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成本高昂，而且只有少数药物效果不大 可以采用药物治疗。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，并且随着动物变得 依赖酒精，这些变化会导致病理性饮酒。例如，药理学 研究表明，反复接触酒精会增加促肾上腺皮质激素释放因子（CRF）信号传导 CeA，促进啮齿类动物过度饮酒。然而，这个 CRF 的来源并不是 CRF 信号传导的增加是肯定的，但也不可能是饮酒量增加的唯一驱动因素 在这些动物中。该项目将检验以下假设：侧脑区神经元是这种现象的来源 CRF 以及这些神经元产生额外的神经肽，导致过量饮酒。这个 假设将通过检查单独下调饮酒行为后的饮酒行为来检验 使用 BAC 转基因 Crh-Cre 大鼠和 CeA-CRF 神经元中的神经递质 CRF、强啡肽或 GABA Cre 依赖性 RNA 干扰。该项目还将采用公正的方法来探索酒精引起的 通过使用单核 RNA-seq 来识别神经元群体和 CeA 所有神经元中的基因表达 当动物变得酒精依赖时，这些群体中受影响最严重的基因。结果 这项工作将带来关于特定蛋白质的新的、可测试的假设，这些蛋白质可以有针对性地减少 酒精依赖者过度饮酒。 培训：这项研究将对申请人进行多种实验技术的培训，包括啮齿动物饮水 行为、遗传工具的测试和验证、单核测序、RNA 定量以及使用 生物信息学分析工具。通过这次培训，申请人将发展分子技术应用方面的专业知识 操纵基因表达的工具和分析转录组的生物信息学方法。培训内容 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 进一步实现申请人成为独立学术神经科学家的目标。

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.