A novel proteomics approach to identify alcohol-induced changes in synapse-specific presynaptic protein interactions.

一种新的蛋白质组学方法，用于识别酒精引起的突触特异性突触前蛋白质相互作用的变化。

• 批准号: 10651991
• 负责人: Brady Atwood
• 金额: $ 22.05万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-05 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10651991
• 关键词: Ablation; Address; Adult; Affect; Alcohol consumption; Alcohol dependence; Alcohols; Animals; Anterior; Behavior; Biochemical; Biotin; Biotinylation; Brain; Brain region; Cell Culture Techniques; Cell physiology; Compulsive Behavior; Corpus striatum structure; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Data; Development; Disease; Disease model; Dorsal; Drug Addiction; Drug Modelings; Drug abuse; Electrophysiology (science); Ethanol; Female; Future; Genetic; Glutamates; Goals; Infusion procedures; Injections; Intervention; Knowledge; Label; Ligase; Long-Term Depression; Mass Spectrum Analysis; Mediating; Mental Depression; Methodology; Methods; Modeling; Molecular; Molecular Target; Mus; Naltrexone; Pathologic; Pharmaceutical Preparations; Pharmacology Study; Physiological; Post-Translational Protein Processing; Predisposition; Presynaptic Terminals; Proteins; Proteome; Proteomics; Receptor Signaling; Research Personnel; Resistance; Saline; Signal Pathway; Signal Transduction; Signaling Protein; Slice; Streptavidin; Substance Use Disorder; Synapses; Synaptic Potentials; Synaptic plasticity; Techniques; Testing; Thalamic structure; Therapeutic; Tissues; Translations; Viral Vector; Work; addiction; adeno-associated viral vector; alcohol abuse therapy; alcohol behavior; alcohol effect; alcohol exposure; alcohol misuse; alcohol use disorder; cognitive control; differential expression; dorsal proteins; drug of abuse; experimental study; genetic manipulation; habit learning; in vivo; innovation; insight; male; mu opioid receptors; neurophysiology; new therapeutic target; novel; novel strategies; novel therapeutic intervention; pharmacologic; presynaptic; protein expression; protein protein interaction; receptor; response; restoration; socioeconomics; symptom management; synaptic depression; therapeutic target; tool; vesicular release

Project Summary Alcohol misuse and addiction is a growing and increasingly destructive socioeconomic national and global crisis. There is a critical need for novel therapeutic interventions that, rather than managing symptoms, reverse alcohol-induced neurophysiological changes. One effect that alcohol, and myriad other addicitive drugs, produce in the brain, is the disruption of long-term synaptic depression (LTD), particularly the presynaptic form of LTD that occurs at corticostriatal synapses. The striatum is associated with compulsive and habitual behaviors, including drug addiction. Drug-induced disruption of LTD in dorsolateral striatum (DLS) is associated with increased habitual behavior whereas LTD restoration restores drug-disrupted goal-directed behavior. We have found that alcohol disrupts striatal presynaptic mu opioid receptor (MOR)-mediated LTD that, in the DLS, occurs exclusively at anterior insular cortex inputs (AIC-DLS synapses). MOR-short-term depression at thalamic striatal inputs are unaffected by alcohol exposure. Identifying the molecular changes that occur presynaptically following alcohol exposure at AIC-DLS synapses, but not at thalamostriatal synapses, may reveal new targets for pharmacological interventions in alcohol use disorder and addiction. The specific problem though is that identifying presynaptic-specific changes in protein-protein interactions using pharmacological and genetic tools is laborious, inefficient, and relies on a priori knowledge of signaling pathways, likely missing key molecular players that could be affected. Based on our preliminary data, our central hypothesis is that in vivo alcohol exposure alters PKA-dependent presynaptic protein organization that is required for MOR-mediated LTD induction. We will capitalize on the combined physiological and proteomic expertise of the investigators to develop a novel biochemical methodology that is able to selectively determine the precise ex vivo and in vivo changes that occur in the protein interactome precisely within AIC synaptic terminals (but do not occur within thalamostriatal terminals) in DLS during the induction of MOR-mediated LTD and following in vivo exposure to alcohol. The comparison between these alcohol-sensitive AIC-DLS synapses and the alcohol-insensitive thalamic inputs will allow us to determine what makes some synapses susceptible to alcohol’s deleterious effects and others resistant. This methodology employs mouse brain slice electrophysiology, in vivo alcohol exposure, and viral vector-driven expression of proteins that allow for proximity labeling within presynaptic terminals with subsequent proteomic analyses. Using this new tool we will open the “black box” of presynaptic proteome changes that we predict will uncover previously unidentified synaptic plasticity protein-protein interactions that are disrupted by alcohol and other addictive drugs. Furthermore, we aim to identify novel protein interactions that may prove to be therapeutic targets for treating alcohol use disorder and addiction.

项目摘要 酒精滥用和成瘾是一个日益严重和破坏性越来越大的国家和全球社会经济危机。 迫切需要新的治疗干预措施，而不是管理症状，逆转 酒精引起的神经生理学变化。酒精和无数其他兴奋剂产生的一个效果是， 在大脑中，是长期突触抑制（LTD）的中断，特别是LTD的突触前形式 发生在皮质纹状体突触上纹状体与强迫性和习惯性行为有关， 包括毒瘾药物诱导的背外侧纹状体（DLS）LTD的破坏与以下因素相关： 增加的习惯性行为，而LTD恢复恢复药物中断的目标导向行为。我们有 发现酒精破坏纹状体突触前μ阿片受体（莫尔）介导的LTD，在DLS中， 仅在前岛叶皮层输入（AIC-DLS突触）。MOR-丘脑纹状体的短期抑制 输入不受酒精暴露的影响。识别突触前发生的分子变化， 酒精暴露在AIC-DLS突触，而不是在丘脑纹状体突触，可能揭示新的目标， 酒精使用障碍和成瘾的药物干预。但具体问题是， 使用药理学和遗传学工具鉴定蛋白质-蛋白质相互作用中的突触前特异性变化 是费力的，低效的，并且依赖于信号传导途径的先验知识，可能缺少关键分子， 可能受到影响的球员。根据我们的初步数据，我们的中心假设是，体内酒精 暴露改变了MOR介导的LTD所需的PKA依赖性突触前蛋白质组织 诱导我们将利用研究人员的生理学和蛋白质组学专业知识， 开发一种新的生物化学方法，能够选择性地确定精确的离体和体内 发生在蛋白质相互作用体中的变化精确地在AIC突触末梢内（但不发生在AIC突触末梢内）。 在诱导MOR介导的LTD过程中以及在体内暴露于 酒精这些酒精敏感的AIC-DLS突触和酒精不敏感的AIC-DLS突触之间的比较 丘脑的输入将使我们能够确定是什么使一些突触容易受到酒精的有害影响 其他人则抵抗。该方法采用小鼠脑切片电生理学，体内酒精暴露， 以及病毒载体驱动的蛋白质表达，其允许在突触前末端内进行邻近标记， 随后的蛋白质组学分析。利用这一新工具，我们将打开突触前蛋白质组的“黑匣子 我们预测这些变化将揭示以前未被发现的突触可塑性蛋白质-蛋白质相互作用， 被酒精和其他成瘾药物破坏。此外，我们的目标是确定新的蛋白质相互作用 这可能被证明是治疗酒精使用障碍和成瘾的治疗靶点。