Investigating cytoskeletal dynamics in astrocyte structure and cocaine seeking behavior

研究星形胶质细胞结构和可卡因寻求行为的细胞骨架动力学

• 批准号: 10607513
• 负责人: Janay Franklin
• 金额: $ 3.86万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10607513
• 关键词: Abstinence; Actins; Acute; Address; Affect; Area; Astrocytes; Automobile Driving; Behavior; Behavioral; Bioinformatics; Brain; Cessation of life; Chronic; Cocaine; Cocaine Abuse; Cocaine use disorder; Collaborations; Cytoskeletal Proteins; Cytoskeleton; Data; Data Analyses; Development; Down-Regulation; Elements; Extinction (Psychology); FDA approved; Functional disorder; Gene Expression; Genes; Genetic; Heroin; Home; Immunohistochemistry; Impairment; Individual; Intervention; Investigation; Knowledge; Learning; Measures; Mediating; Messenger RNA; MicroRNAs; Microinjections; Microscopy; Molecular; Neuroglia; Neurons; Nucleus Accumbens; Pathway Analysis; Pharmaceutical Preparations; Phosphorylation; Property; Proteins; Public Health; Publishing; Rattus; Relapse; Reporting; Research; Resolution; RiboTag; Role; Saline; Scientist; Self Administration; Structure; Surface; Synapses; Synaptic Transmission; Synaptic plasticity; Testing; Training; United States; Validation; Viral; Withdrawal; cocaine exposure; cocaine relapse; cocaine self-administration; cocaine use; computerized tools; confocal imaging; differential expression; drug of abuse; effective therapy; experience; experimental study; ezrin; insight; mRNA Expression; overdose death; pre-clinical; psychostimulant; reward circuitry; skills; stimulant use disorder; symposium; synaptic function; therapeutic candidate; therapeutic target; tool; transcriptome; transcriptome sequencing; treatment group; treatment strategy; virus genetics

ABSTRACT Cocaine abuse presents a significant public health concern across the United States, as the number of cocaine-related deaths in the United States has almost tripled since 2013. Despite an urgent need for intervention, an FDA-approved treatment for Cocaine Use Disorder is lacking. Hence, there is a considerable need for investigations into the mechanisms that drive relapse vulnerability. Recent advances indicate that cocaine-induced structural adaptations in astrocytes may contribute to relapse vulnerability. Astrocytes are the most abundant glial cell in the brain and regulate varied critical functions, including synaptic transmission and plasticity. Research in the Reissner lab has revealed that astrocytes in the nucleus accumbens are significantly structurally impaired following cocaine self-administration and extinction. In addition, preliminary data from our lab indicate that rat long-access (LgA, 6h/day) self-administration followed by prolonged abstinence (45d) leads to a significant ~40% decrease in astrocyte volume, surface area, and synaptic colocalization. However, the mechanisms driving these observations are unknown. I hypothesize that downregulation of astrocyte cytoskeletal dynamics is a major contributor to these effects. Accordingly, this proposal will examine how astrocyte cytoskeletal dynamics are altered following chronic cocaine self-administration and prolonged abstinence, and how manipulation of cytoskeletal proteins can influence cocaine seeking. Of note, phosphorylation of ezrin, an actin-cytoskeleton linker protein that is abundantly and preferentially expressed in astrocytes, is downregulated by acute cocaine exposure. Thus, the overarching hypothesis of this proposal is that cocaine and abstinence-induced astrocyte structural deficits result from alterations in astrocyte cytoskeletal dynamics, which further contribute to cocaine-seeking behaviors that are exacerbated following long-term abstinence. To test this hypothesis, in Aim 1 I will measure the effects of cocaine self-administration and abstinence on cytoskeletal proteins, including ezrin, and I will use an astrocyte-specific AAV to manipulate ezrin expression, to examine its role astrocyte structure and further in cocaine-seeking behaviors. In Aim 2, I will use a RiboTag AAV, to isolate astrocyte-specific mRNAs in conjunction with RNAseq as an unbiased approach to measure cocaine and abstinence-induced changes in the astrocyte transcriptome that may contribute to cocaine-seeking behavior. I will specifically analyze results to measure relative expression profiles of cytoskeletal proteins among these astrocyte mRNAs. These results will provide a greater understanding of how dysfunctions in astrocyte structure actively contribute to cocaine abuse and relapse.

摘要 可卡因滥用在美国引起了严重的公共卫生问题，因为 自2013年以来，美国与可卡因有关的死亡人数几乎增加了两倍。尽管迫切需要 由于缺乏干预，FDA批准的治疗可卡因使用障碍的方法缺乏。因此，有一个相当大的 需要调查导致易复发的机制。最近的进展表明， 可卡因诱导的星形胶质细胞结构适应可能导致复发脆弱性。星形胶质细胞是 神经胶质细胞是大脑中最丰富的神经胶质细胞，调节各种关键功能，包括突触传递和 可塑性Reissner实验室的研究表明，中脑核中的星形胶质细胞显著地 可卡因自我给药和消失后结构受损。此外，我们的初步数据显示， 实验室表明大鼠长期（LgA，6小时/天）自我给药，随后延长禁欲（45天） 导致星形胶质细胞体积、表面积和突触共定位显著减少~40%。然而，在这方面， 驱动这些观察结果的机制尚不清楚。我假设星形胶质细胞的下调 细胞骨架动力学是这些效应的主要贡献者。因此，本提案将审查如何 星形胶质细胞细胞骨架动力学在慢性可卡因自我给药后改变， 禁欲，以及如何操纵细胞骨架蛋白质可以影响可卡因寻求。值得注意的是， 埃兹蛋白是一种肌动蛋白-细胞骨架连接蛋白， 星形胶质细胞，被急性可卡因暴露下调。因此，这一提议的首要假设是 可卡因和戒断诱导星形胶质细胞结构缺陷是由于星形胶质细胞 细胞骨架动力学，这进一步有助于可卡因寻求行为， 长期禁欲为了验证这一假设，在目标1中，我将测量可卡因自我给药的效果。 和禁欲的细胞骨架蛋白，包括埃兹蛋白，我将使用星形胶质细胞特异性的AAV来操纵 ezrin表达，以检查其在星形胶质细胞结构中的作用，并进一步在可卡因寻求行为中。在目标2中，我 将使用RiboTag AAV，与RNAseq一起分离星形胶质细胞特异性mRNA，作为无偏倚的 一种测量可卡因和戒断诱导的星形胶质细胞转录组变化的方法， 导致可卡因成瘾我将专门分析结果，以衡量相对表达谱 在这些星形胶质细胞mRNA中的细胞骨架蛋白。这些结果将使我们更好地了解 星形胶质细胞结构功能障碍如何积极促进可卡因滥用和复发。