Intermittent Access Research Articles

Distinct Chrna5 mutations link excessive alcohol use to types I/II vulnerability profiles and IPN GABAergic neurons.

Genome wide association and animal studies have implicated genetic variations in CHRNΑ5, encoding the α5 subunit-containing nicotinic acetylcholine receptors (α5*nAChRs), as a risk factor for developing alcohol use disorders (AUDs). To understand how α5*nAChR mutations may influence alcohol (EtOH) drinking behavior, we used a two-bottle choice procedure with intermittent access to alcohol in male and female transgenic mice expressing either the highly frequent human single nucleotide polymorphism (α5SNP/rs16969968) or a deletion of the Chrna5 gene (α5KO). AUDs-related preconsommatory traits (anxiety, sensation-seeking and impulsivity) were assessed with a battery of relevant tasks (elevated-plus maze, novel place preference and step-down inhibitory avoidance). The implication of the α5-expressing IPN GABAergic neurons in AUDs and related behavioral traits was verified using neurospecific lentiviral (LV)-induced reexpression of the α5 subunit in α5KOxGAD-Cre mice. Both α5SNP and α5KO mice showed over-consumption of EtOH, but displayed opposite vulnerability profiles consistent with Cloninger's subtypes of human AUDs. α5SNP mice showed Type I-like characteristics, i.e., high anxiety, novelty avoidance, whereas α5KOs exhibited Type II-like features such as low anxiety and high impulsivity. LV re-expression of the α5 subunit in IPN GABAergic neurons restored the control of EtOH intake and improved the impulsive phenotype. We demonstrate that the SNP (rs16969968) or null mutation of Chrna5 result in increased volitional EtOH consumption but opposite effects on anxiety, novelty-seeking and impulsive-like behaviors that match Cloninger type I and II of AUDs, including sex-related variations. IPN GABAergic neurons expressing α5*nAChRs play a key role in limiting both EtOH drinking and motor impulsivity.

Behavioral changes and transcriptional regulation of mesolimbic dopaminergic genes in a mouse model of binge eating disorder by diet intermittent access.

Binge Eating Disorder (BED) is among the most prevalent eating disorders worldwide. It is characterized by recurrent episodes of excessive consumption of palatable foods in short periods, accompanied by a sense of loss of control and distress around the episode, which tends to worsen over time. The mesolimbic dopaminergic system influences on reinforcement and reward-seeking behaviors is implicated in the disorder's pathogenesis. Animal models that replicate the clinical conditions observed in humans, including the disorder progression, are essential for understanding the underlying physiological mechanisms of BED. This study aimed to evaluate binge eating behavior induced by intermittent High Sugar and Butter (HSB) diet access in mice, their phenotypes, transcriptional regulation of mesolimbic dopaminergic system genes, and behavior. Thus, mice were subdivided into three groups: CHOW (maintenance diet only), HSB-i (maintenance diet with thrice-weekly access to HSB), and HSB (continuous access to HSB). Animals were subjected to marble-burying and light-dark box behavioral tests, and transcriptional regulation was evaluated by RT-qPCR. The results indicated that the HSB-i group established a feeding pattern of significantly more kilocalories on days when HSB was available and reduced intake on non-HSB days similar to human binge eating. Over time, binge episodes intensified, potentially indicating a tolerance effect. Additionally, these animals behave differently towards preferring the HSB diet and exhibited altered transcriptional regulation of the Drd1, Slc6a3, and Lrrk2 genes. Our study provides a mouse model that reflects human BED, showing a progression in binge episodes and mesolimbic dopamine pathway involvement, suggesting targets for future therapeutic interventions.

Ethanol drinking sex-dependently alters cortical IL-1β synaptic signaling and cognitive behavior in mice.

