Cell-type specific central amygdala neurotransmission in alcohol dependence

项目摘要/摘要： 研究：酒精使用障碍非常普遍且代价高昂，只有少数有效 药物治疗是可用的。目前的证据表明，反复饮酒 诱导中央杏仁核(CEA)神经元内基因表达的变化，随着动物成为 酒精依赖，这些变化推动了病理性的酒精消费。例如，药理学 研究表明，反复酒精暴露会增加促肾上腺皮质激素释放因子(CRF)信号转导。 促进啮齿动物过度饮酒的CEA。然而，此CRF的来源不是 当然，CRF信号的增加也不太可能是酒精摄入量增加的唯一驱动因素 在这些动物身上。这个项目将检验这样一个假设，即外侧CEA内的神经元是这一现象的来源 CRF和这些神经元产生额外的神经肽，驱动过度饮酒。这 假设将通过检查饮酒行为来检验，在个别下调 BAC转基因CRH-CRE大鼠CEA-CRF神经元中CRF、强啡肽或GABA的神经递质 依赖Cre的RNA干扰。该项目还将使用一种公正的方法来探索酒精诱导的 用单核RNA-SEQ方法检测CEA中所有神经元的基因表达 当动物变得依赖酒精时，这些种群中受影响最大的基因。结果是 这项工作将导致关于特定蛋白质的新的、可检验的假说，这些蛋白质可能被靶向地减少 酒精依赖者过度饮酒。 培训：这项研究将对申请者进行几项实验技术的培训，包括啮齿动物饮酒 遗传工具的行为、测试和验证、单核测序、RNA定量以及 生物信息学分析工具。通过这次培训，申请者将发展分子使用方面的专业知识。 操纵基因表达的工具，以及在生物信息学方法中分析转录本。培训 该计划还将培养专业技能，包括科学写作和演示以及编程，以 此外，申请者的目标是成为一名独立的学术神经学家。

Project Summary/Abstract: Research: Alcohol use disorder is highly prevalent and costly, and only a few modestly effective pharmacological treatments are available. Current evidence indicates that repeated consumption of alcohol induces changes in gene expression within neurons in the central amygdala (CeA), and as animals become alcohol dependent, these changes drive pathological alcohol consumption. For example, pharmacological studies have shown that repeated alcohol exposure increases corticotropin-releasing factor (CRF) signaling in the CeA, which promotes excessive alcohol consumption in rodents. However, the source of this CRF is not certain and it is also unlikely that increased CRF signaling is the sole driver of increased alcohol consumption in these animals. This project will test the hypothesis that neurons within the lateral CeA are the source of this CRF and that these neurons produce additional neuropeptides that drive excessive alcohol drinking. This hypothesis will be tested by examining drinking behavior after individually downregulating the neurotransmitters CRF, dynorphin, or GABA in CeA-CRF neurons using a BAC transgenic Crh-Cre rat and Cre-dependent RNA interference. The project will also use an unbiased approach to explore alcohol-induced gene expression in all neurons of the CeA by using single nuclei RNA-seq to identify neuronal populations and the genes within these populations that are most affected as animals become alcohol dependent. The results of this work will lead to new, testable hypotheses about specific proteins that could be targeted to reduce excessive drinking in alcohol dependent individuals. Training: This research will train the applicant in several experimental techniques including rodent drinking behavior, testing and validation of genetic tools, single-nuclei sequencing, RNA quantification, and use of bioinformatics analysis tools. Through this training, the applicant will develop expertise in the use of molecular tools to manipulate gene expression and in bioinformatics approaches to analyze transcriptomes. The training program will also develop professional skills, including scientific writing and presentation, and programming, to further the applicant’s goal of becoming an independent academic neuroscientist.