) signaling pathway. Pretreatment with an acute subanesthetic dose of ketamine suppresses binge-like ethanol consumption in female but not male mice. Lastly, DID-exposure reduces spontaneous glutamatergic synaptic transmission in the PLC of both sexes, but synaptic transmission is rescued by ketamine selectively in female mice. Thus, ketamine may have therapeutic potential as an ethanol binge suppressing agent selectively in female subjects.

Characterization of the Hippocampal Neuroimmune Response to Binge-Like Ethanol Consumption in the Drinking in the Dark Model

Objectives: Alcohol dependence leads to dysregulation of the neuroimmune system, but the effects of excessive alcohol consumption on key players of the neuroimmune response after episodic binge drinking in nondependence has not been readily assessed. These studies seek to determine how the neuroimmune system within the hippocampus responds to binge-like consumption prior to dependence or evidence of brain damage. Methods: C57BL/6J mice underwent the drinking in the dark (DID) paradigm to recapitulate binge consumption. Immunohistochemical techniques were employed to determine the effects of ethanol on cytokine and astrocyte responses within the hippocampus. Astrocyte activation was also assessed using qRT-PCR. Results: Our results indicated that binge-like ethanol consumption resulted in a 3.6-fold increase in the proinflammatory cytokine interleukin (IL)-1β immunoreactivity in various regions of the hippocampus. The opposite effect was seen in the anti-inflammatory cytokine IL-10. Binge-like consumption resulted in a 67% decrease in IL-10 immunoreactivity but had no effect on IL-4 or IL-6 compared with the water-drinking control group. Moreover, astrocyte activation occurred following ethanol exposure as GFAP immunoreactivity was increased over 120% in mice that experienced 3 cycles of ethanol binges. PCR analyses indicated that the mRNA increased by almost 4-fold after one cycle of DID, but this effect did not persist in abstinence. Conclusions: Altogether, these findings suggest that binge-like ethanol drinking prior to dependence causes dysregulation to the neuroimmune system. This altered neuroimmune state may have an impact on behavior but could also result in a heightened neuroimmune response that is exacerbated from further ethanol exposure or other immune-modulating events.