Chronic Stress Induces Neuroimmune Modulated Primary Muscle Afferent Sensitization

总结 许多慢性疼痛患者伴随着压力和焦虑水平的增加。多 报告指出免疫系统是压力和慢性疼痛发展之间的潜在联系。应力 激活免疫细胞，增加促炎细胞因子，可调节外周致敏 并通过初级感觉神经元中的基因表达变化来调节延长的超敏反应。我们 目的是表征应激诱导的调节初级传入的神经免疫相互作用 致敏并导致长时间缺血性肌肉疼痛的发展。宏观经济是关键之一， 免疫细胞参与这些调节外周敏化的神经免疫相互作用，但 巨噬细胞在应激诱导的肌肉痛觉过敏和超敏反应的发展中的作用仍然未知。 这项提议以一种新的方式探索了压力对周围神经系统的影响--选择性地抑制 或增强巨噬细胞介导的应激炎症反应， 肌肉疼痛相关的行为它将使用并验证以前从未使用过的环境富集损失 (LOE)不使用有害物理刺激的潜在混杂物的应激小鼠模型。 有趣的是，这种模式复制了COVID-19大流行期间经常经历的压力源- LOE 因为社交距离。我们的初步数据显示背根神经节基因表达的变化 (DRG)缺血性损伤（I/R）后肌肉传入神经的神经组织调制反应特性 部分由促炎细胞因子信号调节。LOE诱导的压力导致疼痛增加- 损伤后相关行为和骨骼肌中巨噬细胞浸润增加。我们假设 LOE诱导的应激通过免疫细胞依赖性机制诱导外周致敏， 调节缺血性肌痛的发展。我们将使用转基因和化学遗传学方法， 用LOE相关应激加缺血/再灌注（I/R）损伤的小鼠，我们的离体后爪肌肉传入神经 记录策略和疼痛相关行为分析。目标1将确定压力是否调节外周 损伤后（I/R）通过消耗巨噬细胞Fas中的巨噬细胞浸润而致敏， 诱导凋亡（MaFIA）小鼠暴露于LOE，有或没有I/R。电生理实验将是 与压力下DRG转录组的详细分析配对。目标2将评估压力的作用- 通过使用表达designer的转基因小鼠在I/R后诱导外周致敏的免疫改变 受体专门激活的设计师药物（DREADDs）驱动的Cre重组酶表达，从 溶菌酶-2启动子（LysM，仅表达骨髓单核细胞，如巨噬细胞）。 应激诱导的调节初级传入敏感化的神经免疫相互作用的机制将 阐明巨噬细胞在应激诱导的肌肉痛觉过敏和超敏反应中的作用。

SUMMARY Many patients with chronic pain concomitantly present with increased levels of stress and anxiety. Multiple reports point to the immune system as a potential link between stress and chronic pain development. Stress activates immune cells and increases pro-inflammatory cytokines, which can modulate peripheral sensitization and regulate prolonged hypersensitivity through gene expression changes in primary sensory neurons. Our objective is to characterize the stress-induced neuroimmune interactions that modulate primary afferent sensitization and lead to the development of prolonged ischemic muscle pain. Macrophages are one of the key immune cells involved in these neuroimmune interactions that regulate peripheral sensitization, but the role of macrophages in development of stress-induced muscle hyperalgesia and hypersensitivity remains unknown. This proposal explores effects of stress in the peripheral nervous system in a new way -- selectively inhibiting or enhancing macrophage -mediated inflammatory responses to stress and testing its role in the development muscle pain-related behaviors. It will use and validate a never before used, loss of environmental enrichment (LOE) mouse model of stress that does not employ the potential confounder of noxious physical stimulation. Interestingly, this paradigm replicates a stressor frequently experienced during the COVID-19 pandemic - LOE due to social distancing. Our preliminary data showed gene expression changes in the dorsal root ganglia (DRG) and innervated tissue modulated response properties of muscle afferents after ischemic injury (I/R) modulated, in part, by proinflammatory cytokine signaling. LOE induced stress resulted in increased pain- related behaviors after injury and increased macrophage infiltration in skeletal muscle. We hypothesize that LOE-induced stress induces peripheral sensitization via immune cell-dependent mechanisms to modulate the development of ischemic myalgia. We will use transgenic and chemogenetic approaches in mice with LOE-related stress plus ischemic/reperfusion (I/R) injury, our ex vivo hind paw muscle afferent recording strategies and pain-related behavioral assays. Aim 1 will identify if stress modulates peripheral sensitization after injury (I/R) through macrophage infiltration by depleting macrophages in macrophage fas- induced apoptosis (MaFIA) mice exposed to LOE with or without I/R. Electrophysiological experiments will be paired with detailed profiling of the transcriptome of the DRG under stress. Aim 2 will evaluate the role of stress- induced immune alterations in peripheral sensitization after I/R by using transgenic mice expressing designer receptors exclusively activated by designer drugs (DREADDs) driven by Cre-recombinase expression from the Lysozyme-2 promotor (LysM, only expressed myelomonocytic cells, such as macrophages).Characterization of the mechanisms of stress-induced neuroimmune interactions that modulate primary afferent sensitization will elucidate the role of macrophages in development of stress-induced muscle hyperalgesia and hypersensitivity.