Neuronal basis of OPRM1 A118G polymorphism in alcohol use disorders

酒精使用障碍中 OPRM1 A118G 多态性的神经元基础

基本信息

批准号：
9146155
负责人：
Apoorva Halikere
金额：
$ 3.35万
依托单位：
RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2018-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9146155
关键词：
Acute Address Adenine Affect Affinity Agonist Alcohol abuse Alcohol consumption Alcoholism Alcohols Alleles Amino Acid Substitution Animals Asparagine Aspartate Behavior Biochemical Bioinformatics Brain imaging Cell Line Cells Chronic D Aspartate Data Dependence Development Dopamine Dose Economics Electrophysiology (science)Enkephalins Environmental Risk Factor Ethnic Origin Ethnic group Etiology European Exhibits Genes Genetic Polymorphism Genetic Variation Genetic study Genotype Goals Guanine Heritability Human Immunohistochemistry Individual Intoxication Life Ligand Binding Ligands Mediating Molecular Morphology N-Glycosylation Site Naltrexone Neurons Nicotine Dependence Opioid Opioid Receptor Patients Pattern Physiological Positioning Attribute Predisposition Property Receptor Activation Receptor Signaling Recycling Regulation Reporting Research Risk Signal Transduction Single Nucleotide Polymorphism Societies Synapses Synaptic Transmission System Techniques Technology Testing Therapeutic Effect Western Blotting Work alcohol exposure alcohol reinforcement alcohol response alcohol reward alcohol sensitivity alcohol use disorder base brain tissue effective therapy excitatory neuron genetic variant glycosylation induced pluripotent stem cell inhibitory neuron innovation insight mu opioid receptors nervous system disorder novel presynaptic problem drinker public health relevance receptor receptor expression receptor function relating to nervous system research study social stem cell biology synaptic function synaptic inhibition synaptogenesis trafficking

项目摘要

DESCRIPTION (provided by applicant): Alcohol use disorders (AUDs) imposing enormous economic and social burdens on society today. AUDs are now understood as a chronic nervous system disorder characterized by a physiological dependence on alcohol beyond voluntary control. While environmental factors contribute to alcohol use, recent genetic studies reveal that the heritability of alcoholism is around 50%, illustrating that common genetic -variation contributes significantly to the etiology of alcoholism. However, the major target mediating alcohol reward signaling and the synaptic mechanisms of alcohol reward are poorly understood. Recent genetic studies have identified alcoholism associated gene variants in patients, one being the single nucleotide polymorphism (SNP) of an adenine to a guanine at position 118 of human OPRM1, a gene encoding the mu opioid receptor (MOR). This results in a nonsynonymous substitution of asparagine (N) at residue 40 to an aspartate (D). Although heterologous expression systems and animal studies suggest altered receptor trafficking and ligand binding affinities, there have been no conclusive studies determining the underlying cellular and molecular mechanisms of the N40D SNP on alcohol reinforcement, especially in a human neuronal context. To better understand the functional consequences of this SNP, the objective of this study is to investigate the interaction of alcohol and opioid signaling in human neurons carrying allelic variants for MOR N40 or D40 at the cellular and synaptic level. Using the novel induced pluripotent stem (iPS) cell and induced neuron (iN) cell techniques, it is possible to elucidate how the N40D gene variant alters alcohol reward signaling in human neurons. Using iPS cell lines from multiple individuals carrying homozygous MOR D40 or N40 alleles, human neurons of excitatory, inhibitory, and dopaminergic subtypes will be derived using the iN cell technology. To characterize the functional impact of this SNP, biochemical and morphological analyses examining changes in MOR expression, localization/trafficking, and degrees of glycosylation will be conducted using western blotting and immunohistochemistry. To give insight into interactions between alcohol exposure and MOR activation, presynaptic and dendritic markers will be analyzed to quantify synapse numbers and dendritic branching in alcohol-naïve, 1-day and 7-day alcohol and/or DAMGO treated iNs. I will also use electrophysiology to elucidate how the MOR N40 or D40 genotype differentially affects the more detailed functional parameters of synaptic transmission in iNs with or without alcohol and/or MOR activation. Our system using iPS cells carrying human gene variants provides the unique opportunity to dissect the interactions of alcohol exposure and MOR activation on synapse functionality in human neurons. As the long-term goal of this project is to understand the effect of human gene variants on alcohol sensitivity and synaptic function, results of this research will likely provide novel information about how to devise therapies for a heterogeneous group of AUDs.

描述（由申请人提供）：酒精使用障碍 (AUD) 给当今社会带来了巨大的经济和社会负担，AUD 现在被认为是一种慢性神经系统疾病，其特征是对酒精的生理依赖超出了自愿控制，而环境因素则导致了酒精。最近的遗传学研究表明，酒精中毒的遗传率约为 50%，这表明常见的遗传变异对酒精中毒的病因学有显着影响，然而，调节酒精奖赏信号和酒精中毒的主要目标。最近的遗传学研究发现了患者中与酒精中毒相关的基因变异，其中之一是人类 OPRM1（编码 mu 阿片受体的基因）第 118 位的腺嘌呤到鸟嘌呤的单核苷酸多态性 (SNP)。 (MOR)。这导致残基 40 处的天冬酰胺 (N) 被非同义取代为天冬氨酸 (D)，尽管异源表达系统和动物研究表明受体运输发生了改变。和配体结合亲和力，目前还没有确凿的研究来确定 N40D SNP 对酒精强化的潜在细胞和分子机制，特别是在人类神经背景下。为了更好地了解该 SNP 的功能后果，本研究的目的是使用新型诱导多能干 (iPS) 细胞和诱导神经元 (iN) 在细胞和突触水平上研究携带 MOR N40 或 D40 等位基因变体的人类神经元中酒精和阿片类信号传导的相互作用。通过细胞技术，可以阐明 N40D 基因变体如何改变人类神经元中的酒精奖赏信号传导，使用来自多个携带纯合 MOR D40 或 N40 等位基因的个体的 iPS 细胞系，可以使用来自兴奋性、抑制性和多巴胺能亚型的人类神经元。为了表征该 SNP 的功能影响，我们对 MOR 表达、定位/运输和程度的变化进行了生化和形态学分析。将使用蛋白质印迹和免疫组织化学进行糖基化，以深入了解酒精暴露和 MOR 激活之间的相互作用，将分析突触前和树突标记，以量化未接触酒精、1 天和 7 天酒精和酒精中的突触数量和树突分支。 /或 DAMGO 处理的 iNs，我还将使用电生理学来阐明 MOR N40 或 D40 基因型如何差异影响更详细的情况。我们的系统使用携带人类基因变异的 iPS 细胞，提供了剖析酒精暴露和 MOR 激活对人类神经元突触功能的相互作用的独特机会。该项目的长期目标是了解人类基因变异对酒精敏感性和突触功能的影响，这项研究的结果可能会提供有关如何为异质性 AUD 群体设计治疗方法的新信息。