Acute Effects Of Ethanol Research Articles

Increased anxiolytic effect in aged female rats and increased motoric behavior in aged male rats to acute alcohol administration: Comparison to younger animals

The population of most countries in the world is increasing and understanding risk factors that can influence the health of the older population is critical. Older adults consume alcohol often in a risky, binge manner. Previous work has demonstrated that aged rats are more sensitive to many of the effects of acute ethanol. In the current project aged, adult, and adolescent female and male rats were tested on the elevated plus maze and open field following either a 1.0 g/kg alcohol injection or a saline injection. We report sex- and age-dependent effects whereas aged female rats, but not aged male rats, showed an increased anxiolytic effect of alcohol in the elevated plus maze while aged male rats, but not aged female rats, showed increased stimulatory movement in the open field. In addition, significant age effects were found for both female and male rats. It is proposed that the sex- and age-dependent effects reported in the current studies may be due to differential levels of alcohol-induced allopregnanolone for the anxiolytic effects and differential levels of alcohol-induced dopamine for the stimulatory effects. The current work provides insights into factors influencing alcohol consumption in older adults.

Ethanol-induced atrial fibrillation results from late INa and can be prevented by ranolazine

Abstract Background Ethanol consumption is the most important self-triggered cause of atrial fibrillation (AF). As abstinence often fails, alternative therapies are needed. CaMKII-activation upon ethanol with consecutive Ca2+-leak from the sarcoplasmic reticulum (SR) appears to be an important pathomechanism for ethanol-induced AF, but is not a druggable target. CaMKII has previously been shown to induce late sodium current (late INa) which could be proarrhythmogenic. Purpose We hypothesized that late INa might be involved in the development of AF upon acute ethanol exposition and that thus, the antianginal drug ranolazine (an inhibitor of late INa) could be repurposed to prevent ethanol-induced AF. Methods Acute effects of ethanol were investigated in vivo and in vitro as compared to vehicle. The occurrence of AF in vivo was assessed by programmed electrical stimulation in mice. To test the role of late INa, we used ranolazine in vivo and in vitro, as well as tetrodotoxin in vitro. We measured late INa and cytosolic sodium concentration, recorded stimulated action potentials and spontaneous delayed afterdepolarizations (DAD), and assessed SR Ca2+-leak by recording Ca2+-sparks. Mechanistically, we inhibited CaMKII using AIP and used NaV1.5 S571A mice to specifically investigate the role of CaMKII-mediated late INa. Human biopsies were acquired during surgery and were used for cell isolation or chunk incubation. Results Ethanol acutely induced late INa, sodium overload, SR Ca2+-leak and delayed afterdepolarizations in isolated atrial cardiomyocytes, as well as atrial fibrillation in vivo (10 of 10 mice). All of these cellular ethanol effects were prevented by pharmacologic inhibition of late INa. Importantly, ranolazine prevented the development of AF upon ethanol exposure in vivo. Mechanistically, CaMKII-activation and, accordingly, CaMKII-dependent NaV1.5 phosphorylation (S571) were increased by ethanol exposure. Indeed, the phosphoresistant NaV1.5 S571A mutation prevented late INa, SR Ca2+-leak and in vivo AF upon ethanol, demonstrating a critical role of CaMKII-mediated NaV1.5 phosphorylation for acute ethanol effects. Conclusion We show that ethanol acutely induces late INa upstream of arrhythmogenic SR Ca2+-leak and delayed afterdepolarizations, dependent on NaV1.5 S571 phosphorylation, leading to atrial fibrillation in vivo. Importantly, these effects as well as AF in vivo can be prevented using the late INa inhibitor ranolazine. We suggest that this antianginal drug could be repurposed to prevent ethanol-induced AF.Late INa (Patch-Clamp)In vivo AF (programmed stimulation)

Adolescent Alcohol Exposure Produces Sex-Specific Long-term Hyperalgesia via Changes in Central Amygdala Circuit Function

