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.

项目概要 酒精滥用和成瘾是一场日益严重、破坏性越来越大的国家和全球危机。 迫切需要新的治疗干预措施，而不是控制症状，而是逆转 酒精引起的神经生理变化是酒精和无数其他成瘾药物产生的一种影响。 在大脑中，是长期突触抑制 (LTD) 的破坏，特别是突触前形式的 LTD 发生在皮质纹状体突触处的纹状体与强迫性和习惯性行为有关， 包括药物成瘾引起的背外侧纹状体 (DLS) 的 LTD 破坏与此相关。 增加了习惯性行为，而LTD恢复则恢复了药物破坏的目标导向行为。 发现酒精会破坏纹状体突触前 mu 阿片受体 (MOR) 介导的 LTD，该受体在 DLS 中发生 仅在前岛叶皮质输入（AIC-DLS 突触）处。 输入不受酒精暴露的影响 识别突触前发生的分子变化。 AIC-DLS 突触的酒精暴露，但丘脑纹状体突触的酒精暴露，可能会揭示新的目标 酒精使用障碍和成瘾的药物干预具体问题是。 使用药理学和遗传工具识别蛋白质-蛋白质相互作用的突触前特异性变化 费力、低效，并且依赖于信号通路的先验知识，可能缺少关键分子 根据我们的初步数据，我们的中心假设是体内酒精。 暴露会改变 MOR 介导的 LTD 所需的 PKA 依赖性突触前蛋白组织 我们将利用研究人员的生理学和蛋白质组学专业知识来进行诱导。 开发一种新的生化方法，能够选择性地确定精确的离体和体内 精确地在 AIC 突触末端内发生的蛋白质相互作用组的变化（但不发生在 MOR 介导的 LTD 诱导期间以及体内暴露后 DLS 中的丘脑纹状体末端） 酒精敏感的 AIC-DLS 突触与酒精不敏感的突触之间的比较。 丘脑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 这种方法采用小鼠脑切片电生理学，在酒精体内暴露， 和病毒载体驱动的蛋白质表达，允许在突触前末梢内进行邻近标记 使用这个新工具，我们将打开突触前蛋白质组的“黑匣子”。 我们预测的变化将揭示以前未识别的突触可塑性蛋白质-蛋白质相互作用 此外，我们的目标是识别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目标。