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Stress is a risk factor that plays a considerable role in the development and maintenance of alcohol (ethanol) abuse and relapse. Preclinical studies examining ethanol-stress interactions have demonstrated elevated ethanol drinking, cognitive deficits, and negative affective behaviors in mice. However, the neural adaptations in prefrontal cortical regions that drive these aberrant behaviors produced by ethanol-stress interactions are unknown. In this study, male C57BL/6J mice were exposed to chronic intermittent ethanol (CIE) and repeated forced swim stress (FSS). After two cycles of CIE x FSS, brain slices containing the prelimbic (PrL) and infralimbic (IfL) cortex were prepared for analysis of adaptations in dendritic spines and synaptic plasticity. In the PrL cortex, total spine density was increased in mice exposed to CIE. Immediately following induction of long-term potentiation (LTP), the fEPSP slope was increased in the PrL of CIE x FSS treated mice, indicative of a presynaptic adaptation on post-tetanic potentiation (PTP). In the IfL cortex, CIE exposure regardless of FSS experience resulted in an increase in spine density. FSS alone or when combined with CIE exposure increased PTP following LTP induction. Repeated FSS episodes increased IfL cortical paired-pulse facilitation, a second measure of presynaptic plasticity. In summary, CIE exposure resulted in structural adaptations while repeated stress exposure drove metaplastic changes in presynaptic function, demonstrating distinct morphological and functional changes in PrL and IfL cortical neurons. Thus, the structural and functional adaptations may be one mechanism underlying the development of excessive drinking and cognitive deficits associated with ethanol-stress interactions.

Decades of work in Aplysia californica established the general rule that principles of synaptic plasticity and their molecular mechanisms are evolutionarily conserved from mollusks to mammals. However, an exquisitely sensitive, activity-dependent homosynaptic mechanism that protects against the depression of neurotransmitter release in Aplysia sensory neuron terminals has, to date, not been uncovered in other animals, including mammals. Here, we discover that depression at a mammalian synapse that is implicated in habit formation and habit learning acceleration by ethanol, the fast-spiking interneuron (FSI) to medium spiny principal projection neuron (MSN) synapse of the dorsolateral striatum, is subject to this type of synaptic protection. We show that this protection against synaptic depression is calcium- and PDZ domain interaction-dependent. These findings support activity dependent protection against synaptic depression as an Aplysia-like synaptic switch in mammals that may represent a leveraging point for treating alcohol use disorders.

Exposure to ethanol during the last trimester equivalent of human pregnancy causes apoptotic neurodegeneration in the developing brain, an effect that is thought to be mediated, in part, by inhibition of NMDA receptors. However, NMDA receptors can rapidly adapt to the acute effects of ethanol and are ethanol resistant in some populations of developing neurons. Here, we characterized the effect of ethanol on NMDA and non-NMDA receptor-mediated synaptic transmission in the retrosplenial cortex (RSC), a brain region involved in the integration of different modalities of spatial information that is among the most sensitive regions to ethanol-induced neurodegeneration. A single 4-h exposure to ethanol vapor of 7-day-old transgenic mice that express the Venus fluorescent protein in interneurons triggered extensive apoptosis in the RSC. Slice electrophysiological recordings showed that bath-applied ethanol inhibits NMDA and non-NMDA receptor excitatory postsynaptic currents (EPSCs) in pyramidal neurons and interneurons; however, we found no evidence of acute tolerance development to this effect after the 4-h in-vivo ethanol vapor exposure. Acute bath application of ethanol reduced action potential firing evoked by synaptic stimulation to a greater extent in pyramidal neurons than interneurons. Submaximal inhibition of NMDA EPSCs, but not non-NMDA EPSCs, mimicked the acute effect of ethanol on synaptically-evoked action potential firing. These findings indicate that partial inhibition of NMDA receptors by ethanol has sizable effects on the excitability of glutamatergic and GABAergic neurons in the developing RSC, and suggest that positive allosteric modulators of these receptors could ameliorate ethanol intoxication-induced neurodegeneration during late stages of fetal development.

