TRANSCRIPTOME AND EPIGENOME MAPPING IN DOPAMINE NEURONS FROM THE OPIOID EXPOSED HUMAN BRAIN

尽管许多细胞和神经回路明显导致鸦片类药物和其他物质滥用障碍，但 药物成瘾的途径是通过中脑多巴胺能神经元。虽然是一种罕见的细胞类型(据估计 人类大脑中每200,000个神经元中只有1个是多巴胺能表型)， 多巴胺能神经传递被认为在成瘾的不同阶段发挥作用，从急性奖赏 机制和目标导向的行动，对习惯性行为的发展和增加的突显 与吸毒有关的线索，以及与戒毒有关的快感缺乏和烦躁不安。 令人惊讶的是，人们实际上对基因表达的持续变化知之甚少，这些变化可能是 阿片类药物和其他滥用药物导致的脑多巴胺系统功能障碍。 我们的项目围绕三个具体目标展开。在目标1中，我们将从免疫标记中提取染色质 150例正常人和150例正常人中脑多巴胺能神经元核团的荧光分选 诊断为阿片类药物滥用的病例，然后在全基因组范围内描述转录组和开放 染色质景观和启动子-增强子环以及其他类型的染色体构象( ‘3D基因组’)以特定细胞类型的方式。在目标2中，我们将应用综合基因组学方法和 利用Aim#1尸检脑数据和由提供的人群规模的基因型别和表型 百万退伍军人计划和精神病学基因组学联盟建立因果概率网络和 预测多巴胺能系统的调控非编码DNA空间内的关键驱动因素。在目标3中，我们将 用小RNA引导的表观基因组验证成瘾相关的顺式调控序列(来自目标#1、#2) 培养的人多巴胺能神经元的编辑系统。总的来说，我们的中脑多巴胺能神经元- 重点项目将填补人类成瘾研究和人类神经基因组学领域的关键空白 首次开始对奖励和治疗中的关键细胞群之一进行深入的表观基因组学评估 成瘾回路。

Although many cells and neural circuits clearly contribute to opiate and other substance abuse disorder, the path to drug addiction travels through midbrain dopaminergic neurons. Though a rare cell type (it is estimated that a mere 1 of every 200,000 neurons in the human brain is of a dopaminergic phenotype), changes in dopaminergic neurotransmission are thought to play a role in various stages of addiction, from acute reward mechanisms and goal-directed actions, to the development of habitual behavior and increased salience of cues associated with drug use, as well as the anhedonia and dysphoria associated with drug withdrawal. Surprisingly little is actually known about persistent changes in gene expression that presumably underlie the dysfunction of dopamine systems in brain exposed to opiates and other drug of abuse. Our project is centered on three Specific Aims. In Aim #1,we will extract chromatin from immunotagged midbrain dopaminergic neuron nuclei collected by fluorescence-activated sorting from 150 controls and 150 cases diagnosed with opiate abuse and then profile, on a genome-wide scale, the transcriptome and open chromatin landscapes and promoter-enhancer loopings and other types of chromosomal conformations (the ‘3D genome’) in cell type-specific manner. In Aim #2, we will apply integrative genomics approaches and leverage Aim #1 postmortem brain data with population-scale genotypes and phenotypes provided by the Million Veterans Project and the Psychiatric Genomics Consortium to build causal probabilistic networks and predict key drivers within the regulatory non-coding DNA space of the dopaminergic system. In Aim #3, we will validate addiction-relevant cis-regulatory sequences (from Aim #1, #2) with small RNA-guided epigenomic editing systems in cultured human dopaminergic neurons. Collectively, our midbrain dopaminergic neuron- focused project will fill critical voids in the field of human addiction research and human neurogenomics and embark, for the first time, on a deep epigenomic assessment of one of the key cell populations in reward and addiction circuitry.