Sickle cell disease gut dysbiosis effects on CNS pain processing

研究总结 疼痛是镰状细胞病(SCD)患者最常见的并发症。患有SCD的患者患有 与血管闭塞性发作相关的剧烈急性疼痛和慢性疼痛，后者通常没有 很明显的病态。阿片类药物是治疗SCD疼痛的主要疗法，尽管它们有长期的负面影响 效果、缺乏疗效以及相关的治疗障碍。为了开发更好的治疗SCD疼痛的方法， 需要对SCD疼痛的神经生物学基础有更多的机械性理解。很少有研究发现 表征了大脑在SCD疼痛中的作用，尽管患者报告了中枢敏感化和情感协同作用 发病率，两者都与SCD疼痛强度和频率增加相关。中央核 杏仁核(CEA)是一个边缘大脑区域，由急性疼痛、慢性疼痛和情感障碍激活； 在转基因SCD小鼠中发现CEA活性增加。我们的长期目标是确定与SCD相关的因素 增加CEA神经元活性和痛样行为，以开发新的SCD疼痛疗法。这 该提案将研究肠道微生物群如何影响SCD的CEA活性。来自无菌小鼠的数据显示 一个完整的肠道微生物群对杏仁核的正常功能至关重要。患者和小鼠肠道微生物群的研究 SCD模型与健康对照组不同(即表现出生物失调)。反或益生菌操纵 肠道微生物群改变了SCD小鼠的疼痛行为和CEA信号。这样做的具体目的是 该提案将研究这些变化背后的神经机制。在具体目标1中，我们将使用光纤 光度法在以下条件下测量本底和疼痛诱发的CEA活性：(A)假 无菌小鼠用SCD粪便材料重新克隆，(B)接受青霉素纵向治疗的SCD小鼠， 和(C)接受迷走神经切断术的SCD小鼠。这些实验将使我们能够确定 以及SCD肠道菌群失调影响CEA活性的机制。在具体目标2中，我们将直接 操纵选定的CEA神经元群体，以确定每个神经元如何参与与生物失调相关的疼痛。我们 将首次使用光遗传学来抑制SCD重新克隆的假无菌小鼠中选定的CEA神经元的活动 排泄物。然后我们将确定CEA中的5HT2A受体活性是否与SCD相关的生物失调有关 疼痛。最后，我们将确定新的神经元信号通路，通过这些通路，SCD肠道微生物组可以 让人痛苦不已。这些实验将成为我的独立研究实验室的基础，它将 研究伤害性感觉的解剖学和化学基础，以开发个性化治疗方法 患有SCD和其他慢性疼痛障碍的个人。

Research Summary Pain is the most common complication for patients with sickle cell disease (SCD). Patients with SCD suffer from intense acute pain that is associated with vaso-occlusive episodes and chronic pain which frequently has no obvious pathology. Opioids are the main therapy used to treat SCD pain, despite their negative long-term side effects, lack of efficacy, and associated treatment barriers. In order to develop better therapies for SCD pain, a more mechanistic understanding of the neurobiological basis of SCD pain is needed. Very few studies have characterized the role of the brain in SCD pain, despite patient reports of central sensitization and affective co- morbidities, both of which are correlated with increased SCD pain intensity and frequency. The central nucleus of the amygdala (CeA) is a limbic brain region activated by acute pain, chronic pain, and affective disturbances; increased CeA activity is noted in transgenic SCD mice. Our long-term goal is to identify SCD-related factors that increase CeA neuronal activity and pain-like behaviors in order to develop novel SCD pain therapies. This proposal will examine how the gut microbiome influences CeA activity in SCD. Data from germ-free mice show that an intact gut microbiome is critical for normal amygdala function. The gut microbiome of patients and mouse models with SCD differs from that of healthy controls (i.e. exhibit dysbiosis). Anti- or probiotic manipulation of the gut microbiome changes pain-like behaviors and CeA signaling in SCD mice. The specific aims of this proposal will examine the neuronal mechanisms underlying these changes. In Specific Aim 1, we will use fiber photometry to measure background and pain-evoked CeA activity in the following conditions: (A) pseudo- germfree mice recolonized with SCD fecal material, (B) SCD mice that received longitudinal penicillin treatment, and (C) SCD mice that underwent vagotomy surgery. These experiments will allow us to determine the extent and mechanism through which SCD gut dysbiosis affects CeA activity. In Specific Aim 2, we will directly manipulate select populations of CeA neurons to determine how each is involved in dysbiosis-related pain. We will first use optogenetics to inhibit activity of select CeA neurons in pseudo-germfree mice recolonized with SCD fecal material. We will then determine if 5HT2A receptor activity in the CeA contributes to SCD dysbiosis-related pain. Finally, we will identify novel neuronal signaling pathways through which the SCD gut microbiome could be driving pain. These experiments will be the foundation for my independent research laboratory which will study the anatomical and chemical basis of nociception in order to develop personalized therapeutics for individuals suffering from SCD and other chronic pain disorders.