Individuals with alcohol use disorder (AUD) struggle with inhibitory control, decision making, and emotional processing. These cognitive symptoms reduce treatment adherence, worsen clinical outcomes, and promote relapse. Neuroimmune activation is a key factor in the pathophysiology of AUD, and targeting this modulatory system is less likely to produce unwanted side effects compared to directly targeting neurotransmitter dysfunction. Notably, the cytokine interleukin-1β (IL-1β) has been broadly associated with the cognitive symptoms of AUD, though the underlying mechanisms are not well understood. Here we investigated how chronic intermittent 24-hour access two bottle choice ethanol drinking affects medial prefrontal cortex (mPFC)-related cognitive function and IL-1 synaptic signaling in male and female C57BL/6J mice. In both sexes, ethanol drinking decreased reference memory and increased mPFC IL-1 receptor 1 (IL-1R1) mRNA levels. In neurons, IL-1β can activate either pro-inflammatory or neuroprotective intracellular pathways depending on the isoform of the accessory protein (IL-1RAcP) recruited to the IL-1R1 complex. Moreover, ethanol drinking sex-dependently shifted mPFC IL-1RAcP isoform gene expression and IL-1β regulation of mPFC GABA synapses, both of which may contribute to female mPFC resiliency and male mPFC susceptibility. This type of signaling bias has become a recent focus of rational drug development. Therefore, in addition to increasing our understanding of how IL-1β sex-dependently contributes to mPFC dysfunction in AUD, our current findings also support the development of a new class of pharmacotherapeutics based on biased IL-1 signaling.

Binge ethanol consumption can be attenuated by systemic administration of minocycline and is associated with enhanced neuroinflammation in the central amygdala

Alcohol use disorder (AUD) has a complicated pathophysiology. Binge ethanol intoxication may produce long-lasting changes throughout extended amygdala neurocircuitry including neuroinflammation, often leading to relapse. Therefore, understanding the role of binge drinking induced neuroinflammation on extended amygdala neurocircuitry is critically important for treatment. We sought to understand the role of neuroinflammation in a naturalized form of rodent binge ethanol drinking (Drinking in the Dark (DID)). In a 5-week DID paradigm, we demonstrate that acute intraperitoneal (IP) injection of the anti-inflammatory drug minocycline significantly reduced binge drinking repeatedly in male and female Cx3CR1-GFP and C57BL/6J mice. Importantly, IP administration transiently decreased intermittent access sucrose consumption, was not observed on the second IP injection, but did not significantly alter food or water consumption, suggesting that minocycline may produce initial acute aversive effects and may not alter long-term consumption of natural rewards. Examination of rodent behaviors post ethanol binge drinking reveals no lasting effects of minocycline treatment on locomotion or anxiety-like behavior. To assess neuroinflammation, we developed a novel analysis method using a Matlab image analysis script, which allows for non-biased skeletonization and evaluation of microglia morphology to determine a possible activation state in Cx3CR1-GFP knock-in mice after repeated DID. We observed significant morphological changes of microglia within the CeA, but no differences in the BLA. Taken together, this study demonstrates repeated binge ethanol consumption can produce significant levels of microglia morphology changes within the CeA, and that immunomodulatory therapies may be an intriguing pharmacological candidate for the treatment of AUD.

Expression of sensitized β2 nAChR subunits in VTA neurons enhances intravenous nicotine self-administration in male rats

Ventral tegmental area (VTA) nicotinic acetylcholine receptors (nAChRs) are important for nicotine reinforcement. To determine whether and to what extent these receptors are sufficient for nicotine reinforcement, we expressed β2Leu9′Ser (i.e. sensitized) nAChR subunits in the VTA of adult male rats and assessed the nicotine dose-response relationship in intravenous self-administration (SA). β2Leu9′Ser rats self-administered nicotine doses 50–100 fold lower than the lowest doses that control rats would respond for. Expression of WT β2 subunits confirmed that this enhanced sensitivity to nicotine was due to the Leu9′Ser mutation in β2. Higher unit doses were associated with strong escalation in β2Leu9′Ser rats over 17 fixed ratio sessions. Escalation was minimal or absent in control rats at the same unit doses. In progressive ratio SA, β2Leu9′Ser rats exhibited higher breakpoints than control rats when the nicotine unit dose was 1.5 μg/kg/inf or higher. In intermittent access SA, β2Leu9′Ser rats exhibited response patterns very similar to control rats. By adding nicotine dose-response data, progressive ratio assays, and intermittent access results that rule out stereotypy, these data significantly extend our previous finding that nicotine activation of the mesolimbic dopamine pathway is sufficient for nicotine reinforcement.