BackgroundExposure to alcohol during adolescence produces many effects that last well into adulthood. Acute alcohol use is analgesic, and people living with pain report drinking alcohol to reduce pain, but chronic alcohol use produces increases in pain sensitivity. MethodsWe tested the acute and lasting effects of chronic adolescent intermittent ethanol (AIE) exposure on pain-related behavioral and brain changes in male and female rats. We also tested the long-term effects of AIE on synaptic transmission in midbrain (ventrolateral periaqueductal gray [vlPAG])-projecting central amygdala (CeA) neurons using whole-cell electrophysiology. Finally, we used circuit-based approaches (DREADDs [designer receptors exclusively activated by designer drugs]) to test the role of vlPAG-projecting CeA neurons in mediating AIE effects on pain-related outcomes. ResultsAIE produced long-lasting hyperalgesia in male, but not female, rats. Similarly, AIE led to a reduction in synaptic strength of medial CeA cells that project to the vlPAG in male, but not female, rats. Challenge with an acute painful stimulus (i.e., formalin) in adulthood produced expected increases in pain reactivity, and this effect was exaggerated in male rats with a history of AIE. Finally, CeA-vlPAG circuit activation rescued AIE-induced hypersensitivity in male rats. ConclusionsOur findings are the first, to our knowledge, to show long-lasting sex-dependent effects of adolescent alcohol exposure on pain-related behaviors and brain circuits in adult animals. This work has implications for understanding the long-term effects of underage alcohol drinking on pain-related behaviors in humans.

Acute and Chronic Ethanol Effects during Adolescence on Neuroimmune Responses: Consequences and Potential Pharmacologic Interventions.

Heavy ethanol consumption during adolescence has been linked to neuroimmune response dysregulation and cognitive deficits in the developing adolescent brain. During adolescence, the brain is particularly susceptible to the pharmacological effects of ethanol that are induced by acute and chronic bouts of exposure. Numerous preclinical rodent model studies have used different ethanol administration techniques, such as intragastric gavage, self-administration, vapor, intraperitoneal, and free access, and while most models indicated proinflammatory neuroimmune responses in the adolescent brain, there are various factors that appear to influence this observation. This review synthesizes the most recent findings of the effects of adolescent alcohol use on toll-like receptors, cytokines, and chemokines, as well as the activation of astrocytes and microglia with an emphasis on differences associated with the duration of ethanol exposure (acute vs. chronic), the amount of exposure (e.g., dose or blood ethanol concentrations), sex differences, and the timing of the neuroimmune observation (immediate vs. persistent). Finally, this review discusses new therapeutics and interventions that may ameliorate the dysregulation of neuroimmune maladaptations after ethanol exposure.

CD5 knockout modulates effects of alcohol consumption in mouse models (P1-12.008)

<h3>Objective:</h3> We seek to elucidate the impact of cluster of differentiation 5 (CD5) expression on the effects of acute ethanol (EtOH) exposure. <h3>Background:</h3> CD5 is a transmembrane glycoprotein expressed on the surface of T and B1 cells. It has been previously shown that CD5 expression is altered by chronic EtOH consumption. Additionally, GABA neurons in the ventral tegmental area (VTA) are inhibited by EtOH, while dopamine neurons are excited. Given that some CD5+ lymphocytes may be involved in the regulation of inhibitory GABA synapses, we hypothesize that CD5 knockout may modulate the behavioral and physiological effects of EtOH consumption. <h3>Design/Methods:</h3> CD5 knockout (CD5KO) mice were compared to C57BL/6J wild type (WT) mice. CD5KO was confirmed with flow cytometry. Behavioral changes were measured via open field behavior, loss of righting reflex, and 24-hour access two bottle choice drinking. Neuronal dynamics were assessed using single-unit recordings, loose patch clamp recordings, fast scan cyclic voltammetry, and qRTPCR. <h3>Results:</h3> CD5KO mice displayed reduced EtOH consumption versus WT mice in two bottle choice drinking as well as decreased reduction in locomotor activity at low dose (0.5–2.0g/Kg) EtOH and decreased sedation at high dose (4.0g/Kg). Paired-pulse frequency response measure of dopamine release also differed between CD5KO and WT strains. Additionally, decreased levels of dopamine transporter (DAT) RNA were found in the nucleus accumbens. <h3>Conclusions:</h3> CD5KO mice may be less sensitive to some of the effects of EtOH, suggesting that CD5 may play a role in EtOH consumption and sedation. <b>Disclosure:</b> Mr. Amendolara has nothing to disclose. Dr. Payne has nothing to disclose. Dr. Obray has nothing to disclose. Mr. Williams has nothing to disclose. Ms. Small has nothing to disclose. The institution of Dr. Yorgason has received research support from American Osteopathic Association. Dr. Edwards has nothing to disclose. Dr. Weber has nothing to disclose. Dr. Steffensen has stock in Syzygy Innovations, LLC. Dr. Steffensen has received intellectual property interests from a discovery or technology relating to health care.

