The effect of alcohol withdrawal on molecular and behavioral circadian rhythms

描述（由申请人提供）：本研究培训项目的重点是多次慢性酒精暴露-戒断周期对分子和行为昼夜节律的影响。睡眠可能是受大脑昼夜节律调节的最关键的生理过程。虽然人们对睡眠的具体机制及其重要性知之甚少，但睡眠障碍如果长期存在，可能会造成毁灭性的生理后果，包括死亡。睡眠问题是戒酒者最常见的抱怨之一，也是酗酒复发的常见原因之一。最近的研究强调了T型电压门控钙通道在调节分子“时钟”中的重要性，这些“时钟”被认为在大脑中产生昼夜节律。我们的实验室已经确定了t型钙通道的一种特殊变体-称为Ca(v)3.2 -其表达与慢性酒精暴露和戒断相关的昼夜节律紊乱密切相关。该项目的目的是评估多种慢性酒精暴露-戒断周期对小鼠正常分子和行为昼夜节律的影响，在小鼠模型中，T型钙通道要么被药理学抑制，要么被基因沉默。该方法将提供分子和行为实验室技术方面的培训，包括使用定量逆转录聚合酶链反应（qRT-PCR）分析和Western blot技术进行基因表达的相对定量，以及使用连续脑电图（EEG）监测进行行为分析和睡眠评分。此外，对结果的分析将提供生理节律数学建模的培训，以及对后续发现的统计分析和解释。导师、研究助理和博士后在实验室的指导和培训将帮助申请人解决实验问题，分析陷阱和意外结果，并开发创新的方法来回答在完成项目过程中出现的每个问题。总之，该项目将提供必要的培训和技能，以便在分子和行为水平上对与酒精滥用有关的生理问题进行严格的研究。

DESCRIPTION (provided by applicant): The focus of the research training project is the effect of multiple chronic alcohol exposure-withdrawal cycles on molecular and behavioral circadian rhythms. Sleep is possibly the most critical physiologic process regulated by circadian rhythms of activity in the brain. While not much is understood regarding the specific mechanisms underlying sleep and its importance, sleep disturbances can have devastating physiologic consequences, including death, if they persist over time. Problems with sleep are one of the most common complaints from individuals undergoing withdrawal from alcohol and also one of the more common reasons given for relapse into alcohol abuse. Recent research has highlighted the importance of the T -type voltage gated calcium channels in the regulation of the molecular "clock" believed to generate circadian rhythms in the brain. Our lab has identified a specific variant of the T-type calcium channels - called Ca(v)3.2 - whose expression correlates strongly with the circadian disturbances associated with chronic alcohol exposure and withdrawal. The aims of the project seek to evaluate the effect of multiple chronic alcohol exposure-withdrawal cycles on normal molecular and behavioral circadian rhythms in a mouse model in which this T -type calcium channel has been either pharmacologically inhibited or genetically silenced. The approach will provide training in both molecular and behavioral laboratory techniques, including relative quantitation of gene expression using quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis and Western blot techniques, as well as behavioral analysis and sleep scoring using continuous electroencephalographic (EEG) monitoring. Furthermore, the analysis of the results will provide training in the mathematical modeling of physiologic rhythms, as well as the statistical analysis and interpretation of subsequent findings. The guidance and training provided by the advisor, research associates, and postdoctoral fellows in the lab will assist the applicant in troubleshooting experiments, analyzing pitfalls and unexpected results, and developing innovative approaches to answering the questions that arise at each point in the process of completing the project. In summary, this project will provide the training and skills necessary to develop and carry out a rigorous study of a physiologic problem related to alcohol abuse on both a molecular and behavioral level. PUBLIC HEALTH RELEVANCE: According to the NIH and CDC, respectively, approximately 43 percent of Americans exhibit moderate-to-high risk drinking habits and 70 percent do not get enough sleep. Furthermore, 36 to 72 percent of chronic drinkers cite sleep problems as a major reason for their relapse into alcoholism from 'withdrawal and at least 30 percent of insomniacs use alcohol as a sleep-aid, putting them at increased risk of alcoholism. Hence, the relationship between alcohol and sleep disturbance is a matter of great importance to public health in the United States.