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

摘要 酒精使用障碍(AUD)影响着超过12%的美国人口，是第四大可预防的疾病 在美国的死因。AUD与强迫性饮酒和出现负面情绪有关 在戒酒期间处于戒酒状态，导致三分之二的患者在戒酒后几个月内复发 戒烟治疗。反复的醉酒和禁欲的循环与持续的改变有关 在戒毒后保持很长时间的大脑，这会导致饮酒复发，通常水平比以前高 禁欲。确定导致危险饮酒并导致高饮酒率的分子机制 对于有效设计更好的治疗方法来遏制酗酒，复发率至关重要。我们以前的 研究表明，整合表观遗传学、转录、电路、药理学和行为工具的力量 阐明危险饮酒的分子基础。这些研究，使用基因组- 伏隔核(NAC)广谱DNA甲基化(DNaM)分析比较猕猴酒精-幼稚 和饮酒者，识别出差异dNaM(D-dNaM)信号，映射到以前没有关联的基因 酒精使用，但与突触可塑性调节高度相关。其中两个的实验操作 DNaM相关基因导致谷氨酸和GABA神经传递改变，酒精摄入量改变。 下一个关键步骤是了解反复戒酒/复发周期如何改变dNaM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 在戒酒期间，与长期饮酒有关的因素可能会持续存在，从而增加复发的风险。此外，De 在反复戒断/复发周期中产生的Novo D-dNaM信号可能会进一步加剧复发 风险。为了识别与戒酒相关的dNaM，我们将使用高度翻译的猕猴酒精自身 给药模式和我们成熟的全基因组方法在猕猴NAC中识别D-dNaM 经过12个月的长期自我饮酒，然后进行3个周期的强制戒酒(每个周期： 禁食1个月，开放获取3个月)。在整合了基因和转录本的变体表达后 从相同的受试者和组织产生的D-dNaM数据，引人注目的新目标的子集将是 用药理学和遗传操作方法选择用于慢性肾衰小鼠的功能研究 间歇乙醇(CIE)模型。这种啮齿动物CIE模型已被广泛用于酒精循环的模拟 接触和戒断会导致复发期间饮酒的升级。除了记录乙醇 摄取，靶向操作对神经传递的影响将使用膜片钳进行评估 电生理分析。总的来说，这些研究将有助于我们理解分子机制(S) 确定复发的风险。这些信息将对推动有效疗法的发展至关重要 治疗酗酒和防止复发。

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.