Intermittent access to sugary drinks associated with fasting induces overeating and depressive-like behavior in female C57BL/6J mice

Binge eating disorder is the most prevalent eating disorder, affecting both sexes but more commonly found in women. Given the frequent co-occurrence of psychiatric disorders, this study aimed to establish a standardized experimental intermittent protocol to investigate overeating associated with depression. A 10-day protocol induced uncontrolled eating behavior in C57BL/6J female mice. The first experiment included the following groups: naive group (chow ad libitum), control group (chow and sucrose solution ad libitum), and fasting groups (16 and 20 h) exposed to an intermittent sucrose solution (10 %) and chow regimen. Subsequently, the feeding test, open field test, elevated plus maze test, tail suspension test, and light/dark conflict test were conducted. Furthermore, monoamine oxidase (MAO) A and B activities in brain structures and plasma corticosterone levels were assessed. Food overconsumption and depressive-like behavior were observed in both sucrose fasting groups, while risk-taking behaviors were specifically observed in the 20-hour fasting sucrose group. While both fasting sucrose groups caused reduced hippocampal MAO-A activity, only the F20 sucrose group inhibited MAO-B in the cortex and hypothalamus. Moreover, both fasting sucrose groups exhibited elevated corticosterone levels. In a separate design (Experiment 2), groups with 16 and 20 h of fasting alone (without sucrose) did not show the same behavioral results as the intermittent fasting sucrose groups, thus avoiding fasting bias. Based on these results, the 20-hour sucrose fasting group was chosen as the ideal protocol for mimicking overeating behavior associated with depression to investigate future therapeutic approaches for this comorbidity.

Effects of early life stress paired with adolescent alcohol consumption on two-bottle choice alcohol drinking behaviors in mice.

In humans, early life stress (ELS) is associated with an increased risk for developing both alcohol use disorder (AUD) and post-traumatic stress disorder (PTSD). We previously used an infant footshock model that produces stress-enhanced fear learning (SEFL) in rats and mice and increases aversion-resistant alcohol drinking in rats to explore this shared predisposition. The goal of the current study was to extend this model of comorbid PTSD and AUD to male and female C57BL/6J mice. Acute ELS was induced using 15 footshocks on postnatal day (PND) 17. In adulthood, alcohol drinking behavior was tested in one of three two-bottle choice drinking paradigms. In continuous access, mice were given 24 h access to 5% or 10% ethanol and water for five consecutive drinking sessions each. In limited access drinking in the dark, mice were given 2 h of access to 15% ethanol and water across 15 sessions 3 h into the dark cycle. In intermittent access, mice were presented with 20% ethanol and water Monday, Wednesday, and Friday, for four consecutive weeks. In a fifth week of intermittent access drinking, increasing concentrations of quinine (10 mg/L, 100 mg/L, and 200 mg/L) were added to the ethanol to test aversion-resistant drinking. Intermittent access drinking was tested with and without a period of adolescent drinking (PND 35). Infant footshock did not alter drinking in the continuous or limited access tasks. Adult consumption and preference were lower in the intermittent access task when adolescent drinking was included and there were ELS-induced differences in consumption of quinine-adulterated ethanol in females. Our results demonstrate that infant footshock followed by a period of adolescent drinking is a viable model of comorbid PTSD and AUD in rats and mice.

Unveiling the power of optimism: Exploring behavioral and neuromolecular correlates of alcohol seeking and drinking in rats with biased judgement