The role of C1473G polymorphism in triptophanhydroxylase-2 gene in the acute ethanol effects on &lt;i&gt;c-fos&lt;/i&gt; gene expression and metabolism of biogenic amines in brain of mice

Tryptophan hydroxylase-2 is a key enzyme in the synthesis of the neurotransmitter serotonin, which plays an important role in the mechanisms of regulation of a wide range of physiological functions and behaviors. In this work, we studied the effect of acute ethanol administration on the expression of the c-fos early response gene and the metabolism of serotonin and catecholamines in the brain structures of B6-1473C and B6-1473G congenic mice, which differ in the C1473G single-nucleotide substitution in the Tph2 gene and enzyme activity. Acute alcoholization led to a significant increase in c-fos gene expression in the frontal cortex and striatum of B6-1473G mice and the hippocampus of B6-1473C mice. The introduction of ethanol caused a decrease in the index of serotonin metabolism in the nucleus accumbens in B6-1473C mice, as well as in the hippocampus and striatum of B6-1473G mice. Also, ethanol led to a decrease in the level of norepinephrine in the hypothalamus in B6-1473C mice. Thus, the C1473G polymorphism in the Tph2 gene has a significant effect on changes in the c-fos gene expression pattern and metabolism of biogenic amines in the brain induced by acute ethanol administration.

The Role of C1473G Polymorphism in Mouse Triptophan Hydroxylase 2 Gene in the Acute Effects of Ethanol on the c-fos Gene Expression and Metabolism of Biogenic Amines in the Brain

Tryptophan hydroxylase 2 is a key enzyme in the synthesis of the neurotransmitter serotonin, which plays an important role in the regulation of behavior and various physiological functions. We studied the effect of acute ethanol administration on the expression of the early response c-fos gene and metabolism of serotonin and catecholamines in the brain structures of B6-1473C and B6-1473G congenic mouse strains differing in the single-nucleotide substitution C1473G in the Tph2 gene and activity of the encoded enzyme. Acute alcoholization led to a significant upregulation of the c-fos gene expression in the frontal cortex and striatum of B6-1473G mice and in the hippocampus of B6-1473C mice and caused a decrease in the index of serotonin metabolism in the nucleus accumbens in B6-1473C mice and in the hippocampus and striatum of B6-1473G mice, as well as to the decrease in the norepinephrine level in the hypothalamus of B6-1473C mice. Therefore, the C1473G polymorphism in the Tph2 gene has a significant effect of acute ethanol administration on the c-fos expression pattern and metabolism of biogenic amines in the mouse brain.

Using animal models to identify clinical risk factors in the older population due to alcohol use and misuse.

