Serotonin Modulation of Trigeminal Nociceptors

描述（由申请人提供）：这项研究培训计划的长期目标是为Dayna R. lloyd博士提供一个培训机制，以发展和磨练作为神经科学家进行转化研究所需的关键技能。lloyd博士将利用博士后研究奖学金和结构化的临床研究硕士（MSCI）项目作为正式培训工具，开展一项关键的研究项目，利用基础科学和临床研究方法来回答颅面疼痛周围机制的重要问题。目前F32应用的总体方法是表征5 -羟色胺（5HT）受体的分布以及5HT对大鼠和人三叉神经辣椒素敏感伤害感受器（颅面疼痛传递感觉神经元）的整体综合作用。瞬时受体电位V1通道（TRPV1）由热和辣椒素（辣椒中的辛辣成分）连接，在三叉神经伤害感受器上高度表达，对疼痛超敏反应的发展至关重要。5HT可能通过三叉神经痛觉感受器增加TRPV1的活性，从而引起疼痛。本提案中概述的实验旨在验证5HT作用于外周5HT受体，增加辣椒素敏感三叉感觉神经元活性的假设。特异性Aim 1将利用荧光原位杂交和免疫组织化学的结合，表征5HT受体在表达trpv1的三叉神经感觉神经元中的表达，这些神经元来自于口腔面部炎症大鼠。特异性Aim 2将通过细胞信号（细胞内钙积累）和功能（促炎肽释放）测定5HT是否激活和/或致敏原代培养中的大鼠辣椒素敏感感觉神经元。特异性目的3将利用荧光免疫组织化学分析正常牙髓与炎症牙髓中表达trpv1的人伤害感受器上5HT受体的表达。此外，目的3将检测5HT在体外增加分离辣椒素敏感的人三叉神经伤害感受器促炎肽释放的能力。所提出的研究结果对于理解外周血清素如何参与调节大鼠和人类三叉神经伤害感受器至关重要。长期以来，血清素被认为是血小板、内皮细胞和免疫细胞在损伤部位释放的促炎介质，然而，血清素作用于感觉神经元调节疼痛的机制尚不清楚。这些知识将提供对周围颅面疼痛机制的见解，这可能会发现以前科学和医学未知的新的治疗靶点。我们的初步数据为假设提供了强有力的支持，结果将具有重大的翻译影响。

DESCRIPTION (provided by applicant): The long-term objectives of this research training plan are to serve as a training mechanism to develop and hone the critical skills necessary for conducting translational research as a neuroscientist for Dr. Dayna R. Loyd. Using a postdoctoral research fellowship and a structured Masters in Clinical Investigation (MSCI) program as a formal training vehicle, Dr. Loyd will conduct a critical research project utilizing both basic science and clinical research methodologies to answer important questions on the peripheral mechanisms of craniofacial pain. The overall approach of the present F32 application is to characterize the distribution of serotonin (5HT) receptors and overall integrated actions of 5HT on rat and human trigeminal capsaicin- sensitive nociceptors (craniofacial pain transmitting sensory neurons). The transient receptor potential V1 channel (TRPV1), gaited by heat and capsaicin (the pungent ingredient in chili peppers), is highly expressed on trigeminal nociceptors and is critical to the development of hypersensitivity to pain. 5HT may increase the activity of TRPV1 as a mechanism to evoke pain via trigeminal nociceptors. Experiments outlined in this proposal aim to test the hypothesis that 5HT, acting at peripheral 5HT receptors, increases the activity of capsaicin-sensitive trigeminal sensory neurons. Specific Aim 1 will characterize the expression of 5HT receptors, utilizing a combination of florescent in situ hybridization and immunohistochemistry, in TRPV1-expressing trigeminal sensory neurons from rats with orofacial inflammation. Specific Aim 2 will determine whether 5HT activates and/or sensitizes rat capsaicin-sensitive sensory neurons in primary culture using cell signaling (intracellular calcium accumulation) and functional (proinflammatory peptide release) assays. Specific Aim 3 will analyze 5HT receptor expression on human TRPV1-expressing nociceptors in normal versus inflamed dental pulp using fluorescent immunohistochemistry. In addition, aim 3 will examine the ability of 5HT to increase proinflammatory peptide release from isolated capsaicin-sensitive human trigeminal nociceptors in vitro. The results of the proposed research are critical to understanding how peripheral serotonin is involved in modulating both rat and human trigeminal nociceptors. Serotonin has long been recognized as a pro- inflammatory mediator released at the site of injury by platelets, endothelial cells, and immune cells, however, the mechanism(s) by which serotonin acts on sensory neurons to modulate pain are unknown. This knowledge will provide insight on peripheral craniofacial pain mechanisms, which may uncover novel therapeutic targets previously unknown to science and medicine. Our preliminary data provide strong support for the hypothesis and the results will have a significant translational impact. PUBLIC HEALTH RELEVANCE: Craniofacial pain, such as migraine, represents a prevalent and expensive healthcare problem. This project will evaluate the potential modulation of rat and human trigeminal sensory neurons by serotonin. If so, then novel drugs targeting peripheral serotonin receptors may be highly effective at treating craniofacial pain.