Alcohol use disorder (AUD) is a common psychiatric condition with substantial global mortality. Despite extensive research into its pathophysiology, the cognitive predispositions driving alcohol dependence are less understood. This study explores whether biased cognition, specifically traits of optimism and pessimism, predicts susceptibility to alcohol-seeking behaviors using an animal model.Rats were initially tested for judgement bias through Ambiguous Cue Interpretation tests. Those identified as ‘optimistic' or ‘pessimistic' were further examined for their tendency to escalate alcohol intake using the intermittent access 2-bottle choice (2BC) paradigm. Additionally, we assessed how judgement bias influenced the development of compulsive alcohol-seeking behavior in a Seeking-Taking (ST) and Seeking-Taking Punishment tasks, alcohol-seeking motivation in the Progressive Ratio Schedule of Reinforcement paradigm, the speed of extinction, and reinstatement after abstinence. Neurochemical analyses were conducted to investigate trait-specific differences in neurotransmitter-related gene expression and receptor densities in the brain. We used TaqMan Gene Expression Array Cards to analyze expression levels of genes linked to serotonergic, dopaminergic, glutamatergic, and GABAergic pathways, and alcohol metabolism in various brain regions. Receptor densities for 5-HT1A, 5-HT2A, and D2 were measured using autoradiography analysis.Behaviorally, ‘optimistic' rats showed significantly lower alcohol consumption in the 2BC paradigm compared to ‘pessimistic' rats. This lowered intake correlated with decreased monoamine oxidase-A (Maoa) expression in the medial prefrontal cortex (mPFC) and increased metabotropic glutamate receptor 2 (Grm2) expression in the amygdala (Amy). Additionally, we observed significant interactions between judgement bias and alcohol intake in the expression of several genes in the mPFC, nucleus accumbens (Nacc), orbitofrontal cortex (OFC), and Amy, as well as in 5-HT2A receptor binding in the Nacc.Overall, these results suggest that optimism is linked to lower alcohol consumption and related neurochemical changes, indicating a potential cognitive mechanism in AUD risk.

Effects of voluntary chronic intermittent access to ethanol on the behavioral performance in adult C57BL/6 J mice

BackgroundChronic alcohol drinking increases the risk of alcohol use disorders, causing various neurological disorders. However, the impact of different ethanol levels on a spectrum of behaviors during chronic drinking remains unclear. In this study, we established an intermittent access to ethanol in a two-bottle choice (IA2BC) procedure to explore the dose-dependent effects of ethanol on the behavioral performance of C57BL/6 J mice. MethodsAdult male C57BL/6 J mice were provided voluntary access to different ethanol concentrations (0 %, 5 %, 10 %, and 20 % ethanol) under a 12-week IA2BC paradigm. A battery of behavioral tests was administered to assess alterations in pain threshold, anxiety-like behaviors, locomotor activity, motor coordination, and cognition. Ethanol consumption and preference were monitored during each session. Moreover, the liver, heart, and lung tissues were examined using pathological microscopy. ResultsThe average (standard deviation) ethanol consumption of mice under the IA2BC paradigm increased dose-dependently to 5.1 (0.2), 8.7 (0.7), and 15.9 (0.8) g/kg/24 h with 5 %, 10 %, and 20 % ethanol, respectively. However, there is no significant difference in ethanol preference among all the ethanol groups. Chronic ethanol drinking caused hyperalgesia, cognitive impairment, and motor incoordination, but caused no changes in body temperature, locomotor activity, or anxiety-like behaviors. Minor histopathological alterations in the liver were detected; however, no major abnormal pathology was observed in the heart or lungs. ConclusionThese findings clarify the link between ethanol dosage and behavioral changes in mice over a 12-week IA2BC paradigm, thereby bridging the knowledge gap regarding the effects of chronic ethanol drinking on neurological disorders.

Intermittent Electrification with Battery Locomotives and the Post-Diesel Future of North American Freight Railroads

Changing attitudes, regulations, public policy, and international treaties concerning fossil fuels are likely to lead freight railroads toward carbon-neutral technologies, yet only electric traction matches the performance of diesel-electric locomotives. The tremendous power requirements of freight trains make efforts to reduce greenhouse gas (GHG) emissions challenging, but the prospect of 7.2 megawatt-hour battery-electric locomotives (BELs) offers promise, and intermittent electrification can facilitate battery charging. Train performance calculations under simulated real-world conditions show four 7.2 megawatt-hour BELs can power 8,000-ton trains up to 230 mainline miles unassisted, with average energy consumption of 12.5 watt-hours/ton-mile. This permits discontinuous electrification of major freight lines, leaving “gaps” of up to 200 mi to reduce capital costs, especially in rugged terrain or where the power grid is sparse. Massive onboard battery arrays and intermittent access to the electrical grid for traction power and recharging provide great energy savings by recycling energy now lost when traveling downhill or braking. A case study of a hypothetical Class I railroad found intermittent electrification with BELs more than 60% more cost-effective than contiguous electric districts, and dramatically reduced engine changes. To reduce railroad GHG emissions, governments must support technical development, show that electrification works in various North American settings, and develop institutional-financial frameworks to incentivize intermittent electrification with BELs, all in the context of massive environmental and capacity upgrades to electrical networks. Given proper assistance and incentives, early 21st century railroads may find discontinuous overhead-wire electrification offers great promise in operating terms.

