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.

项目总结/摘要 偏头痛是一种痛苦的慢性神经系统疾病，是全球第二大致残性疾病。 导致偏头痛的两种机制是免疫细胞激活和降钙素基因相关 肽（CGRP）信号传导。本提案探讨了这两种机制之间的联系。皮质 扩散性抑郁症（CSD），被认为是偏头痛发作前的先兆，在一个亚群中， 与脑膜巨噬细胞活化增加有关。巨噬细胞可以激活 脑膜初级传入神经元并通过释放促炎细胞因子促进偏头痛， 其在偏头痛期间在患者脑脊液（CSF）中增加。CSD还增加CGRP合成 单细胞RNA测序数据支持CGRP受体亚基mRNA在细胞中的表达。 小鼠脑膜免疫细胞。患者血浆和CSF中CGRP升高以及CGRP受体的疗效 作为偏头痛治疗的拮抗剂支持CGRP在偏头痛发生中的中心作用。 然而，这些药物的作用机制还不完全清楚。考虑到CSD对 免疫细胞活化和CGRP释放，免疫细胞上CGRP受体的存在，以及 CGRP受体拮抗剂治疗偏头痛，我假设CGRP受体拮抗剂的疗效 是由它们抑制脑膜中促炎巨噬细胞活化的能力介导的。 在三个具体目标下描述的一组实验中，我将使用解剖学（目标1），功能（目标2）， 和行为（目标3）的方法来测试我的假设的背景下，一个微创模型偏头痛 光遗传扩散性抑制（optogenetic spreading depression，OSD）我将描述CGRP的OSD依赖性变化 使用荧光原位杂交和RT-qPCR检测脑膜免疫细胞中的受体表达（目的1）。我 将评估CGRP受体拮抗剂预防OSD诱导的巨噬细胞变化的能力。 形态和表型与使用实时体内双光子显微镜的激活一致（Aim 2A） 和流式细胞术（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诱导疼痛行为（目的3）。这些实验的结果可能不会 仅揭示了CGRP拮抗剂治疗效果的潜在机制，但可能是新的靶点 以及改善现有偏头痛疗法的机制。在完成这份提案的过程中，我将开发出有价值的 技术技能，并接受成为独立调查员所需的严格的智力培训，以及 我已经组建了一个由专家科学家和临床医生导师组成的团队，以帮助确保我的成功。因此，这 奖学金将使我实现我的长期目标，成为一名学术神经学家，科学家，在 疼痛研究和临床神经学的交叉点。