Function of astrocytes autophagy in brain homeostasis and opioid-induced maladaptive behavior and addiction, in the context of HIV

摘要/摘要 这项提议的首要目标是识别和描述星形胶质细胞的机械性途径。 负责药物诱发的结构和突触可塑性的变化，这是不适应行为的基础 阿片类药物滥用。星形胶质细胞是这些研究的合乎逻辑的焦点，因为它们密切参与各种不同的 神经功能，包括调节突触功能和可塑性，调节 兴奋性神经递质谷氨酸，但仍分泌和反应神经炎性细胞因子， 趋化因子和生长因子。这些过程本身是由自噬调节的：自噬过程 细胞既参与细胞成分的有序降解和回收，也参与能量的平衡 新陈代谢。尽管星形胶质细胞介导的兴奋和炎症与神经适应有关 与寻药行为、星形胶质细胞自噬的作用机制相交 谷氨酸系统和神经免疫信号之间的关系还不是很清楚。我们实验室的发现显示 一种关键的自噬蛋白Beclin1与细胞因子、趋化因子和 神经胶质细胞和神经元释放的生长因子。我们还发现自噬的激活会增加谷氨酸。 摄取与星形胶质细胞表达的谷氨酸转运体一起，共同建立了一个强有力的前提 目前提议的调查。扩展我们最初的研究，这里我们假设自噬- 星形胶质细胞-HIV介导的神经免疫信号和谷氨酸代谢改变神经元回路 促进阿片类药物使用障碍和滥用。由于阿片类药物和艾滋病毒感染是常见的共病， 阿片类药物通常被开给经历疼痛的艾滋病毒感染者，研究将包括 通过在一系列小鼠模型中使用生态艾滋病毒来评估艾滋病毒感染，促进所需的 行为学、神经解剖学和机械学研究，以解决目前的知识差距。以特定的目标 (SA)1我们将使用Becn1+/-和Becn1+/+小鼠来确定有或没有HIV和/或星形胶质细胞自噬的作用 慢性阿片暴露对阿片成瘾、戒断、长期戒断的进展和程度的影响， 并在经过验证的行为模型中复发。在SA 2中，我们将使用从AIM测试的小鼠身上获得的脑组织 1破译兴奋和炎症引起的神经化学和炎症变化之间的相互联系 通过晶状体观察阿片类药物滥用(伴有或不伴有HIV)的神经解剖学可塑性和不良适应行为 星形胶质细胞的自噬。在SA 3中，我们将使用脑细胞来进一步研究Beclin1 与μ-阿片受体和/或相关蛋白相互作用并分析Beclin1活性如何调节 阿片类药物对脑细胞内溶酶体转运和MOR细胞内转运的影响 (感染和不感染艾滋病毒)。预计这些发现将产生对自噬如何调节 星形胶质细胞和神经元回路之间的潜在串扰介导了OUD和滥用行为，导致 在艾滋病毒高危人群中开发新的治疗目标和预防OUD的战略。

SUMMARY/ABSTRACT The overarching goal of this proposal is to identify and characterize mechanistic pathways in astrocytes responsible for changes in drug-evoked structural and synaptic plasticity that underlie the maladaptive behavior in opioid drug abuse. Astrocytes are a logical focus for these studies, as they are intimately involved in diverse neuronal function, including modulation of synaptic function and plasticity, regulating concentrations of the excitatory neurotransmitter glutamate, and yet secrete and response to neuroinflammatory cytokines, chemokines, and growth factors. These processes are themselves regulated by autophagy: the process by which cells both engage in orderly degradation and recycling of cellular components as well as balancing energy metabolism. Although astrocyte-mediated excitation and inflammation have been implicated in neuroadaptations and drug-seeking behavior, the role of astrocytes autophagy in the mechanism underlying the intersection between the glutamate system and neuroimmune signaling, is not well understood. Findings from our lab showed that a key autophagy protein, Beclin1, is strongly associated with the secretion of cytokines, chemokines, and growth factors released from glia and neurons. We also showed that activation of autophagy increases glutamate uptake along with glutamate transporters expressed in astrocytes, collectively establishing a strong premise for the current proposed investigations. Extending our initial studies, here we hypothesize that autophagy- mediated neuroimmune signaling and glutamate metabolism in astrocytes ± HIV alters neuronal circuitry to promote opioid use disorder (OUD) and abuse. Since opioid and HIV-infection are common comorbidities, and opioids are commonly prescribed to HIV-infected individuals experiencing pain, studies will include assessment of HIV-infection through use of EcoHIV in a series of mouse models, facilitating the needed behavioral, neuroanatomical, and mechanistic studies to address the current knowledge gap. In Specific Aim (SA) 1 we will use Becn1+/- and Becn1+/+ mice to define the role of astrocyte autophagy with or without HIV and/or chronic opioid exposure on the progression and magnitude of opioid addiction, withdrawal, long-term abstinence, and relapse in validated behavioral models. In SA 2 we will use brain tissue harvested from mice tested in Aim 1 to decipher the interlink between excitation and inflammation-evoked changes in neurochemical and neuroanatomical plasticity and maladaptive behavior in opioid drug abuse (with or without HIV), through the lens of astrocytes autophagy. In SA 3 we will use brain cells to further investigate the mechanisms by which Beclin1 interacts with the μ-opioid receptor and/or associated proteins and analyze how Beclin1 activity modulates endolysosomal trafficking and degradation of MOR intracellular trafficking in brain cells in response to opioid (with and without HIV). Findings are expected to produce mechanistic insights into how autophagy regulates the underlying crosstalk between astrocytes and neuronal circuitry mediating OUD and abuse behaviors, yielding both novel therapeutic targets for development and strategies for preventing OUD in at risk HIV populations.