Experimental error suppression in Cross-Resonance gates via multi-derivative pulse shaping

While quantum circuits are reaching impressive widths in the hundreds of qubits, their depths have not been able to keep pace. In particular, cloud computing gates on multi-qubit, fixed-frequency superconducting chips continue to hover around the 1% error range, contrasting with the progress seen on carefully designed two-qubit chips, where error rates have been pushed towards 0.1%. Despite the strong impetus and a plethora of research, experimental demonstration of error suppression on these multi-qubit devices remains challenging, primarily due to the wide distribution of qubit parameters and the demanding calibration process required for advanced control methods. Here, we achieve this goal, using a simple control method based on multi-derivative, multi-constraint pulse shaping, which acts simultaneously against multiple error sources. Our approach establishes a two to fourfold improvement on the default calibration scheme, demonstrated on four qubits on the IBM Quantum Platform with limited and intermittent access, enabling these large-scale fixed-frequency systems to fully take advantage of their superior coherence times. The achieved CNOT fidelities of 99.7(1)% on those publically available qubits come from both coherent control error suppression and accelerated gate time.

Identification of novel genetic loci and candidate genes for progressive ethanol consumption in diversity outbred mice.

Mouse behavioral genetic mapping studies can identify genomic intervals modulating complex traits under well-controlled environmental conditions and have been used to study ethanol behaviors to aid in understanding genetic risk and the neurobiology of alcohol use disorder (AUD). However, historically such studies have produced large confidence intervals, thus complicating identification of potential causal candidate genes. Diversity Outbred (DO) mice offer the ability to perform high-resolution quantitative trait loci (QTL) mapping on a very genetically diverse background, thus facilitating identification of candidate genes. Here, we studied a population of 636 male DO mice with four weeks of intermittent ethanol access via a three-bottle choice procedure, producing a progressive ethanol consumption phenotype. QTL analysis identified 3 significant (Chrs 3, 4, and 12) and 13 suggestive loci for ethanol-drinking behaviors with narrow confidence intervals (1-4 Mbp for significant QTLs). Results suggested that genetic influences on initial versus progressive ethanol consumption were localized to different genomic intervals. A defined set of positional candidate genes were prioritized using haplotype analysis, identified coding polymorphisms, prefrontal cortex transcriptomics data, human GWAS data and prior rodent gene set data for ethanol or other misused substances. These candidates included Car8, the lone gene with a significant cis-eQTL within a Chr 4 QTL for week four ethanol consumption. These results represent the highest-resolution genetic mapping of ethanol consumption behaviors in mice to date, providing identification of novel loci and candidate genes for study in relation to the neurobiology of AUD.

Greater inhibition of female rat binge alcohol intake by adrenergic receptor blockers using a novel Two-Shot rat binge drinking model

Binge drinking (BD) contributes strongly to the harms of alcohol use disorder. Most rodent models do not result in binge-level blood alcohol concentrations (BACs), and to better understand individual and sex differences in neurobiological mechanisms related to BD, the use of outbred rat strains would be valuable. Here, we developed a novel BD model where after 3+ months of intermittent access to 20% alcohol Wistar rats drank, twice a week, with two 5-min intake (what we called Two-shot) separated by a 10-min break. Our findings showed during Two-Shot that most animals reached ≥ 80 mg% BAC levels (when briefly food-restricted). However, when increasing alcohol concentrations from 20 to 30%, 40%, or 50%, rats titrated to similar intake levels, suggesting rapid sensing of alcohol effects even when front-loading. Two-Shot drinking was reduced in both sexes by naltrexone (1 mg/kg), validating intake suppression by a clinical therapeutic agent for human problem drinking. Further, both propranolol (β-adrenergic receptor antagonist) and prazosin (α1-adrenergic receptor antagonist) reduced female but not male BD at the lower dose. Thus, our results provide a novel model for BD in outbred rats and suggest that female binging is more sensitive to adrenergic modulation than males, perhaps providing a novel sex-related therapy.

