Mechanisms of Ethanol's Influence on Sleep and Circadian Rhythms

描述（由申请人提供）：两个大脑区域对睡眠的开始和维持特别重要。视交叉上核（SON）是睡眠/觉醒周期的主要发起者，丘脑参与产生和维持睡眠期间发生的大脑节律。以下具体目标将作为培训工具，并将寻求确定乙醇对睡眠和昼夜节律的细胞和分子影响。目的1：长期饮酒者的行为改变。小鼠将被长期给予乙醇饮食（T通道阻滞剂乙氧亚胺）。轮跑活动和录像记录将用于确定睡眠/清醒状态。我假设在长期饮酒者和服用乙氧亚胺的动物中，总睡眠时间减少，昼夜节律变化（延迟）。目的2：慢性饮酒者的分子改变。RTPCR和Western blot技术将用于检测慢性饮酒对T通道和分子钟基因和蛋白质的影响。我假设Aim 1中观察到的睡眠和昼夜节律的变化将与丘脑和SCN T通道亚型中基因（增加）和蛋白质（减少）表达的变化以及SCN分子钟基因和蛋白质（基因和蛋白质表达的明显节律性的衰减和丧失）的变化相关。目的3：慢性饮酒小鼠t通道的电生理特征。小鼠长期给予乙醇饮食将用于产生丘脑离体脑切片。在用适当的电压方案触发t电流时，将进行全细胞贴片记录。我假设在酒精患者中，t电流的神经适应性降低可能是梭形波振荡减少的基础。酒精对睡眠的干扰在酒精中毒的发展中起着关键作用。持续滥用乙醇会导致睡眠回路的长期变化，这种变化会持续到停止使用乙醇之后。这项研究将使我有机会了解慢性乙醇消耗如何改变睡眠回路中断的细胞和分子机制，并希望找到治疗乙醇对正常睡眠中断的新药物靶点。

DESCRIPTION (provided by applicant): Two brain regions are particularly important for the initiation and maintenance of sleep. The suprachiasmatic nucleus (SON) is the primary initiator of sleep/wake cycles and the thalamus is involved in the generation and maintenance of brain rhythms that occur during sleep. The following specific aims will serve as the training vehicle and will seek to determine the cellular and molecular influence of ethanol onsleep and circadian rhythms. Aim 1: Behavioral alterations in chronic drinkers. Mice will be chronically administered an ethanol diet ( the T channel blocker ethosuximide). Wheel running activity and videographic records will be used to determine sleep/waking states. I hypothesize a decrease in total sleep time and shifts in circadian phase (delays) in chronic drinkers and animals administered ethosuximide. Aim 2: Molecular alterations in chronic drinkers. RTPCR and Western blot techniques will be used to examine the effect of chronic alcohol consumption on T channel and molecular clock genes and proteins. I hypothesize that the changes in sleep and circadian rhythms observed in Aim 1, will be correlated with changes in gene (increased) and protein (decreased) expression in both thalamic and SCN T channel isoforms, as well as in SCN molecular clock genes and proteins (attenuation and loss of pronounced rhythmicity of gene and protein expression). Aim 3: Electrophysiological characterization of the T-channel in chronic drinking mice. Mice chronically administered an ethanol diet will be used to generate thalamic in vitro brain slices. Whole cell patch recordings will be made while evoking T-current with appropriate voltage protocols. I hypothesize a neuroadaptational decrease in T-current that may underlie the decrease in spindle wave oscillations reported in alcoholic patients. Sleep perturbations by ethanol play a key role in the progression of alcoholism. Continued abuse of ethanol leads to long-term changes in sleep circuitry that last well beyond the cessation of ethanol administration. This research will afford me the opportunity to understand how chronic ethanol consumption alters the cellular and molecular mechanisms underlying the reported disruptions in sleep circuitry, with the hope of identifying novel drug targets for treatment of ethanol's disruption of normal sleep.