Translational examination of alcohol-associated epigenetic signatures: from primates to rodents

酒精相关表观遗传特征的转化检查：从灵长类动物到啮齿类动物

基本信息

批准号：
10056068
负责人：
Verginia Carmella Cuzon Carlson
金额：
$ 23.52万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1996
资助国家：
美国
起止时间：
1996-12-01 至 2025-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10056068
关键词：
Age Alcohol abuse Alcohol consumption Alcoholic beverage heavy drinker Alcohols American Anatomy Anterior Antibodies Astrocytes Behavioral Biological Assay Brain region Brodmann&apos s area Cause of Death Cells Consumption Corpus striatum structure Cytosine DNA Methylation DNA methylation profiling Data Data Set Development Dorsal Dose Electrophysiology (science)Elements Engineering Epigenetic Process Equilibrium Ethanol Extracellular Matrix FDA approved Female Fluorescence-Activated Cell Sorting Gene Expression Gene Targeting Genes Genomic Segment Glutamates Goals Heavy Drinking Individual Lead Link Macaca Macaca mulatta Maps Methylation Modeling Modification Molecular Monkeys Mus Neurons Nucleic Acid Regulatory Sequences Nucleus Accumbens Pathway Analysis Pharmaceutical Preparations Pharmacology Play Primates Process Regulation Regulator Genes Relapse Rodent Role Sampling Self Administration Signal Transduction Slice Specificity Synapses Technology Testing Tissues Validation Viral Viral Vector Work alcohol effect alcohol response alcohol use disorder annotation system base behavioral genomics bisulfite sequencing brain tissue cell type chronic alcohol ingestion cingulate cortex design drinking behavior effective therapy epigenome epigenomics genetic manipulation genome-wide male methylome mouse genome neural circuit neuroadaptation neurotransmission nonhuman primate novel postsynaptic preference synaptic function tool transcriptome sequencing transcriptomics translational approach transmission process

项目摘要

PROJECT ABSTRACT Although approximately 54% of Americans over the age of 18 consume alcohol, only 6.5% meet the criteria for an alcohol use disorder (AUD). To date, there are three FDA approved medications to treat AUD, each having varying efficacy on an individual basis. Understanding the molecular mechanisms underlying circuit-specific changes that lead to AUD is essential to developing novel, targeted, and effective treatments for AUD. In advancement of this goal, we have begun to identify genome-wide DNA methylation (DNAm) signals within the nucleus accumbens core (NAcC) that distinguish low and heavy ethanol drinking monkeys. A subset of these differential DNAm (D-DNAm) signals were associated with the expression of genes that play a role in modulating neurotransmission. In particular, we found D-DNAm signals in genes that are functionally associated with different compartments of the tetrapartite synapse that includes pre- and postsynaptic elements, astroglial processes, and the extracellular matrix. Glutamatergic inputs into the NAcC arise from brain regions outside of the NAcC, including Brodmann area 32 (A32, in primates; prelimbic cortex (PL) in rodents). By investigating A32 inputs into NAcC, we will begin to elucidate circuit-specific DNAm signals that distinguish low from heavy/very heavy ethanol drinkers. We will use to advantage the highly relevant and well- characterized nonhuman primate (NHP) alcohol drinking model in which rhesus macaques imbibe alcohol daily for over 12 months and self-select into low and heavy drinkers. Genome-wide DNAm sequencing (GW-DNAm) will be used to identify alcohol-dose associated differentially methylated cytosines (DMCs) and regions (DMRs) in the A32 and NAcC. Using a combination of a neuron specific antibody (NeuN) with a fluorescent tag and fluorescence-activated cell sorting, we will begin to elucidate cell specificity (i.e. neuron-specific) in the DNAm signals of the NHP A32. In parallel, we will examine the DNAm state of these same NHP A32 targets using amplicon bisulfite sequencing in PL in high-ethanol preference mice that have consumed alcohol for 3 months and self-select into low and heavy drinkers. These studies will yield gene/regulatory regions as targets that are highly correlated with ethanol dose, based on differences in consumption, conserved across species and play a role in the tetrapartite synapse. Using engineered viral vectors that alter the expression, function or methylation level of gene targets, we will perform a mouse functional assay that will test their role in ethanol drinking (self-administration of ethanol for 3 months) and in the PL-NAcC circuit (tested using ex vivo slice electrophysiology). The most efficacious targets from the mouse functional assay will be tested in ex vivo slices obtained from chronic ethanol drinking NHPs for their ability to alter A32 and NAcC circuitry. In total, this work will identify alcohol dose-dependent DNAm modifications that are specific to the A32/PL-NAcC circuit, and provide functional support for the design of promising new AUD treatments.

项目摘要尽管大约 54% 18 岁以上的美国人饮酒，但只有 6.5% 的人符合饮酒标准酒精使用障碍（AUD）。迄今为止，FDA 批准了三种治疗 AUD 的药物，每种药物都具有功效因人而异。了解特定电路的分子机制导致 AUD 的变化对于开发新颖、有针对性且有效的 AUD 治疗方法至关重要。在为了实现这一目标，我们已经开始识别基因组范围内的 DNA 甲基化 (DNAm) 信号伏隔核核心（NAcC）区分低度和大量饮酒的猴子。其中的一个子集差异 DNAm (D-DNAm) 信号与基因表达相关，这些基因在调节神经传递。特别是，我们在具有功能性的基因中发现了 D-DNAm 信号与四方突触的不同区室相关，包括突触前和突触后元素、星形胶质细胞突起和细胞外基质。 NAcC 的谷氨酸输入来自 NAcC 以外的大脑区域，包括灵长类动物的布罗德曼区 32（A32；灵长类动物的前边缘皮层 (PL)）啮齿类动物）。通过研究 NAcC 的 A32 输入，我们将开始阐明电路特定的 DNAm 信号区分低度和重度/重度乙醇饮用者。我们将利用高度相关且良好的优势恒河猴每天饮用酒精的非人类灵长类动物 (NHP) 饮酒模型超过 12 个月，并自我选择为低度饮酒者和重度饮酒者。全基因组 DNAm 测序 (GW-DNAm) 将用于识别与酒精剂量相关的差异甲基化胞嘧啶（DMC）和区域（DMR）在 A32 和 NAcC 中。结合使用神经元特异性抗体 (NeuN) 和荧光标签荧光激活细胞分选，我们将开始阐明 DNAm 中的细胞特异性（即神经元特异性） NHP A32 的信号。同时，我们将使用以下方法检查这些相同 NHP A32 靶标的 DNAm 状态：对饮酒 3 个月的高乙醇偏好小鼠 PL 进行亚硫酸氢盐扩增子测序并自我选择饮酒量低的人和重度饮酒的人。这些研究将产生基因/调控区域作为目标与乙醇剂量高度相关，基于消耗的差异，在物种和游戏中保守在四方突触中发挥作用。使用工程化病毒载体改变表达、功能或基因靶标的甲基化水平，我们将进行小鼠功能测定，测试它们在乙醇中的作用饮酒（自我施用乙醇 3 个月）和 PL-NAcC 回路（使用离体切片进行测试）电生理学）。小鼠功能测定中最有效的靶标将在离体切片中进行测试从长期饮用乙醇的 NHP 中获得，因为它们能够改变 A32 和 NAcC 电路。这部作品总共将识别 A32/PL-NAcC 电路特有的酒精剂量依赖性 DNAm 修饰，以及为有前景的新 AUD 治疗方法的设计提供功能支持。