The number of people over the age of 65 years old is increasing and understanding health risks associated with the aged population is important. Recent research has revealed that alcohol (ethanol) consumption levels in older demographics remains elevated and often occurs in a dangerous binge pattern. Given ethical constraints on investigating high level or binge pattern alcohol consumption in humans, animal models are often used to study the effects of ethanol. The current review highlights ongoing work revealing that aged rats are often more sensitive to the effects of acute ethanol compared to younger rats. Specifically, aged rats are more sensitive to the motor impairing, hypnotic, hypothermic, and often the cognitive effects of ethanol compared to younger rats. In addition, the development of ethanol tolerance following chronic exposure may have a different temporal pattern in aged rats compared to younger rats. However, the neurobiological mechanisms that cause the increased sensitivity to ethanol in aged animals have yet to be identified. Furthermore, the differential age effects of ethanol highlight clinical risk factors for alcohol misuse in the older human population. Future work is needed to determine underlying CNS mechanisms producing altered effects of ethanol in aged subjects and also the development of educational material concerning ethanol's effects across ages for health care providers working with the aged population.

Influence of early-life adversity on responses to acute and chronic ethanol in female mice.

Stressful early-life experiences increase the risk of developing an alcohol use disorder. We previously found that male C57BL/6J mice reared under limited bedding and nesting (LBN) conditions, a model of early-life adversity, escalate their ethanol intake in limited-access two-bottle choice (2BC) sessions faster than control (CTL)-reared counterparts when exposed to chronic intermittent ethanol (CIE) vapor inhalation. However, the alcohol consumption of female littermates was not affected by LBN or CIE. In the present study, we sought to determine whether this phenotype reflected a general insensitivity of female mice to the influence of early-life stress on alcohol responses. In a first experiment, CTL and LBN females with a history of 2BC combined or not with CIE were tested in affective and nociceptive assays during withdrawal. In a second group of CTL and LBN females, we examined ethanol-induced antinociception, sedation, plasma clearance, and c-Fos induction. In females withdrawn from chronic 2BC, CIE increased digging, reduced grooming, and increased immobility in the tail suspension test regardless of early-life history. In contrast, LBN rearing lowered mechanical nociceptive thresholds regardless of CIE exposure. In females acutely treated with ethanol, LBN rearing facilitated antinociception and delayed the onset of sedation without influencing ethanol clearance rate or c-Fos induction in the paraventricular nucleus of the hypothalamus, paraventricular nucleus of the thalamus, central nucleus of the amygdala, or auditory cortex. CIE withdrawal produced multiple indices of negative affect in C57BL/6J females, suggesting that their motivation to consume alcohol may differ from air-exposed counterparts despite equivalent intake. Contrasted with our previous findings in males, LBN-induced mechanical hyperalgesia in chronic alcohol drinkers was specific to females. Lower nociceptive thresholds combined with increased sensitivity to the acute antinociceptive effect of ethanol may contribute to reinforcing ethanol consumption in LBN females but are not sufficient to increase their intake.

Unraveling the epigenomic and transcriptomic interplay during alcohol-induced anxiolysis

Positive effects of alcohol drinking such as anxiolysis and euphoria appear to be a crucial factor in the initiation and maintenance of alcohol use disorder (AUD). However, the mechanisms that lead from chromatin reorganization to transcriptomic changes after acute ethanol exposure remain unknown. Here, we used Assay for Transposase-Accessible Chromatin followed by high throughput sequencing (ATAC-seq) and RNA-seq to investigate epigenomic and transcriptomic changes that underlie anxiolytic effects of acute ethanol using an animal model. Analysis of ATAC-seq data revealed an overall open or permissive chromatin state that was associated with transcriptomic changes in the amygdala after acute ethanol exposure. We identified a candidate gene, Hif3a (Hypoxia-inducible factor 3, alpha subunit), that had ‘open’ chromatin regions (ATAC-seq peaks), associated with significantly increased active epigenetic histone acetylation marks and decreased DNA methylation at these regions. The mRNA levels of Hif3a were increased by acute ethanol exposure, but decreased in the amygdala during withdrawal after chronic ethanol exposure. Knockdown of Hif3a expression in the central nucleus of amygdala attenuated acute ethanol-induced increases in Hif3a mRNA levels and blocked anxiolysis in rats. These data indicate that chromatin accessibility and transcriptomic signatures in the amygdala after acute ethanol exposure underlie anxiolysis and possibly prime the chromatin for the development of AUD.

