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

虽然许多细胞和神经回路显然有助于阿片类药物和其他物质滥用障碍， 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（估计 人类大脑中每20万个神经元中只有1个是多巴胺能表型）， 多巴胺能神经传递被认为在成瘾的各个阶段（从急性奖赏）发挥作用 机制和目标导向的行动，习惯行为的发展和增加的突出性， 与药物使用相关的线索，以及与药物戒断相关的快感缺乏和烦躁不安。 令人惊讶的是，人们对基因表达的持续变化知之甚少，而这种变化可能是基因表达的基础。 暴露于阿片类药物和其他滥用药物大脑中多巴胺系统功能障碍。 我们的项目围绕三个具体目标。在目标#1中，我们将从免疫标记的 通过荧光激活分选从150名对照和150名 诊断为阿片类药物滥用的病例，然后在全基因组范围内分析转录组和开放 染色质景观和启动子-增强子环和其他类型的染色体构象（ “3D基因组”）。在目标#2中，我们将应用整合基因组学方法， 利用Aim #1死后大脑数据与人口规模的基因型和表型提供的 百万退伍军人项目和精神病基因组学联盟建立因果概率网络， 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指导的表观基因组验证成瘾相关的顺式调节序列（来自目标#1，#2） 在培养的人类多巴胺能神经元中的编辑系统。我们的中脑多巴胺能神经元- 该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， 第一次开始对奖励中的关键细胞群之一进行深入的表观基因组评估， 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.