Investigating the Role of Microglia in Methamphetamine Use Disorder

摘要 物质使用障碍是一种慢性复发性疾病，其特征是反复使用药物，尽管 负面后果。尽管在研究潜在的分子机制方面做出了广泛的努力 甲基苯丙胺使用障碍，目前还没有FDA批准的治疗方法来治疗这种衰弱 疾病。重复使用甲基苯丙胺可导致脑区基因表达的长期变化 与奖励处理和毒品寻找行为有关，最近的证据表明，冰毒- 诱导的神经炎症也可能与药物的行为和分子反应有关。小胶质细胞， 大脑中的常驻免疫细胞是神经炎性反应的主要驱动力，但这些细胞的作用 细胞对冰毒相关行为的调控知之甚少。在这个提案中，我将检查小胶质细胞 静脉注射冰毒小鼠模型中基因表达的动态变化 甲基苯丙胺自身给药(甲基IVSA)和多组体分子分析。AIM的初步数据 1(F99)表明小胶质细胞通过诱导神经炎性基因表达来对冰毒暴露作出反应 并相应地改变它们的细胞形态。此外，由于我们的初步结果表明， 小胶质细胞的耗尽减少了长期戒毒后的寻药行为，我建议进一步研究 小胶质细胞在吸食冰毒和渴求冰毒过程中的转录和表观遗传学变化 寻找。为此，将从甲基IVSA中分离小胶质细胞进行RNA测序和染色质分析 使用剪切和标记进行分析(F99)。为此，我将接受培训，分离小胶质细胞以提取RNA和染色质， 以及数据分析所需的下游生物信息管道。对于目标2(K00)，我将确定一个 博士后导师，与他一起研究早期生活应激对有助于以后生活的小胶质细胞的影响 对精神和物质使用障碍的易感性。这些研究将加深我们对 冰毒强化过程中的小胶质细胞转录组和表观基因组以及这些细胞如何影响冰毒摄取 并寻求行为，同时也培养我成为博士后学者的发展，并最终为 作为一名独立调查员，在学术终身教职轨道上取得成功的职业生涯。

ABSTRACT Substance use disorder is a chronic relapsing disease that is characterized by repeated drug use despite negative consequences. Despite extensive efforts into examining the underlying molecular mechanisms of methamphetamine use disorder, there are currently no FDA approved therapeutics to treat this debilitating disease. Repeated methamphetamine (METH) use induces long-term gene expression changes in brain regions associated with reward processing and drug-seeking behavior, and recent evidence suggests that METH- induced neuroinflammation may also be involved in behavioral and molecular responses to the drug. Microglia, the resident immune cells in the brain, are principal drivers of neuroinflammatory responses, yet the role of these cells in the regulation of METH-related behaviors is poorly understood. In this proposal, I will examine microglial gene expression dynamics during METH-taking and seeking using a mouse model of intravenous methamphetamine self-administration (METH IVSA) and multi-omic molecular analyses. Preliminary data in Aim 1 (F99) demonstrate that microglia respond to METH-exposure by inducing neuroinflammatory gene expression and, accordingly, changing their cellular morphology. Additionally, since our preliminary results suggest that depletion of microglia attenuates drug-seeking after prolonged abstinence, I propose to further investigate the transcriptional and epigenetic changes in microglia in response to METH-taking and during METH craving and seeking. To this end, microglia will be isolated from METH IVSA for RNA sequencing analysis and chromatin profiling using CUT&Tag (F99). For this, I will be trained to isolate microglia for RNA and chromatin extraction, as well as the downstream bioinformatic pipelines required for data analysis. For Aim 2 (K00), I will identify a postdoctoral mentor with whom to study the effect of early life stress on microglia that contribute to later life susceptibility to psychiatric and substance use disorders. These studies will sharpen our understanding of the microglial transcriptome and epigenome during METH reinforcement and how these cells influence METH-taking and seeking behavior, while also fostering my development into a postdoctoral scholar, and ultimately for a successful career as an independent investigator in an academic tenure-track position.