Reducing expression or inhibiting translocation of protein kinase C epsilon (PKCε) prolongs ethanol intoxication and decreases ethanol consumption in mice. However, we do not know if this phenotype is due to reduced PKCε kinase activity or to impairment of kinase-independent functions. In this study, we used a chemical-genetic strategy to determine whether a potent and highly selective inhibitor of PKCε catalytic activity reduces ethanol consumption. We generated ATP analog-specific PKCε (AS-PKCε) knock-in mice harboring a point mutation in the ATP binding site of PKCε that renders the mutant kinase highly sensitive to inhibition by 1-tert-butyl-3-naphthalen-1-ylpyrazolo[3,4-d]pyrimidin-4-amine (1-NA-PP1). Systemically administered 1-NA-PP1 readily crossed the blood brain barrier and inhibited PKCε-mediated phosphorylation. 1-NA-PP1 reversibly reduced ethanol consumption by AS-PKCε mice but not by wild type mice lacking the AS-PKCε mutation. These results support the development of inhibitors of PKCε catalytic activity as a strategy to reduce ethanol consumption, and they demonstrate that the AS- PKCε mouse is a useful tool to study the role of PKCε in behavior.

Stressful life events and chronic stressors have been associated with escalations in alcohol drinking. Stress exposure leads to the secretion of glucocorticoids (cortisol in the human; corticosterone (CORT) in the rodent). To model a period of heightened elevations in CORT, the present work assessed the effects of chronic exposure to the stress hormone CORT on alcohol self-administration. Male Long Evans rats were trained to self-administer a sweetened alcohol solution (2% sucrose/15% alcohol) resulting in moderate levels of daily alcohol intake (0.5–0.7 g/kg). Following stable baseline operant self-administration, rats received CORT in the drinking water for 7 days. A transient increase in alcohol self-administration was observed on the first self-administration session following CORT exposure, and behavior returned to control levels by the second session. Control experiments determined that this increase in alcohol self-administration was specific to alcohol, unrelated to general motor activation, and functionally dissociated from decreased CORT levels at the time of testing. These results indicate that repeated exposure to heightened levels of stress hormone (e.g., as may be experienced during stressful episodes) has the potential to lead to exacerbated alcohol intake in low to moderate drinkers. Given that maladaptive drinking patterns, such as escalated alcohol drinking following stressful episodes, have the potential to put an individual at risk for future drinking disorders, utilization of this model will be important for examination of neuroadaptations that occur as a consequence of CORT exposure in order to better understand escalated drinking following stressful episodes in nondependent individuals.

