Identifying new targets for the treatment of alcohol dependence and relapse: epigenetic analysis of the abstinent brain

总结 酒精使用障碍（AUD）影响超过12%的美国人口，是第四大可预防的疾病。 美国的死因。AUD与强迫性饮酒和负面情绪的出现有关 在戒酒期间，三分之二的受影响的人在完成饮酒后的几个月内复发 停止治疗。反复的中毒和禁欲循环与大脑中的持续变化有关。 大脑在解毒后很长时间内仍然存在，这会导致饮酒的复发，通常比以前更高 禁欲确定导致危险酒精使用的分子机制， 复发率对于有效设计更好的治疗方法以遏制酗酒至关重要。我们以前的 研究说明了整合表观遗传学、转录组学、电路、药理学和行为工具的力量 以阐明导致危险饮酒的分子基础。这些研究，使用基因组- 广泛的DNA甲基化（DNAm）分析的核延髓（NAc），以比较猕猴酒精幼稚 和饮酒者，确定了差异DNAm（D-DNAm）信号，映射到以前没有关联的基因， 酒精的使用，但与突触可塑性调制高度相关。对其中两个的实验操作 DNAm相关基因导致谷氨酸和GABA神经传递的改变以及乙醇摄入量的变化。 下一个关键步骤是了解重复的戒断/复发周期如何改变DNA信号，以及如何 这些变化有助于神经回路的适应。我们假设一些D-DNAm信号 与长期饮酒有关的慢性酒精中毒可能在戒酒期间持续存在，增加复发的风险。此外，de 在重复的戒断/复发周期中产生的新生D-DNAm信号，可能进一步增加复发 风险为了识别禁欲相关的DNA，我们将使用高度翻译的猕猴酒精自我- 管理模式和我们已经证明的全基因组方法来识别猕猴NAc中的D-DNAm 在> 12个月的慢性酒精自我给药后，然后进行3个周期的强制戒酒（每个周期： 1个月的禁欲期和3个月的开放期）。在整合基因和转录变体表达后， 从相同的受试者和组织中产生的D-DNAm数据，一个引人注目的新靶点的子集， 选择用于在小鼠慢性中使用药理学和遗传操作方法进行功能研究， 间歇乙醇（CIE）模型。这种啮齿动物CIE模型已被广泛用于模拟酒精循环 暴露和戒断导致复发期间饮酒升级。除了记录乙醇 摄入后，将使用膜片钳评估靶点操作对神经传递的影响 电生理分析总的来说，这些研究将有助于我们了解分子机制（S） 确定复发的风险。这些信息对于促进有效疗法的发展至关重要 防止酗酒和复发

SUMMARY Alcohol use disorder (AUD) affects more than 12% of the US population and is the fourth leading preventable cause of death in the US. AUD is associated with compulsive drinking and emergence of a negative emotional state during abstinence, resulting in two-thirds of afflicted people relapsing within months of completing alcohol cessation treatment. Repeated cycles of intoxication and abstinence are linked to persistent alterations in the brain that remain long after detoxification, which drive relapse of drinking, often at levels higher than before abstinence. Identifying the molecular mechanisms that lead to hazardous alcohol use and contribute to the high rates of relapse is imperative for the effective design of better treatments to curb alcohol abuse. Our previous studies illustrate the power of integrating epigenetic, transcriptomic, circuitry, pharmacology and behavioral tools to elucidate the molecular underpinnings contributing to hazardous alcohol use. Those studies, using genome- wide DNA methylation (DNAm) analysis of the nucleus accumbens (NAc) to compare macaque alcohol-naive and alcohol-drinkers, identified differential DNAm (D-DNAm) signals, mapping to genes not previously linked to alcohol use, but with high relevance to synaptic plasticity modulation. Experimental manipulation of two of these DNAm-linked genes resulted in altered glutamate and GABA neurotransmission and changes in ethanol intake. The the next critical step is to understand how repeated abstinence/relapse cycles alters DNAm signals and how these changes contribute to adaptations in neurocircuitry. We hypothesize that some D-DNAm signals associated with chronic alcohol use may persist during abstinence, contributing to risk of relapse. In addition, de novo D-DNAm signals generated during repeated cycles of abstinence/relapse, may further heighten relapse risk. To identify abstinence-associated DNAm, we will use the highly translational macaque alcohol self- administration model and our proven genome-wide approach to identify D-DNAm in the NAc of macaques following > 12 months of chronic alcohol self-administration and then 3 cycles of forced abstinence (each cycle: 1 month abstinence and 3 months of open-access). After integrating gene and transcript variant expression and D-DNAm data generated from the same subjects and tissues, a subset of compelling, novel targets will be selected for functional study using pharmacological and genetic manipulation approaches in a mouse chronic intermittent ethanol (CIE) model. This rodent CIE model has been extensively used to model cycles of alcohol exposure and withdrawal leading to an escalation of drinking during relapse. In addition to recording ethanol intake, the effects of target manipulation on neurotransmission will be evaluated using patch-clamp electrophysiological analysis. Overall, these studies will aid in our understanding of the molecular mechanism(s) establishing risk for relapse. This information will be critical to advancing the development of effective therapies for alcohol abuse and to prevent relapse.