Gene Regulation in the Opioid Dependent Human Brain (Project 2)

阿片类药物依赖性人脑的基因调控（项目 2）

• 批准号: 10493706
• 负责人: Peter Christopher Scacheri
• 金额: $ 58.91万
• 依托单位: RESEARCH TRIANGLE INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-15 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10493706
• 关键词: Acute; Addictive Behavior; Address; Amygdaloid structure; Architecture; Autopsy; Basal Ganglia; Behavior; Behavioral; Biological; Brain; Brain region; Cells; ChIP-seq; Chronic; Collection; Complex; DNA Methylation; Data; Data Analyses; Data Set; Dependence; Development; Dimensions; Dorsal; Environmental Risk Factor; Fluorescence-Activated Cell Sorting; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Engineering; Genome; Genomics; Genotype; Goals; Histone Acetylation; Human; Impulsivity; Individual; Injections; Institutes; Intervention; Intoxication; Lateral; Link; Maps; Measurement; Meta-Analysis; Methylation; Motivation; Mus; Neurobiology; Nucleus Accumbens; Opiate Addiction; Opioid; Pathway interactions; Pharmacology; Prefrontal Cortex; Public Health; Quantitative Trait Loci; RNA; Regulator Genes; Regulatory Element; Resources; Rewards; SET gene; Sampling; Single Nucleotide Polymorphism; Site; Stress; Substance Use Disorder; Substance abuse problem; Surveys; Testing; Time; Tissues; Transcript; Validation; Variant; Viral Vector; Withdrawal; Work; addiction; brain tissue; case control; cell type; cohort; compulsion; drug discovery; epigenome; executive function; follow-up; genetic association; genetic variant; genome wide association study; genome-wide; gun homicide; histone modification; human tissue; in vivo; incentive salience; multiple omics; negative affect; novel; novel therapeutics; opioid abuse; opioid epidemic; opioid exposure; opioid mortality; opioid overdose; opioid use; overdose death; relating to nervous system; single cell analysis; single-cell RNA sequencing; synergism; targeted treatment; theories; transcriptome; transcriptome sequencing

PROJECT SUMMARY/ABSTRACT The ongoing opioid epidemic is one of the most important public health crises of our time with overdose fatalities quadrupling since 1999 and surpassing those of gun homicides in recent years. Given the individual and societal burden of opioid dependence, it is necessary to characterize the biological pathways to dependence that can be targeted for treatment and intervention. The work proposed in Project 2 combines multi-omics resources to examine specific hypotheses regarding the functional brain architecture of opioid addiction (OA). Guided by existing theory implicating the prefrontal cortex (PFC), nucleus accumbens (NAc), and amygdala in the cycle of substance abuse, we propose to generate and combine existing data at multiple omics levels in each brain region using postmortem human tissue collected from opioid overdose cases and matched controls. The measurement approaches we use are genome-wide, in each of the three brain regions and in each biological domain: single nucleotide polymorphism (SNP), epigenome (DNA methylation and histone acetylation), and transcript (via RNA-seq) in our combined sample (N=641) with follow-up single-cell validation/replication. We will accomplish these goals across three aims. In Aim 1, in the existing combined sample, we will fill out the brain regions not currently assessed in specific brain/regions or biological domains and perform analyses to detect region-specific and general (across-brain regions) methylation, histone acetylation, and gene expression differences between cases and controls. In addition, we will perform analyses aimed at integrating across those biological domains. In Aim 2, we will perform single-cell follow-up of Aim 1 findings to pinpoint all cell types within the PFC, NAc, and amygdala that drive the differences identified in Aim 1, with validation by RNA-seq in fluorescence activated cell sorting (FACS) separated cells and RNA-FISH. In Aim 3, we will genotype samples where needed and perform genetic association mapping—histone modification, DNA methylation, and gene expression QTL mapping—to identify genetic variants that account for the regulatory differences observed between cases and controls in Aim 1. These QTL approaches will allow us to identify regulatory differences with a genetic basis, versus those somatically acquired as a consequence of chronic opiate use, exposure, or other environmental factors. Our overall goal is to create a framework in which we can map, in brain, the OA functional relevance of genomic sets (genes, pathways, features) and generate results that can be leveraged in Project 1 and the Synergy Core through mapOA, and in Project 3 through selected genes, targeted for functional validation in vivo in the dlPFC and NAc using genetic engineering, viral vector injection, and pharmacology approaches in mice. Through this synergy we will distinguish between predisposing and exposure consequent dysregulation and move toward mechanistic understanding that can drive new treatments of OA.

项目总结/摘要 持续的阿片类药物流行病是我们这个时代最重要的公共卫生危机之一， 自1999年以来，死亡人数翻了两番，近年来超过了枪支凶杀案。鉴于个人 和阿片类药物依赖的社会负担，有必要描述生物学途径， 依赖，可以有针对性的治疗和干预。项目2中提出的工作结合了 多组学资源，以检查有关阿片类药物功能性大脑结构的特定假设 成瘾（OA）。在现有理论的指导下，涉及前额叶皮层（PFC），丘脑核（NAc）， 和杏仁核在药物滥用的循环，我们建议产生和联合收割机现有的数据， 使用从阿片类药物过量收集的死后人体组织，在每个大脑区域中进行多组学水平分析 病例和配对对照。我们使用的测量方法是全基因组的，在每个基因组中， 三个大脑区域和每个生物学领域：单核苷酸多态性（SNP），表观基因组（DNA 甲基化和组蛋白乙酰化）和转录物（通过RNA-seq）在我们的合并样品（N=641）中， 后续单细胞验证/复制。 我们将通过三个目标实现这些目标。在目标1中，在现有的组合样本中，我们将填充 排除目前在特定大脑/区域或生物学领域中未评估的大脑区域， 分析以检测区域特异性和一般（跨脑区域）甲基化、组蛋白乙酰化，以及 病例组和对照组之间的基因表达差异。此外，我们还将进行分析， 整合这些生物领域。在目标2中，我们将对目标1的发现进行单细胞随访 确定PFC，NAc和杏仁核中驱动Aim 1中确定的差异的所有细胞类型， 通过荧光激活细胞分选（FACS）中的RNA-seq分离的细胞和RNA-FISH进行验证。在Aim中 3、我们将在需要的地方对样本进行基因分型，并进行遗传关联图谱-组蛋白修饰， DNA甲基化和基因表达QTL定位-以确定解释基因突变的遗传变异。 在目标1中观察到病例和对照之间的监管差异。这些QTL方法可以让我们 以确定具有遗传基础的调节差异，与那些体细胞获得的结果相比， 慢性阿片类药物使用，暴露或其他环境因素。 我们的总体目标是创建一个框架，在这个框架中，我们可以在大脑中绘制OA功能相关性， 基因组集（基因、途径、特征），并生成可在项目1和 协同核心通过mapOA，并在项目3中通过选定的基因，针对功能验证， 体内dlPFC和NAc使用基因工程，病毒载体注射，和药理学方法， 小鼠通过这种协同作用，我们将区分诱发和暴露后果 调节失调，并朝着机制的理解，可以推动新的治疗OA。