Ethanol inhibits pancreatic projecting neurons in the dorsal motor nucleus of the vagus

Alcohol use disorder (AUD) is a rapidly growing concern in the United States. Current trending escalations of alcohol use are associated with a concurrent rise in alcohol-related end-organ damage, increasing risk for further diseases. Alcohol-related end-organ damage can be driven by autonomic nervous system dysfunction, however studies on alcohol effects on autonomic control of end-organ function are lacking. Alcohol intake has been shown to reduce insulin secretions from the pancreas. Pancreatic insulin release is controlled in part by preganglionic parasympathetic motor neurons residing in the dorsal motor nucleus of the vagus (DMV) that project to the pancreas. How these neurons are affected by alcohol exposure has not been directly examined. Here we investigated the effects of acute ethanol (EtOH) application on DMV pancreatic-projecting neurons with whole-cell patch-clamp electrophysiology. We found that bath application of EtOH (50 mM) for greater than 30 min significantly enhanced the frequency of spontaneous inhibitory post synaptic current (sIPSC) events of DMV pancreatic-projecting neurons suggesting a presynaptic mechanism of EtOH to increase GABAergic transmission. Thirty-minute EtOH application also decreased action potential firing of these neurons. Pretreatment of DMV slices with 20 μM fluoxetine, a selective serotonin reuptake inhibitor, also increased GABAergic transmission and decreased action potential firing of these DMV neurons while occluding any further effects of EtOH application, suggesting a critical role for serotonin in mediating EtOH effects in the DMV. Ultimately, decreased DMV motor output may lead to alterations in pancreatic secretions. Further studies are needed to fully understand EtOH’s influence on DMV neurons as well as the consequences of changes in parasympathetic output to the pancreas.

The ethanol inhibition of basolateral amygdala neuron spiking is mediated by a γ-aminobutyric acid type A-mediated tonic current.

The basolateral nucleus of the amygdala (BLA) plays an important role in the development of fear and anxiety-related behaviors. The BLA receives inputs from all sensory stimuli. After processing those stimuli, BLA neurons signal neurons within the central amygdala and other brain regions, including the ventral and dorsal striatum and frontal cortex. Studies suggest that the BLA is involved in drug dependence and in the reinforcing actions of ethanol. For example, acute exposure to ethanol reduces anxiety, while withdrawal from chronic ethanol exposure alters BLA synaptic transmission, which increases anxiety, a common underlying cause of relapse. Exposure to and withdrawal from chronic alcohol also disrupts many brain areas that connect with the BLA. Despite these important findings, the acute actions of alcohol on the intrinsic excitability of BLA neurons have not been fully characterized. Brain slices containing the BLA were prepared from adult C57BL/6J male mice. Whole-cell and sharp electrode electrophysiological recordings were performed to characterize the effects of acute ethanol on BLA neuronal and astrocyte function, respectively. Ethanol inhibited action potential (AP) firing of BLA neurons but had no effect on BLA astrocyte resting membrane potential. The ethanol-induced inhibition of firing was concentration-dependent (11 to 66 mM) and accompanied by a reduction in the input resistance and an increase in the rheobase of BLA neurons. The inhibitory effect of ethanol was suppressed by picrotoxin, which blocks both γ-aminobutyric acid type A (GABAA ) and glycine receptors, but not by the selective glycine receptor antagonist strychnine, which suggests an involvement of GABAA receptors. Ethanol did not affect spontaneous inhibitory postsynaptic currents suggesting that the inhibition of BLA neuronal excitability by ethanol was not due to an increase in GABAA -mediated synaptic transmission. However, acute ethanol enhanced the amplitude of the holding current of BLA neurons, an effect that was prevented by picrotoxin, which by itself reduced the holding current. These results suggest that BLA neurons express a GABA-mediated tonic current that is enhanced by acute ethanol, which leads to reduced excitability of BLA neurons.

Alcohol Dependence Induces CRF Sensitivity in Female Central Amygdala GABA Synapses.