The Small G-Protein Rac1 in the Dorsomedial Striatum Promotes Alcohol-Dependent Structural Plasticity and Goal-Directed Learning in Mice.

The small G-protein Ras-related C3 botulinum toxin substrate 1 (Rac1) promotes the formation of filamentous actin (F-actin). Actin is a major component of dendritic spines, and we previously found that alcohol alters actin composition and dendritic spine structure in the nucleus accumbens (NAc) and the dorsomedial striatum (DMS). To examine if Rac1 contributes to these alcohol-mediated adaptations, we measured the level of GTP-bound active Rac1 in the striatum of mice following 7 weeks of intermittent access to 20% alcohol. We found that chronic alcohol intake activates Rac1 in the DMS of male mice. In contrast, Rac1 is not activated by alcohol in the NAc and DLS of male mice or in the DMS of female mice. Similarly, closely related small G-proteins are not activated by alcohol in the DMS, and Rac1 activity is not increased in the DMS by moderate alcohol or natural reward. To determine the consequences of alcohol-dependent Rac1 activation in the DMS of male mice, we inhibited endogenous Rac1 by infecting the DMS of mice with an adeno-associated virus (AAV) expressing a dominant negative form of the small G-protein (Rac1-DN). We found that overexpression of AAV-Rac1-DN in the DMS inhibits alcohol-mediated Rac1 signaling and attenuates alcohol-mediated F-actin polymerization, which corresponded with a decrease in dendritic arborization and spine maturation. Finally, we provide evidence to suggest that Rac1 in the DMS plays a role in alcohol-associated goal-directed learning. Together, our data suggest that Rac1 in the DMS plays an important role in alcohol-dependent structural plasticity and aberrant learning.

Mu-opioid receptor knockout on Foxp2-expressing neurons reduces aversion-resistant alcohol drinking.

Mu-opioid receptors (MORs) in the amygdala and striatum are important in addictive and rewarding behaviors. The transcription factor Foxp2 is a genetic marker of intercalated (ITC) cells in the amygdala and a subset of striatal medium spiny neurons (MSNs), both of which express MORs in wild-type mice and are neuronal subpopulations of potential relevance to alcohol-drinking behaviors. For the current series of studies, we characterized the behavior of mice with genetic deletion of the MOR gene Oprm1 in Foxp2-expressing neurons (Foxp2-Cre/Oprm1fl/fl). Male and female Foxp2-Cre/Oprm1fl/fl mice were generated and heterozygous Cre+ (knockout) and homozygous Cre- (control) animals were tested for aversion-resistant alcohol consumption using an intermittent access (IA) task, operant responding for a sucrose reward, conditioned place aversion (CPA) to morphine withdrawal, and locomotor sensitization to morphine. The results demonstrate that deletion of MOR on Foxp2-expressing neurons renders mice more sensitive to quinine-adulterated ethanol (EtOH). Mice with the deletion (vs. Cre- controls) also consumed less alcohol during the final sessions of the IA task, responded less for sucrose under an FR3 schedule, and were less active at baseline and following morphine injection. Foxp2-MOR deletion did not impair the ability to learn to respond for reward or develop a conditioned aversion to morphine withdrawal. Together, these investigations demonstrate that Foxp2-expressing neurons may be involved in escalation of alcohol consumption and the development of compulsive-like alcohol drinking.

Escalating caffeine dose-dependently increases alcohol consumption in adult male, but not female, C57BL/6J mice