La consommation d'alcool est un phénomène répandu dans le monde entier. Cependant, la façon dont la consommation d'alcool à des fins récréatives évolue en trouble de la consommation d'alcool (AUD) reste mal comprise. Le gène Smpd3 et sa protéine codée sphingomyélinase neutre (NSM) sont associés à la consommation d'alcool chez l'homme et aux comportements liés à l'alcool chez la souris, suggérant un rôle potentiel dans cette transition. À l'aide de l'imagerie par résonance magnétique multiparamétrique, nous avons caractérisé le rôle de la NSM dans les effets aigus et chroniques de l'alcool sur l'anatomie et la fonction cérébrales chez la souris femelle. La consommation volontaire chronique d'alcool (16 % en volume pendant au moins 6 jours) a affecté l'anatomie cérébrale chez les souris WT, réduisant le volume de la structure régionale principalement dans les régions corticales. Une activité NSM atténuée a empêché ces changements anatomiques. L'IRM fonctionnelle a lié ces adaptations anatomiques aux changements fonctionnels : la consommation chronique d'alcool chez la souris a modulé de manière significative la connectivité fonctionnelle de l'état de repos (FC RS) en réponse à une provocation aiguë à l'éthanol (bolus i.p. de 2 g kg-1) chez les souris hétérozygotes knock-out NSM (Fro), mais pas chez les souris WT. L'administration aiguë d'éthanol chez des souris WT naïves d'alcool a significativement diminué le FC RS dans les régions corticales et du tronc cérébral, une découverte clé qui a été amplifiée chez les souris FRO. En ce qui concerne les effets pharmacologiques directs, l'administration aiguë d'éthanol a augmenté le volume sanguin cérébral régional (rCBV) dans de nombreuses zones du cerveau. Ici, la consommation chronique d'alcool a autrement atténué la réponse aiguë au rCBV chez les souris WT mais l'a améliorée chez les souris FRO. Au total, ces résultats suggèrent un rôle différentiel du NSM dans les réponses cérébrales fonctionnelles aiguës et chroniques à l'alcool. Par conséquent, le ciblage de la NSM peut être utile dans la prévention ou le traitement de l'AUD. El consumo de alcohol es un fenómeno generalizado en todo el mundo. Sin embargo, la forma en que el consumo recreativo de alcohol evoluciona hacia el trastorno por consumo de alcohol (AUD) sigue siendo poco conocida. El gen Smpd3 y su proteína codificada esfingomielinasa neutra (NSM) están asociados con el consumo de alcohol en humanos y comportamientos relacionados con el alcohol en ratones, lo que sugiere un papel potencial en esta transición. Utilizando imágenes de resonancia magnética multiparamétrica, caracterizamos el papel de NSM en los efectos agudos y crónicos del alcohol en la anatomía y función cerebral en ratones hembra. El consumo voluntario crónico de alcohol (16 vol.% durante al menos 6 días) afectó la anatomía cerebral en ratones WT, reduciendo el volumen de la estructura regional predominantemente en las regiones corticales. La actividad atenuada de NSM evitó estos cambios anatómicos. La MRI funcional vinculó estas adaptaciones anatómicas con cambios funcionales: el consumo crónico de alcohol en ratones moduló significativamente la conectividad funcional en estado de reposo (RS FC) en respuesta a un desafío agudo con etanol (bolo i.p. de 2 g kg-1) en ratones knockout NSM heterocigotos (Fro), pero no en ratones WT. La administración aguda de etanol en ratones WT sin tratamiento previo con alcohol disminuyó significativamente la FC RS en las regiones cortical y del tronco encefálico, un hallazgo clave que se amplificó en ratones Fro. En cuanto a los efectos farmacológicos directos, la administración aguda de etanol aumentó el volumen sanguíneo cerebral regional (rCBV) en muchas áreas del cerebro. En este caso, el consumo crónico de alcohol atenuó la respuesta aguda de rCBV en ratones WT, pero la mejoró en ratones Fro. En conjunto, estos hallazgos sugieren un papel diferencial de la NSM en las respuestas cerebrales funcionales agudas y crónicas al alcohol. Por lo tanto, dirigirse a la NSM puede ser útil en la prevención o el tratamiento de la AUD. Alcohol consumption is a widespread phenomenon throughout the world. However, how recreational alcohol use evolves into alcohol use disorder (AUD) remains poorly understood. The Smpd3 gene and its coded protein neutral sphingomyelinase (NSM) are associated with alcohol consumption in humans and alcohol-related behaviors in mice, suggesting a potential role in this transition. Using multiparametric magnetic resonance imaging, we characterized the role of NSM in acute and chronic effects of alcohol on brain anatomy and function in female mice. Chronic voluntary alcohol consumption (16 vol.% for at least 6 days) affected brain anatomy in WT mice, reducing regional structure volume predominantly in cortical regions. Attenuated NSM activity prevented these anatomical changes. Functional MRI linked these anatomical adaptations to functional changes: Chronic alcohol consumption in mice significantly modulated resting state functional connectivity (RS FC) in response to an acute ethanol challenge (i.p. bolus of 2 g kg-1) in heterozygous NSM knockout (Fro), but not in WT mice. Acute ethanol administration in alcohol-naïve WT mice significantly decreased RS FC in cortical and brainstem regions, a key finding that was amplified in Fro mice. Regarding direct pharmacological effects, acute ethanol administration increased the regional cerebral blood volume (rCBV) in many brain areas. Here, chronic alcohol consumption otherwise attenuated the acute rCBV response in WT mice but enhanced it in Fro mice. Altogether, these findings suggest a differential role for NSM in acute and chronic functional brain responses to alcohol. Therefore, targeting NSM may be useful in the prevention or treatment of AUD. استهلاك الكحول ظاهرة واسعة الانتشار في جميع أنحاء العالم. ومع ذلك، لا تزال كيفية تطور تعاطي الكحول الترفيهي إلى اضطراب تعاطي الكحول (AUD) غير مفهومة بشكل جيد. يرتبط جين Smpd3 وأنزيم السفينغوميليناز المحايد للبروتين المشفر (NSM) باستهلاك الكحول في البشر والسلوكيات المتعلقة بالكحول في الفئران، مما يشير إلى دور محتمل في هذا الانتقال. باستخدام التصوير بالرنين المغناطيسي متعدد المعلمات، قمنا بتمييز دور NSM في التأثيرات الحادة والمزمنة للكحول على تشريح الدماغ ووظيفته في إناث الفئران. يؤثر استهلاك الكحول الطوعي المزمن (16 ٪ لمدة 6 أيام على الأقل) على تشريح الدماغ في فئران WT، مما يقلل من حجم الهيكل الإقليمي في الغالب في المناطق القشرية. منع نشاط NSM الموهن هذه التغييرات التشريحية. ربط التصوير بالرنين المغناطيسي الوظيفي هذه التكيفات التشريحية بالتغيرات الوظيفية: قام استهلاك الكحول المزمن في الفئران بتعديل الاتصال الوظيفي لحالة الراحة بشكل كبير (RS FC) استجابة لتحدي الإيثانول الحاد (جرعة 2 جم كجم -1) في الضربة القاضية غير المتجانسة في NSM (FRO)، ولكن ليس في فئران WT. قلل إعطاء الإيثانول الحاد في فئران WT الساذجة الكحولية بشكل كبير من RS FC في المناطق القشرية وجذع الدماغ، وهو اكتشاف رئيسي تم تضخيمه في فئران Fro. فيما يتعلق بالتأثيرات الدوائية المباشرة، أدى تناول الإيثانول الحاد إلى زيادة حجم الدم الدماغي الإقليمي (rCBV) في العديد من مناطق الدماغ. هنا، أدى استهلاك الكحول المزمن إلى تخفيف استجابة rCBV الحادة في فئران WT ولكنه عززها في فئران Fro. إجمالاً، تشير هذه النتائج إلى دور تفاضلي لـ NSM في استجابات الدماغ الوظيفية الحادة والمزمنة للكحول. لذلك، قد يكون استهداف NSM مفيدًا في الوقاية من AUD أو علاجه.