Alcohol use disorder (AUD) is a chronically relapsing disease characterized by loss of control in seeking and consuming alcohol (ethanol) driven by the recruitment of brain stress systems. However, AUD differs among the sexes: men are more likely to develop AUD, but women progress from casual to binge drinking and heavy alcohol use more quickly. The central amygdala (CeA) is a hub of stress and anxiety, with corticotropin-releasing factor (CRF)-CRF1 receptor and Gamma-Aminobutyric Acid (GABA)-ergic signaling dysregulation occurring in alcohol-dependent male rodents. However, we recently showed that GABAergic synapses in female rats are less sensitive to the acute effects of ethanol. Here, we used patch-clamp electrophysiology to examine the effects of alcohol dependence on the CRF modulation of rat CeA GABAergic transmission of both sexes. We found that GABAergic synapses of naïve female rats were unresponsive to CRF application compared to males, although alcohol dependence induced a similar CRF responsivity in both sexes. In situ hybridization revealed that females had fewer CeA neurons containing mRNA for the CRF1 receptor (Crhr1) than males, but in dependence, the percentage of Crhr1-expressing neurons in females increased, unlike in males. Overall, our data provide evidence for sexually dimorphic CeA CRF system effects on GABAergic synapses in dependence.

Sex-related differences in pattern of ethanol drinking under the intermittent-access model and its impact on exploratory and anxiety-like behavior in Long-Evans rats.

While men in the United States consume more alcohol than women, rates of drinking are converging. Nevertheless, females remain underrepresented in preclinical alcohol research. Here, we examined rats' sex-related differences in patterns of ethanol (EtOH) drinking and the effects of this drinking on exploratory and anxiety-like behavior. Adult male and female Long-Evans rats were given 20% ethanol under the intermittent-access two-bottle-choice paradigm. Their intake was measured daily for the first 7weeks. During the eighth week, intake was measured over the 24h of daily access. During the ninth week, they, along with EtOH-naive controls, were tested prior to daily access in a novel chamber, light-dark box, and hole board apparatus. During the tenth week, blood ethanol concentration (BEC) was assessed after 30 to 40min of access. Females overall demonstrated higher ethanol intake and preference across all access weeks than males, although only half of females drank significantly more than males. Across 24h of daily access, both sexes had their highest intake in the first 30min and their lowest in the middle of the light phase of the light/dark cycle. Despite their greater ethanol intake, females did not show significantly different BECs than males. In behavioral tests, females showed less vertical time in a novel activity chamber, more movement between chambers in a light-dark box, and more nose pokes in a hole-board apparatus than males. While a history of ethanol drinking led to a trend for lower vertical time in the activity chamber and greater chamber entries in the light-dark box, the effects were not sex-dependent. These results suggest that female and male rats could both be tested for acute effects of ethanol after 30min of daily access, but that nuanced considerations are needed in the design of these experiments and the interpretation of their findings.

Sex differences in the acute ethanol effects on object recognition memory: influence of estrous cycle.

Effects of acute ethanol (EtOH) on memory depend on several factors, including type of behavioral task. Sex differences in EtOH effects have been reported in humans and animals, and recognition memory can be influenced by circulating sex hormones. The aim of this study was to investigate the influence of sex and estrous cycle in the acute effects of EtOH on novel object recognition memory in rats. Male and female Wistar rats were part of one of the groups: control, 0.6-g/kg EtOH and 1.8-g/kg EtOH (administered intraperitoneally before the training session). The estrous cycle was evaluated by vaginal smear. The task was conducted in an open field arena. During training, animals were exposed to two identical objects, and test sessions were performed 1 h (short-term) and 24 h (long-term) later. One of the objects was changed in each test. Increased novel object exploration was shown by male and female controls in the short- and long-term tests, respectively. In the short-term test, females did not show preference for the novel object, and EtOH 1.8 g/kg impaired performance in males. In the long-term test, both sexes showed object discrimination, and 1.8-g/kg EtOH reduced preference for the new object in male rats. The phase of the cycle, the performance on proestrus was worse compared with other phases, and EtOH failed to impair performance mainly on estrous. In conclusion, while male rats displayed ethanol-induced recognition memory deficit, female rats were unaffected by EtOH impairing effects. In addition, the performance of female rats was influenced by the estrous cycle phases.