Although previous research has illustrated the effects of the consumption of alcohol and caffeine individually, less research has focused on the popular combination of the two drugs. The increase in alcohol consumption when combined with caffeine has led to the idea that the stimulant effects of caffeine may mask the depressant effects of alcohol, and this may contribute to increased binge drinking as the individual feels more awake and stimulated. Preclinical research has shown various effects of combined alcohol and caffeine where several studies show decreased alcohol consumption and others show increased alcohol consumption and even binge-like drinking. Results from a previous study in our lab indicate that intermittent access (IA) to steady levels of low (0.015 %) but not moderate (0.03 %) caffeine increased alcohol consumption in male C57BL/6J mice. The current studies further investigated the sex and dose differences in adult mice receiving varying concentrations of caffeine on combined alcohol intake. In Experiment 1, adult mice (n = 50, 25 males and 25 females) had IA to one of the following experimental bottles throughout the 4 week period: water, alcohol (10 % v/v), caffeine (0.015 % w/v), or 10 % alcohol +0.015 % caffeine. In Experiment 2, adult mice (n = 70, 35 males and 35 females) were given IA to one of the following experimental bottles: water, alcohol (10 % v/v; steady, maintained throughout the 4 weeks), caffeine (increasing 0.01 % to 0.015 % to 0.02 % to 0.03 % weekly), or 10 % alcohol+increasing caffeine (at the previously mentioned concentrations). When both caffeine and alcohol concentrations remained steady throughout the 4 weeks, there was no change in alcohol consumption. Chronic exposure to IA caffeine led to increased locomotor activity and decreased freezing episodes when tested in the open field test approximately 6 h after removal of the bottles. In Experiment 2, caffeine dose-dependently increased alcohol co-consumption in male mice whereas female mice consumed less alcohol when it was presented in conjunction with caffeine. The results in males are in line with clinical literature suggesting that the combination of alcohol and caffeine may lead to increased stimulation and alcohol drinking. Additionally, these studies provide evidence that the escalation of caffeine is crucial when investigating alcohol and caffeine co-consumption using the IA paradigm.

Assessing the Influence of Intermittent Alcohol Access on Acrylamide-Induced Neuronal Toxicity in an Experimental Rat Model.

Tobacco and alcohol have been identified as health risk behaviors associated with significant unfavorable health consequences, ranking within the list of the top ten causes of mortality and disability-adjusted life years (DALY). The combustion of tobacco leads to the formation of acrylamide (ACR), which is well known for its neurotoxic effects. Similarly, alcohol consumption has also been widely recognized for its neurotoxic effects. Both substances can affect neurons and neuroglia cells through various pathways. This study sought to examine the impacts of co-administration of ACR and intermittent-access ethanol (IAE) consumption over a period of one month. The experimental group received 20 mg/kg of ACR, administered orally, along with IAE of 20% ethanol sessions lasting 24 h, three times per week. The cognitive outcomes were assessed utilizing the elevated plus maze (EPM), which was employed as a means of assessing the capability to learn and remember, the novel object recognition (NOR) test, which was employed to assess recognition memory, and the Y-maze, which was used to explore a new environment and navigate. Additionally, ELISA assays were performed to examine underlying mechanisms, including markers associated with inflammation (NF-κB, PGE2, and TNF-α), apoptosis (Bcl2, Bax, and Caspase-3), and oxidative stress (MDA, catalase, and GSH). These markers were assessed in the brain homogenate as part of the investigation. Furthermore, a histopathological study was conducted. The findings indicated that NF-κB levels increased significantly in the combination of ACR and IAE groups (ACR + IAE) compared to either the ACR-alone or IAE-alone groups. However, parallel changes were observed in TNF-α, PGE2, Bax, Bcl-2, Caspase-3, GSH, and CAT levels when comparing the ACR + IAE group to the ACR-alone group. Comparable alterations were noted between the ACR + IAE treatment and IAE-alone groups in TNF-α, Bcl-2, MDA, GSH, and CAT levels. Moreover, the histopathological analysis revealed significant changes between the ACR + IAE and the ACR- or IAE-alone groups. Regarding memory parameters assessed using tests including EPM, NOR, and Y-maze, considerable changes were observed across all treatment groups as opposed to the control. Surprisingly, there were no notable differences in the NOR and Y-maze tasks between the alone and combination treatment. Further study is necessary to explore the long-term alteration of co-administering ACR and IAE on behavior, memory, and neurotoxicity-related mechanisms, in order to elucidate their combined effects more clearly.

Greater inhibition of female rat binge alcohol intake by adrenergic receptor blockers using a novel Two-Shot rat binge drinking model.