The dopamine (DA) mesolimbic pathway, which originates from DA cell bodies within the ventral tegmental area (VTA), has been shown by various studies to play a role in the mediation of various drugs of abuse including alcohol (EtOH). It has been suggested that the VTA's control of EtOH reward is mediated in part by the D2 receptors within the VTA. These receptors may be under the regulation of reciprocal GABAergic inputs from forebrain components of the mesolimbic path such as the nucleus accumbens (NAcc), a classic EtOH reward substrate, and the bed nucleus of the stria terminalis, a substrate recently implicated in EtOH reinforcement, forming a self-regulating feedback loop. To test this hypothesis, D2 regulation of EtOH self-administration (SA) was evaluated by the microinfusion of the D2 antagonist eticlopride into the VTA of P rats, which produced profound reductions in EtOH SA in the highest (20.0 and 40.0microg) doses tested in both BST/VTA and NAcc/VTA implanted P rats. To determine the role of GABA in the mediation of EtOH SA, a 32.0ng dose the non-selective GABA antagonist SR 95531 was microinfused into the BST producing no effect on responding for EtOH and into the NAcc which lead to a reduction in EtOH responding. Finally, the hypothesis that GABA innervation of the VTA from the mesolimbic forebrain may influence EtOH SA was examined by the simultaneous infusion of eticlopride (40.0microg) into the VTA and SR 95531 (32.0ng) into either the BST or NAcc. This combination infusion completely attenuated the reduction in EtOH SA observed with the 40.0microg dose of eticlopride alone in both groups of animals. These results suggest that while the D2 receptors within the VTA regulate EtOH-motivated behaviors, this is modulated by GABAergic input from the mesolimbic forebrain, specifically from the BST and NAcc.

Alcoholism is a chronic relapsing disorder characterized by periods of heavy alcohol consumption and unsuccessful attempts at abstinence. Relapse is one of the most problematic aspects in the treatment of alcoholism and is triggered by ethanol-associated cues. Extinction-based cue exposure therapies have proven ineffective in the treatment of alcoholism. However, positive allosteric modulation of mGlu5 with CDPPB enhances the extinction learning of alcohol-seeking behavior. The current study investigated the impact of chronic alcohol exposure on the extinction of ethanol-seeking behavior. Adult Wistar rats were trained to self-administer alcohol with a light/tone stimulus serving as the alcohol cue. After training, one group of rats was exposed to chronic intermittent ethanol (CIE) daily for a period of 2 weeks to induce ethanol dependence. Control rats were exposed to air for the same period of time. Both groups were then retrained to self-administer ethanol and subsequently tested for changes in extinction learning. CIE exposed rats consumed more ethanol compared to their pre-CIE levels and to control rats. During extinction training, CIE rats responded significantly more on the previously active lever and required more sessions to reach extinction criteria compared to control rats. Treatment with CDPPB facilitated extinction in control rats and attenuated the increased resistance to extinction in CIE-exposed rats. These results demonstrate that chronic ethanol exposure not only alters ethanol intake, but also the extinction of ethanol-seeking behaviors. The ability to attenuate deficits through modulation of mGlu5 provides a potential target for pharmacological manipulation that could ultimately reduce relapse in alcoholics.