Psychoactive Effects of Lactobacillus johnsonii BS15 on Preventing Memory Dysfunction Induced by Acute Ethanol Exposure Through Modulating Intestinal Microenvironment and Improving Alcohol Metabolic Level.

The negative effects of ethanol (EtOH) abuse on the body have been widely reported in recent years. Building on the microbiota-gut-brain axis hypothesis, our study aimed to demonstrate the potential psychobiotic role of Lactobacillus johnsonii BS15 in the preventive effects of acute EtOH intake on memory impairment. We also determined whether L. johnsonii BS15 intake could effectively improve resistance to acute drinking and alleviate the adverse effects of EtOH. Male mice were fed L. johnsonii BS15 orally with (Probiotic group) or without (Control and Alcohol groups) daily dose of 0.2 × 109 CFU/ml per mouse for 28 days. Gavage with L. johnsonii BS15 significantly modified the ileal microbial ecosystem (assessed by 16S rRNA gene sequencing) in favor of Firmicutes and Lactobacillus, indicating the ability of BS15 to restore the gut microbiota. The acute EtOH exposure model (7 g/kg EtOH per mice) was established by gavage, which was administered to the alcohol and probiotic groups on day 28 of the experiment. The L. johnsonii BS15 intake effectively reduced alcohol unconsciousness time, blood alcohol concentration, and serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels. Meanwhile, the improvement of ethanol resistance time and the activities of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) in the liver were shown by BS15 in acute alcohol-induced mice. We found that acute EtOH exposure reduced the exploration ratio (assessed by the novel object recognition test), escape latency, number of errors (assessed by passive avoidance test), and spontaneous exploration (assessed by T-maze test) in mice, which were obviously improved by L. johnsonii BS15. In the hippocampus, L. johnsonii BS15 significantly reversed the decrease in antioxidant capacity of superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione (GSH) and mRNA expression of memory-related functional proteins of brain-derived neurotrophic factor (BDNF) and cyclic ampresponse element binding protein (CREB) in the hippocampal tissue after acute EtOH exposure. In conclusion, L. johnsonii BS15 intake appears as a promising psychoactive therapy to ameliorate alcohol-mediated memory impairment by increasing EtOH metabolic levels.

Behavioral effects of ethanol in the Red Swamp Crayfish (Procambarus clarkii).

Alcohol abuse remains one of the primary preventable sources of mortality in the United States. Model species can be used to evaluate behavioral and other biological changes associated with alcohol and to identify novel treatments. This report describes methods for evaluating the behavioral effects of ethanol (EtOH) in crayfish. Crayfish (Procambarus clarkii) were immersed in ethanol concentrations ranging from 0.1 to 1.0 molar, for 10-30 min. Studies evaluated hemolymph alcohol concentration, locomotion in an open field and anxiety-like behavior using a Light/Dark transfer approach. EtOH immersion produced dose-dependent increases in hemolymph EtOH (up to 249 mg/dL) and reductions in open field locomotion that depended on EtOH concentration or exposure duration. Untreated crayfish exhibit avoidance of the open parts of the locomotor arena and a preference for a covered portion. Acute EtOH immersion decreased time spent in the covered portion of the Light/Dark arena, consistent with a decrease in anxiety-like behavior. Daily EtOH immersion for 5 days did not alter locomotor responses, however, activity was increased 3 days after the repeated EtOH regimen. Overall, this study shows that this inexpensive, easily maintained species can be used for behavioral pharmacological experiments designed to assess the acute and repeated effects of EtOH.

Effects of acute ethanol and/or diazepam exposure on immediate and delayed hippocampal metabolite levels in rats anesthetized with isoflurane.