Binge drinking (BD) contributes strongly to the harms of alcohol use disorder. Most rodent models do not result in binge-level blood alcohol concentrations (BACs), and to better understand individual and sex differences in neurobiological mechanisms related to BD, the use of outbred rat strains would be valuable. Here, we developed a novel BD model where after 3+ months of intermittent access to 20% alcohol Wistar rats drank, twice a week, with two 5-minute intake (what we called Two-shot) separated by a 10-minute break. Our findings showed during Two-Shot that most animals reached ≥ 80mg% BAC levels (when briefly food-restricted). However, when increasing alcohol concentrations from 20% to 30%, 40%, or 50%, rats titrated to similar intake levels, suggesting rapid sensing of alcohol effects even when front-loading. Two-Shot drinking was reduced in both sexes by naltrexone (1mg/kg), validating intake suppression by a clinical therapeutic agent. Further, both propranolol (β adrenergic receptor antagonist) and prazosin (α1 adrenergic receptor antagonist) reduced female but not male BD at the lower dose. Thus, our results provide a novel model for BD in outbred rats and suggest that female binging is more sensitive to adrenergic modulation than males, perhaps providing a novel sex-related therapy.

Parvalbumin interneuron mGlu5 receptors govern sex differences in prefrontal cortex physiology and binge drinking.

Despite established sex differences in the prevalence and presentation of psychiatric disorders, little is known about the cellular and synaptic mechanisms that guide these differences under basal conditions. The proper function of the prefrontal cortex (PFC) is essential for the top-down regulation of motivated behaviors. The activity of the PFC is tightly controlled by parvalbumin-expressing interneurons (PV-INs), a key subpopulation of fast-spiking GABAergic cells that regulate cortical excitability through direct innervations onto the perisomatic regions of nearby pyramidal cells. Recent rodent studies have identified notable sex differences in PV-IN activity and adaptations to experiences such as binge drinking. Here, we investigated the cellular and molecular mechanisms that underlie sex-specific regulation of PFC PV-IN function. Using whole-cell patch-clamp electrophysiology and selective pharmacology, we report that PV-INs from female mice are more excitable than those from males. Moreover, we find that mGlu1 and mGlu5 metabotropic glutamate receptors regulate cell excitability, excitatory drive, and endocannabinoid signaling at PFC PV-INs in a sex-dependent manner. Genetic deletion of mGlu5 receptors from PV-expressing cells abrogates all sex differences observed in PV-IN membrane and synaptic physiology. Lastly, we report that female, but not male, PV-mGlu5-/- mice exhibit decreased voluntary drinking on an intermittent access schedule, which could be related to changes in ethanol's stimulant properties. Importantly, these studies identify mGlu1 and mGlu5 receptors as candidate signaling molecules involved in sex differences in PV-IN activity and behaviors relevant to alcohol use.

Relapse after intermittent access to cocaine: Discriminative cues more effectively trigger drug seeking than do conditioned cues.

When people with drug addiction encounter cues associated with drug use, this can trigger cravings and relapse. These cues can include conditioned stimuli (CSs) signaling drug delivery and discriminative stimuli (DSs) signaling drug availability. Compared to CS effects, DS effects are less explored in preclinical studies on cue-induced relapse. We compared CS and DS effects on reward seeking following abstinence from intermittent-access cocaine (or sucrose) self-administration. During 15-20 intermittent-access sessions, rats self-administered i.v. cocaine or sucrose pellets paired with a light-tone CS. Cocaine/sucrose was available for 5-min (signalled by a light; DS+) and unavailable for 25min (signalled by different lighting conditions; DS-), and this cycled for 4h/session. Following abstinence, we measured cocaine/sucrose seeking under extinction triggered by CS and DS presentation, and instrumental responding reinforced by these cues. Across intermittent-access sessions, rats increased lever pressing for cocaine or sucrose during DS+ periods and decreased responding during DS- periods. On days 2 and 21 of abstinence, only presentation of the DS+ or DS+ and CS combined elicited increased cocaine/sucrose-seeking behaviour (i.e., increased active lever presses). Presenting the DS- alongside the DS+ suppressed the increased cocaine-seeking behaviour otherwise produced by the DS+ . Finally, on day 21 of abstinence, rats showed equivalent levels of lever pressing reinforced by the DS+ , CS and by the DS+ and CS combined, suggesting comparable conditioned reinforcing value. After intermittent self-administration, cocaine-associated DSs and CSs acquire similar conditioned reinforcing properties, but DSs more effectively trigger increases in drug seeking.