Role of GluN2A and MMPs in the CeA in Dependence-Induced Escalation of Etoh Drinking

CeA 中 GluN2A 和 MMP 在 Etoh 饮酒依赖性升级中的作用

基本信息

批准号：
10675691
负责人：
JOHN J. WOODWARD
金额：
$ 17.93万
依托单位：
MEDICAL UNIVERSITY OF SOUTH CAROLINA
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-02 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10675691
关键词：
Acute Adolescent Adult Air Alcohol consumption Alcohol dependence Alcohol withdrawal syndrome Alcohols Amino Acids Amygdaloid structure Animals Anxiety Applications Grants Area Bilateral Brain CRISPR/Cas technology Calcium-Sensing Receptors Cannulas Cell Nucleus Chronic Clinical Research Data Dendritic Spines Dependence Effectiveness Electrophysiology (science)Ethanol Extracellular Matrix FDA approved Female Fiber Future Gelatinase B Gelatinases Genes Glutamates Guide RNA Health Heavy Drinking Human Implant Individual Infusion procedures Intoxication Ion Channel Gating Knock-in Mouse Knock-out Laboratories Learning Legal Status Ligands Light Link Matrix Metalloproteinase Inhibitor Matrix Metalloproteinases Measures Mediating Mediator Memory Molecular Mus N-Methyl-D-Aspartate Receptors N-Methylaspartate NMDA receptor A1 Neurons Outcome Permeability Pharmaceutical Preparations Pharmacological Treatment Phenotype Photometry Play Population Proteins Resistance Rewards Role Signal Transduction Single Nucleotide Polymorphism Slice Synapses Synaptic plasticity Testing Time Transgenic Mice Transmembrane Domain Vertebral column Viral Virus Western Blotting Wild Type Mouse Withdrawal Work Zinc addiction alcohol abuse therapy alcohol exposure alcohol measurement alcohol sensitivity alcohol use disorder binge drinking cell type cohort drinking drinking behavior effective therapy endonuclease in vivo inhibitor knock-down learned behavior male negative affect neurotransmission new growth novel postoperative recovery preclinical study prevent recruit reduce symptoms social

项目摘要

Project Summary The legal status and widespread use of alcohol among the US population is associated with adverse health outcomes in both adolescents and adults. Nearly 88% of the US population have used alcohol at least once during their lifetime and rates of binge drinking and heavy alcohol use continue to be of concern. Pharmacological treatments for alcohol abuse show limited effectiveness and only three medications are FDA approved for treating alcohol dependence. One factor that underlies this problem is a lack of understanding of the mechanisms that contribute to excessive drinking. Recent results from our previous studies show that knock-in mice expressing ethanol resistant GluN2A N-methyl-D-aspartate receptors (NMDARs) drink the same as wild-type mice under baseline conditions but, unlike their wild-type counterparts, fail to escalate their drinking following repeated cycles of chronic intermittent ethanol exposure. These findings suggest that GluN2A NMDARs play a key role for in the loss of control over drinking observed in alcohol dependent subjects. Importantly, a similar lack of escalated drinking is observed following administration of inhibitors of zinc-dependent matrix metalloproteinases (MMPs) either i.c.v. or directly into the central nucleus of the amygdala (CeA). MMPs are key mediators of NMDA-mediated forms of synaptic plasticity where they remodel the extracellular matrix to support new growth and enlargement of glutamatergic dendritic spines. In this proposal we test the overarching hypothesis that MMPs and ethanol-sensitive GluN2A NMDARs in the CeA are critically involved in the transition to heavy drinking that is a hallmark of alcohol dependence. In Aim 1, we use slice electrophysiology, in vivo zymography and western blot analysis to test the hypothesis that CIE-induced changes in CeA neural signaling and MMP activity are absent in the GluN2A knock-in mice. In Aim 2, we use Crispr expressing AAVs and guide RNAs targeting the GluN2A gene to test the hypothesis that the CIE-induced increases in drinking and MMP activity requires functional GluN2A NMDARs in the CeA. The findings from these novel studies will provide critical data to support a future R01 grant application focused on determining which CeA cell types drive the altered drinking phenotype of the GluN2A knock-in mice and whether their expression in other key addiction-related brain areas contribute to this effect.

项目概要美国人口中酒精的合法地位和广泛使用与不良健康有关青少年和成人的结果。近 88% 的美国人至少饮酒过一次在他们的一生中，酗酒和大量饮酒的比率仍然令人担忧。药理作用酒精滥用的治疗方法效果有限，只有三种药物获得 FDA 批准治疗酒精依赖。造成这一问题的一个因素是缺乏对机制的了解导致过度饮酒。我们之前研究的最新结果表明，基因敲入小鼠表达乙醇抗性 GluN2A N-甲基-D-天冬氨酸受体 (NMDAR) 的饮酒方式与野生型相同小鼠在基线条件下，但与野生型小鼠不同，小鼠的饮酒量未能增加慢性间歇性乙醇暴露的重复周期。这些发现表明 GluN2A NMDAR 发挥着在酒精依赖受试者中观察到的饮酒失控的关键作用。重要的是，类似的缺乏服用锌依赖性基质抑制剂后观察到饮酒量增加金属蛋白酶 (MMP) 或 i.c.v.或直接进入杏仁核中央核（CeA）。 MMP 是关键 NMDA 介导的突触可塑性形式的介质，它们重塑细胞外基质以支持谷氨酸能树突棘的新生长和扩大。在此提案中，我们测试了总体假设 CeA 中的 MMP 和乙醇敏感的 GluN2A NMDAR 与转变密切相关酗酒是酒精依赖的标志。在目标 1 中，我们使用切片电生理学，体内酶谱分析和蛋白质印迹分析检验 CIE 诱导 CeA 神经信号传导变化的假设 GluN2A 敲入小鼠中不存在 MMP 活性。在目标 2 中，我们使用 Crispr 表达 AAV 并引导靶向 GluN2A 基因的 RNA 用于检验 CIE 诱导饮酒和 MMP 增加的假设活性需要 CeA 中的功能性 GluN2A NMDAR。这些新颖研究的结果将提供重要的支持未来 R01 拨款申请的数据，重点是确定哪些 CeA 细胞类型驱动改变 GluN2A 敲入小鼠的饮酒表型及其在其他关键成瘾相关中的表达是否大脑区域有助于这种效果。