Deciphering the neural basis of alcohol use disorders using human and mouse neurons

利用人类和小鼠神经元破译酒精使用障碍的神经基础

• 批准号: 9217533
• 负责人: ZHIPING P. PANG
• 金额: $ 35.21万
• 依托单位: RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-02-05 至 2021-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9217533
• 关键词: Acute; Address; Affect; Agonist; Alcohol abuse; Alcoholism; Alcohols; Amino Acid Substitution; Asparagine; Aspartate; Behavior; Biochemical; Biochemistry; Bioinformatics; Cell physiology; Chronic; Consumption; Data; Defect; Development; Enkephalins; Ethanol; Ethnic group; Genes; Genetic study; Goals; Health; High Prevalence; Human; Human Genetics; Impairment; Individual; Label; Mediating; Methods; Microfabrication; Microfluidics; Midbrain structure; Modeling; Molecular; Mus; N-Glycosylation Site; Naltrexone; Neuraxis; Neurobiology; Neurons; Opioid; Opioid Receptor; Physiology; Receptor Activation; Receptor Signaling; Regulation; Research; Rewards; Signal Transduction; Single Nucleotide Polymorphism; Slice; Specific qualifier value; Stem cells; Synapses; Synaptic Transmission; System; Technology; Testing; Therapeutic Effect; Variant; Ventral Tegmental Area; addiction; alcohol effect; alcohol sensitivity; alcohol use disorder; cell type; design; dopamine system; dopaminergic neuron; drinking behavior; effective therapy; endogenous opioids; genetic variant; innovation; insight; mouse model; mu opioid receptors; novel; problem drinker; protein transport; public health relevance; receptor function; receptor internalization; relating to nervous system; socioeconomics; stem cell biology; synaptic function; trafficking

 DESCRIPTION (provided by applicant): Alcoholism is a serious health and socioeconomic problem in the U.S. Understanding how alcohol produces reward, motivates further consumption, and eventually leads to addiction is necessary to design effective treatments for alcohol use disorders (AUDs). Synaptic transmission mediates all of these behaviors, however, little is known about the effect of alcohol on synaptic transmission in the context of human neurons. The single nucleotide polymorphism (SNP) rs1799971 (OPRM1 A118G) produces a non-synonymous amino acid substitution in the mu-opioid receptor (MOR), in which Asparagine 40 (MOR N40) is replaced with Aspartate (MOR D40), and is associated with AUDs in specific ethnic groups. Importantly, Naltrexone, a nonselective MOR antagonist, has potent therapeutic effects in alcoholic individuals with MOR D40. We generated human neurons from 7 subjects carrying either homozygous MOR N40 or MOR D40. Our preliminary data suggest that human neurons carrying D40 show defective re-sensitization after MOR activation by DAMGO ([D-Ala2, NMe-Phe4, Gly-ol5]-enkephalin), suggesting defective trafficking of MORs. Supporting this, bioinformatic analyses and mouse models of human MOR N40D suggest that D40 disrupts an N-glycosylation site on MOR. However, the mechanism by which MOR protein trafficking defects affect the interaction between ethanol and opioids is not known. It is particularly important to reveal the molecular mechanisms underlying the function of N40D MOR variants in their native neuronal context because previous studies performed in heterologous systems have revealed inconclusive and confusing results. Moreover, a species-specific trafficking mechanism of MORs has been suggested. The objective of this proposal is to study the impact of alcohol and opioid signaling in both mouse and human neurons carrying both the N40 and D40 MOR allelic variants, focusing on the synaptic mechanisms that likely underlie behavior. The central hypothesis is that defective D40 MOR trafficking results in an altered effect of the interaction between alcohol and opioids on synaptic function in the reward neurocircuitry. We will first examine the effect of alcohol on synaptic function in a defined neurocircuitry composed of human neurons carrying these gene variants. Next, we will use a mouse model of human N40D, and study the synaptic mechanism in the reward neurocircuitry, i.e. the midbrain ventral tegmental area (VTA), in relation to MOR function and ethanol. The proposed research is innovative, because we will combine recent developments in stem cell biology, the state-of-the-art synaptic physiology, and novel microfabrication technologies to directly probe the impact of alcohol and opioid signaling on synaptic function in both mouse and human neuronal networks carrying OPRM1 gene variants. We expect to unravel a species- and cell type-specific mechanism of MOR N40D variants that may provide novel information for understanding AUDs.

 描述（由申请人提供）：酒精中毒是一个严重的健康和社会经济问题在美国了解酒精如何产生奖励，激励进一步消费，并最终导致成瘾是必要的，以设计有效的治疗酒精使用障碍（AUDs）。突触传递介导了所有这些行为，然而，关于酒精对人类神经元突触传递的影响知之甚少。单核苷酸多态性（SNP）rs 1799971（OPRM 1 A118 G）在μ阿片受体（莫尔）中产生非同义氨基酸取代，其中天冬酰胺40（莫尔N40）被天冬氨酸（莫尔D40）取代，并与特定种族群体中的AUD相关。重要的是，纳洛酮，一种非选择性莫尔拮抗剂，在患有莫尔D40的酒精个体中具有有效的治疗作用。我们从7名携带纯合莫尔N40或莫尔D40的受试者中产生了人神经元。我们的初步数据表明，携带D40的人神经元在DAMGO（[D-Ala 2，NMe-Phe 4，Gly-ol 5]-脑啡肽）激活莫尔后显示出缺陷的再敏化，表明MOR的缺陷运输。支持这一点的是，人莫尔N40 D的生物信息学分析和小鼠模型表明，D40破坏了莫尔上的N-糖基化位点。然而，莫尔蛋白运输缺陷影响乙醇和阿片类药物之间相互作用的机制尚不清楚。揭示N40 D莫尔变体在其天然神经元环境中的功能的分子机制是特别重要的，因为先前在异源系统中进行的研究揭示了不确定和混乱的结果。此外，MORs的物种特异性运输机制已被提出。本提案的目的是研究酒精和阿片类药物信号传导在携带N40和D40莫尔等位基因变体的小鼠和人类神经元中的影响，重点关注可能构成行为基础的突触机制。核心假设是，缺陷的D40莫尔运输导致酒精和阿片类药物之间的相互作用对奖赏神经回路中突触功能的影响改变。我们将首先研究酒精对由携带这些基因变体的人类神经元组成的确定的神经回路中突触功能的影响。接下来，我们将使用人类N40 D的小鼠模型，并研究与莫尔功能和乙醇相关的奖赏神经回路（即中脑腹侧被盖区（VTA））中的突触机制。拟议的研究是创新的，因为我们将结合联合收割机在干细胞生物学，最先进的突触生理学和新的微加工技术的最新发展，直接探测酒精和阿片类药物信号传导对突触功能的影响在小鼠和人类神经网络携带OPRM 1基因变异。我们希望能够阐明莫尔N40 D变异的物种和细胞类型特异性机制，这可能为理解AUDs提供新的信息。