The role of meningeal immune cells in the efficacy of CGRP-based migraine therapies

脑膜免疫细胞在 CGRP 偏头痛疗法疗效中的作用

• 批准号: 10604482
• 负责人: Talia Adi
• 金额: $ 4.77万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10604482
• 关键词: Afferent Neurons; Anatomy; Attenuated; Auras; Behavior; Behavioral; Behavioral Assay; Calcitonin; Calcitonin Gene-Related Peptide; Calcitonin-Gene Related Peptide Receptor; Caring; Cells; Cephalic; Cerebrospinal Fluid; Chronic; Classic Migraine; Clinical; Coculture Techniques; Data; Development; Disease; Ensure; Event; Fellowship; Flow Cytometry; Fluorescent in Situ Hybridization; Future; Goals; Headache; Hypersensitivity; Immune; Immune Targeting; Inflammation Mediators; Inflammatory; Macrophage; Macrophage Activation; Mechanics; Mediating; Meningeal; Meninges; Mentors; Messenger RNA; Migraine; Modeling; Morphology; Mus; Neurologic; Neurologist; Neurology; Neurons; Outcome; Outcome Study; Pain; Pain Measurement; Pain Research; Patients; Peptide Signal Sequences; Peptide Synthesis; Phenotype; Physicians; Physiology; Plasma; RNA; Reporting; Research; Research Personnel; Role; Scientist; Spreading Cortical Depression; Structure of trigeminal ganglion; Technical Expertise; Testing; Time; Training; Treatment Efficacy; Work; antagonist; career; clinically relevant; cytokine; drug action; experimental study; immune activation; improved; in vivo; intravenous administration; mechanical allodynia; migraine treatment; minimally invasive; mouse model; nervous system disorder; neural; novel; optogenetics; pain behavior; pain reduction; prevent; receptor; receptor expression; receptor-activity-modifying protein; single-cell RNA sequencing; spreading depression; success; two photon microscopy

Project Summary/Abstract Migraine is a painful, chronic neurological disorder that represents the second most disabling illness worldwide. Two of the mechanisms that to contribute to migraine pain are immune cell activation and calcitonin gene-related peptide (CGRP) signaling. This proposal investigates the connection between these two mechanisms. Cortical spreading depression (CSD), thought to underlie the aura that precedes migraine attack in a subpopulation of migraineurs, is associated with increased meningeal macrophage activation. Macrophages can activate meningeal primary afferent neurons and promote migraine pain through release of pro-inflammatory cytokines, which are increased in patient cerebrospinal fluid (CSF) during migraine. CSD also increases CGRP synthesis and release, and single-cell RNA sequencing data support the expression of CGRP receptor subunit mRNA in mouse meningeal immune cells. Elevated CGRP in patient plasma and CSF and the efficacy of CGRP receptor antagonists as migraine therapies support the central role of CGRP in the development of migraine pain. However, the mechanism of action of these drugs is incompletely understood. Given the effect of CSD on immune cell activation and CGRP release, the presence of CGRP receptor on immune cells, and the use of CGRP receptor antagonists to treat migraine, I hypothesize that the efficacy of CGRP receptor antagonists is mediated by their ability to inhibit the activation of pro-inflammatory macrophages in the meninges. In a set of experiments described under three specific aims, I will use anatomical (Aim 1), functional (Aim 2), and behavioral (Aim 3) approaches to test my hypothesis in the context of a minimally invasive model of migraine with aura (optogenetic spreading depression, OSD). I will characterize OSD-dependent changes in CGRP receptor expression in meningeal immune cells using fluorescent in situ hybridization and RT-qPCR (Aim 1). I will assess the ability of CGRP receptor antagonists to prevent OSD-induced changes in macrophage morphology and phenotype consistent with activation using real-time in vivo two-photon microscopy (Aim 2A) and flow cytometry (Aim 2B). Finally, I will assess the contribution of macrophage CGRP receptors to OSD- induced pain behavior using a selective depletion strategy (Aim 3). The outcomes of these experiments may not only reveal a mechanism underlying the therapeutic efficacy of CGRP antagonists but potentially novel targets and mechanisms to improve existing migraine therapies. In completing this proposal, I will develop valuable technical skills and receive rigorous intellectual training necessary to becoming an independent investigator, and I have assembled a team of expert scientist and clinician mentors to help ensure my success. Thus, this fellowship will allow me to achieve my long-term goal of becoming an academic neurologist-scientist working at the intersection of pain research and clinical neurology.

项目摘要/摘要 偏头痛是一种痛苦的，慢性的神经系统疾病，代表了全球第二大疾病。 有助于偏头痛疼痛的两种机制是免疫细胞激活和降钙素基因相关的 肽（CGRP）信号传导。该提案研究了这两种机制之间的联系。皮质 散布抑郁症（CSD），被认为是偏头痛攻击之前的光环的基础 偏头痛与脑膜巨噬细胞激活增加有关。巨噬细胞可以激活 脑膜原发性传入神经元并通过释放促炎细胞因子来促进偏头痛疼痛， 偏头痛期间患者脑脊液（CSF）的增加。 CSD还增加了CGRP合成 和释放，单细胞RNA测序数据支持CGRP受体亚基mRNA在 小鼠脑膜免疫细胞。患者血浆和CSF的CGRP升高以及CGRP受体的功效 作为偏头痛疗法的拮抗剂支持CGRP在偏头痛疼痛发展中的核心作用。 但是，这些药物的作用机理尚未完全理解。考虑到CSD对 免疫细胞激活和CGRP释放，免疫细胞上的CGRP受体的存在以及使用 CGRP受体拮抗剂治疗偏头痛，我假设CGRP受体拮抗剂的功效 它们的抑制能力抑制脑膜中促炎性巨噬细胞的活化的能力。 在三个特定目标下描述的一组实验中，我将使用解剖学（AIM 1），功能（AIM 2）， 和行为（AIM 3）的方法来检验我的假设，即偏头痛的微创模型的背景 带有光环（光学扩散抑郁症，OSD）。我将表征CGRP的OSD依赖性更改 使用荧光原位杂交和RT-QPCR在脑膜免疫细胞中的受体表达（AIM 1）。我 将评估CGRP受体拮抗剂防止OSD诱导的巨噬细胞变化的能力 形态和表型与实时在体内两光子显微镜下激活一致（AIM 2A） 和流式细胞仪（AIM 2B）。最后，我将评估巨噬细胞CGRP受体对OSD的贡献 使用选择性耗竭策略引起疼痛行为（AIM 3）。这些实验的结果可能不是 仅揭示了CGRP拮抗剂的治疗功效的基础机制，但可能是新的靶标 和改善现有偏头痛疗法的机制。在完成这项建议时，我将发展有价值 技术技能并接受成为独立研究人员所必需的严格智力培训，并且 我已经组建了一个由专家科学家和临床医师导师组成的团队，以帮助确保自己的成功。因此，这个 奖学金将使我能够实现成为一名学术神经科医生科学家的长期目标 疼痛研究与临床神经病学的交集。