Microglia-Mediated Astrocyte Activation in the Acute-to-Chronic Pain Transition

项目总结 虽然急性疼痛是对损伤组织作出反应的重要生物信号，但慢性疼痛发生在 疼痛信号持续时间长于最初的损伤，并对健康和生活质量产生有害影响。慢性疼痛 代表着巨大的公共卫生负担，几乎没有治疗选择。慢性疼痛不同于急性疼痛 疼痛具有几个独特的特征，包括长时间激活星形胶质细胞。星形胶质细胞是中枢神经系统细胞，具有 多种功能包括能量平衡，调节血脑屏障，清除 神经递质和突触传递的调节，所有这些都在激活状态下改变。 防止星形胶质细胞的激活是一个关键的治疗目标。小胶质细胞是人类免疫系统中的常驻免疫细胞 中枢神经系统被认为是星形胶质细胞激活的关键介质。通过这种方式，小胶质细胞的操纵 可以提供一种工具来防止或改变星形胶质细胞的激活，这反过来可能会防止疼痛成为 慢性病。我们实验室之前的研究表明，小胶质细胞从 从急性疼痛到慢性疼痛可以预防慢性疼痛。然而，一旦慢性疼痛导致小胶质细胞耗尽， 已经确定的是，疼痛类行为只有短暂的改善。对这些不同之处的一种解释 影响是，小胶质细胞可能通过触发星形胶质细胞激活而间接导致慢性疼痛 向慢性疼痛的过渡。然而，一旦星形胶质细胞被激活，小胶质细胞就不再在 疼痛信号和维持慢性疼痛的脊髓回路的改变是由于 星形胶质细胞的功能。我假设，在急性到慢性的转变过程中，小胶质细胞是必要的，也是充分的 激活星形胶质细胞，小胶质细胞在慢性疼痛中的作用完全依赖于星形胶质细胞的激活。在……里面 目的1，我将在致痛性外周损伤的小鼠模型中表征星形胶质细胞的激活 急性到慢性疼痛转变过程中小胶质细胞的选择性耗竭。我将进一步使用DREADS来 在急性到慢性转变中外源性激活星形胶质细胞以确定外源性激活是否 在小胶质细胞耗尽的情况下，星形胶质细胞足以维持向慢性疼痛的过渡。在目标2中，我 将利用幼稚小鼠的外源性小胶质细胞激活来确定小胶质细胞的激活是否足以 激活星形胶质细胞，如果星形胶质细胞的这种激活导致疼痛行为。最后，在《目标3》中我将 确定在慢性疼痛诱导过程中，小胶质细胞的哪些信号对星形胶质细胞的激活是重要的 使用细胞间相互作用分析来自星形胶质细胞和小胶质细胞的单核RNA测序数据。vbl.使用 在这项创新实验的研究建议中，我将揭示相关的作用和贡献 小胶质细胞和星形胶质细胞对慢性疼痛的作用，并为疼痛治疗产生新的靶点。建议数 研究是全面培训计划的一部分，将为我提供广泛的技术知识 尖端技术和分析。此外，这项提议的事业和科学目标将服务于 作为我未来K99/R00独立之路奖和学术研究生涯的基础。

PROJECT SUMMARY While acute pain is an important biological signal in response to injured tissue, chronic pain occurs when the pain signaling outlasts the initial injury and has deleterious effects on health and quality of life. Chronic pain represents an enormous public health burden with few therapeutic options. Chronic pain is distinct from acute pain with several unique features including long-lasting activation of astrocytes. Astrocytes are CNS cells with diverse functions including energy homeostasis, regulation of the blood brain barrier, clearance of neurotransmitters, and regulation of synaptic transmission, all of which are altered in activated states. Preventing activation of astrocytes represents a key therapeutic target. Microglia, the resident immune cells of the CNS, have been implicated as key mediators of astrocyte activation. In this way, microglia manipulation may provide a tool to prevent or alter astrocyte activation, which in turn may prevent pain from becoming chronic. Previous research from our lab has shown that depletion of microglia at the time of transition from acute pain to chronic pain prevents chronic pain. However, when microglia are depleted once chronic pain is established, there are only transient improvements in pain-like behaviors. One explanation for these different effects is that microglia may be contributing indirectly to chronic pain by triggering astrocyte activation during the transition to chronic pain. However, once astrocytes are activated, microglia cease to have an active role in pain signaling and the alterations in spinal cord circuits maintaining chronic pain are due to changes in astrocyte function. I hypothesize that at the acute-to-chronic transition microglia are necessary and sufficient to activate astrocytes and that microglia effects in chronic pain are entirely dependent on astrocyte activation. In Aim 1, I will characterize astrocyte activation in a mouse model of pain-producing peripheral injury after selective depletion of microglia at the acute-to-chronic pain transition. I will further use DREADDs to exogenously activate astrocytes at the acute-to-chronic transition to determine if exogenous activation of astrocytes is sufficient to maintain the transition to chronic pain in the context of microglia depletion. In Aim 2, I will use exogenous activation of microglia in a naïve mouse to determine if activation of microglia is sufficient to activate astrocytes and if this activation of astrocytes leads to pain-like behaviors. Finally, in Aim 3 I will determine which signals from microglia are important for astrocyte activation in the induction of chronic pain using cell-cell interaction analyses of single nuclei RNA-Sequencing data from astrocytes and microglia. Using the innovative experiments in this research proposal, I will uncover the relative roles and contributions of microglia and astrocytes to chronic pain and generate new targets for pain therapeutics. The proposed research is part of a comprehensive training plan that will provide me with extensive technical knowledge in cutting-edge technologies and analyses. In addition, the career and scientific goals of this proposal will serve as the foundation for my future K99/R00 Pathway to Independence award and an academic research career.