Alcohol and benzodiazepines are psychoactive substances frequently associated in voluntary drug intoxications that share common mechanisms of action, including facilitation of GABAergic transmission. This study aimed to assess the separate and combined effects of ethanol and diazepam acute exposure on hippocampal metabolite levels, as well as on delayed cognitive performance, in rats anesthetized with isoflurane. Adult male Wistar rats received one intraperitoneal injection containing either saline solution ("CTL" group, N = 15), a 5-mg/kg dose of diazepam ("DIA" group, N = 16), a 2-g/kg dose of ethanol ("EtOH" group, N = 18), or a 5-mg/kg dose of diazepam + a 2-g/kg dose of ethanol ("DIA + EtOH" group, N = 24). The levels of brain metabolites in the hippocampal region were assessed using in vivo magnetic resonance spectroscopy (MRS) before and after injection. Behavioral testing, including working memory and visual recognition memory assessment, was performed at week 3, while a new MRS acquisition was conducted 4 weeks after the injection. In the hour following acute exposure, a decrease in glutamate levels was found in the DIA + EtOH group only. Four weeks after injection, a decrease in GABA and glutamate levels and an increase in NAA levels were found in the EtOH group only. No significant between-group differences were found in the behavioral assessment. While the initial decrease in glutamate levels in the DIA + EtOH group suggests an early potentiation effect between ethanol and diazepam, the long-term modifications found only in the EtOH group suggest a possible downregulation of ethanol's effect by diazepam at 4 weeks.

Assessment of BMP responses in an in vitro model of acute ethanol toxicity

The assay presented here was designed to assess the immediate effects of ethanol (EtOH) exposure on intracellular signaling activated by BMPs (Bone Morphogenetic Proteins). Previous reports of the relationship between EtOH exposure and BMP-dependent signaling have primarily assessed the expression of individual BMPs, changes in BMP target genes or effects on the phosphorylation level of key downstream mediators after days or weeks of in vivo EtOH exposure. What happens to BMP-stimulated signaling immediately following exposure to EtOH remains largely unexplored. Here, the early events of BMP-evoked intracellular signaling were examined in an in vitro model of acute EtOH toxicity. The BMP/Ethanol Stimulation Assay involved first stimulating cultured cells with recombinant BMPs. BMP-evoked intracellular signaling was then allowed to develop for 30 minutes. Next, the cells were exposed to a range of EtOH concentrations for an additional 30 minutes. Finally, the cultures were processed for Western blot analysis or immunofluorescent labeling.This short-term assay:• Permits investigation of EtOH exposure during the initial signaling events downstream of BMP receptor activation• Enables assessment of how the presence of BMPs might protect against cellular injury caused by toxic EtOH levels

Astrocytes play a critical role in mediating the effect of acute ethanol on central amygdala glutamatergic transmission

The Central Amygdala (CeA) has been heavily implicated in many aspects of alcohol use disorder. Ethanol (EtOH) has been shown to modulate glutamatergic transmission in the lateral subdivision of the CeA, however, the exact mechanism of this modulation is still unclear. EtOH exposure is associated with increased pro-inflammatory cytokines in the CeA, and inhibition of neuroimmune cells (microglia and astrocytes) has previously been shown to reduce EtOH drinking in animal models. Since neuroimmune activation seems to be involved in many of the effects of EtOH, we hypothesized that acute EtOH exposure will increase excitatory glutamatergic transmission in the CeA via modulation of neuroimmune cells. Using ex vivo brain slice whole-cell patch clamp electrophysiology, it was found that a physiologically relevant concentration of EtOH (20 mM) significantly increased presynaptic glutamatergic transmission in the CeA. Pharmacologic and chemogenetic inhibition of astrocyte function significantly reduced the ability of EtOH to modulate CeA glutamatergic transmission with minimal impact of microglia inhibition. This finding prompted additional studies examining whether direct neuroimmune activation through lipopolysaccharide (LPS) might lead to an increase in the glutamatergic transmission in the CeA. It was found that LPS modulation of glutamatergic transmission was limited by microglia activation and required astrocyte signaling. Taken together these results support the hypothesis that acute EtOH enhances lateral CeA glutamatergic transmission through an astrocyte mediated mechanism.This article is part of the special Issue on ‘Neurocircuitry Modulating Drug and Alcohol Abuse'.