Role of Cell Type-Specific Molecular Rhythm Disruption in Alcohol Use Disorder

项目摘要/摘要 饮酒是200多种健康状况的原因之一，据估计占全球负担的5.1% 疾病和伤害。尽管对公众健康造成了巨大的影响，但我们仍然缺乏对 导致酒精使用障碍(AUD)的机制。AUD的一个显著特征是昼夜节律 紊乱，包括生理节律和睡眠/清醒周期的改变。这种关系是双向的，因为 节律紊乱会加剧饮酒和正常昼夜节律基因的多态性 与酒精使用量的增加有关。然而，人们对这些现象背后的机制知之甚少。 关系，特别是在分子水平上，患有澳门氏症的个体。分子节律是 由转录-翻译反馈环产生，以控制昼夜节律依赖(近24小时)的基因 表情。值得注意的是，AUD与正常昼夜节律基因的分子节律紊乱有关。 外周和啮齿动物的研究表明，纹状体分子节律的破坏，特别是 伏隔核(NAC)与奖赏调节的改变和物质使用风险的增加有关。 然而，人类死后大脑分子节律的测量从历史上来说一直是具有挑战性的 每个大脑代表一个昼夜节律的时间点。一项创新的分析使用了死亡时间(TOD)来匹配 通过组合来自所有受试者的表达数据，有可能重建分子 人类死后大脑的节律模式。为了研究与AUD相关的分子节律变化， 我们进行了一项初步研究，检测了受试者大量nac组织中的大规模转录变化。 将澳元与对照组进行比较。值得注意的是，核心昼夜节律基因(例如，CRY1、PER2)显示出节律性 在对照组中的表达，在AUD中是节律性的，表明分子与行为节律有关 在AUD患者中观察到的变化。然而，NAC是由各种转录上不同的细胞组成的 在饮酒行为中扮演不同角色的类型，以及分子节律在这些细胞类型中的作用 之前没有被调查过。这个R21提议的中心假设是分子节律是 在AUD受试者的NAC中以细胞类型特定的方式被干扰，并且这些干扰有助于 过度饮酒。为了验证这一假设，我们将使用人类的单核rna测序(snrna-seq)。 死后NAC组织以确定AUD异质细胞群体中的分子节律如何改变 (目标1)。然后，我们将操纵特定NAC细胞类型的节律，以确定节律的因果作用 饮酒中断(目标2)。总之，这些研究将是第一个检查特定细胞类型的研究 人类纹状体的节律，它们在AUD中是如何改变的，以及它们对过度饮酒的贡献。 这些研究对于理解澳门氏症昼夜节律紊乱的机制至关重要，并可能 导致发现新的治疗靶点和/或未来治疗的依赖于时间的策略。

PROJECT SUMMARY/ABSTRACT Alcohol use is a causal factor in over 200 health conditions and is estimated to account for 5.1% of global burden of disease and injury. Despite the enormous public health impact, we still lack a basic understanding of the mechanisms that contribute to alcohol use disorder (AUD). A prominent feature of AUD is circadian rhythm disturbances, including altered physiological rhythms and sleep/wake cycles. The relationship is bidirectional, as rhythm disruptions can exacerbate alcohol consumption and polymorphisms in canonical circadian genes associate with increased alcohol use. However, very little is known about the mechanisms underlying these relationships, especially at the molecular level in the brains of individuals with AUD. Molecular rhythms are generated by transcriptional-translational feedback loops to control circadian-dependent (near 24-hour) gene expression. Notably, AUD is associated with disrupted molecular rhythms of canonical circadian genes in the periphery and rodent studies have shown that disruptions to molecular rhythms in the striatum, particularly the nucleus accumbens (NAc), are associated with altered reward regulation and increased risk for substance use. Measures of molecular rhythms in human postmortem brain, however, have historically been challenging since each brain represents a single circadian timepoint. An innovative analysis uses “time of death” (TOD) to fit subjects on a 24-hr ‘clock’; by combining expression data from all subjects, it is possible to reconstruct molecular rhythm patterns in the human postmortem brain. To investigate AUD-associated molecular rhythm alterations, we performed a preliminary study examining large-scale transcriptional changes in bulk NAc tissue from subjects with AUD compared to controls. Notably, core circadian genes (e.g., CRY1, PER2), which display rhythmic expression in control subjects, were arhythmic in AUD, suggesting a molecular link to behavioral rhythm alterations observed in AUD patients. However, the NAc is composed of a variety of transcriptionally distinct cell types with differential roles in alcohol drinking behavior, and the role of molecular rhythms in these cell types has not been previously investigated. The central hypothesis of this R21 proposal is that molecular rhythms are disrupted in a cell-type specific manner in the NAc of AUD subjects, and that these disruptions contribute to excessive drinking. To test this hypothesis, we will use single nucleus RNA-sequencing (snRNA-seq) of human postmortem NAc tissue to determine how molecular rhythms are altered in heterogenous cell populations in AUD (Aim 1). We will then manipulate rhythms in specific NAc cell types to determine a causal role for rhythm disruption in alcohol drinking (Aim 2). Together, these studies will be the first to examine cell type-specific rhythms in human striatum, how they are altered in AUD, and their contribution to excessive alcohol consumption. These studies are central to understanding the mechanisms underlying circadian disruptions in AUD and may result in the discovery of novel therapeutic targets and/or time-dependent strategies for future treatments.