Functional Analysis of Distinct Nociceptor Populations

描述（申请人提供）：触觉在日常生活中是必不可少的。此外，触觉过敏是一种与慢性神经性疼痛和持续性炎症性疼痛相关的常见问题。尽管它很重要，但令人惊讶的是，人们对触觉的分子和细胞机制知之甚少，也不知道不同的转导机制是否介导了无害和疼痛的机械刺激。瞬时受体电位锚蛋白1 （TRPA1）是一个很有前途的机械转导候选者。大量证据表明，TRPA1参与了组织炎症或神经损伤后体感觉神经元的机械转导及其对机械刺激的敏化。此外，有争议的证据表明，TRPA1参与神经损伤和炎症后的冷转导和冷致敏。然而，关于TRPA1在机械转导和冷转导中的具体功能作用，仍然存在一些重要的问题。1)我们和其他实验室的证据表明，TRPA1在非神经元角质形成细胞和感觉神经末梢中表达，但TRPA1在两种细胞类型中对皮肤初级传入神经元对机械力或低温的净反应的贡献尚不清楚。本提案中的实验将使用两种新的转基因小鼠系，其中TRPA1从角质形成细胞或感觉神经元中选择性地删除，以确定两种细胞类型中TRPA1对机械和冷刺激的行为和初级传入纤维反应的贡献。2)感觉神经元或角质形成细胞中的TRPA1是否介导质膜对机械力或冷的反应尚不清楚。本应用中的实验将在全细胞膜片钳记录期间使用创新的定量焦点机械刺激器来确定TRPA1对感觉神经元和角质形成细胞膜中机械电流的贡献。机械性超敏反应是与神经性疼痛和炎症性疼痛相关的最常见、最具破坏性的症状之一。TRPA1在神经损伤或炎症后初级传入纤维对机械和冷刺激的敏化中的作用尚不清楚。本实验将确定感觉神经元或角化细胞中的TRPA1是否介导发生在神经性疼痛模型和持续性外周炎症模型中的机械或冷致敏。在本研究中，使用细胞、纤维和行为分析的互补特异性目标将提供多方面的方法来确定TRPA1在正常组织、神经性和炎症性疼痛中的感觉传导功能。

DESCRIPTION (provided by applicant): Touch sensation is essential to daily life. Further, hypersensitivity to touch is a frequent problem associated with chronic neuropathic pain and persistent inflammatory pain. Despite its importance, amazingly little is known about the molecular and cellular mechanisms underlying touch sensation and whether different transduction mechanisms mediate innocuous versus painful mechanical stimuli. A promising mechanotransduction candidate is the Transient Receptor Potential Ankyrin 1 (TRPA1). Significant evidence suggests that TRPA1 contributes to mechanotransduction in somatosensory neurons and their sensitization to mechanical stimuli after tissue inflammation or nerve injury. Further, controversial evidence suggests that TRPA1 is involved in cold transduction and cold sensitization after nerve injury and inflammation. A number of important questions remain, however, about the specific functional roles of TRPA1 in mechanotransduction and cold transduction. 1) Evidence from our laboratory and others indicates that TRPA1 is expressed by non- neuronal keratinocytes, as well as sensory nerve terminals, yet the contribution of TRPA1 in either cell type to the net responses of cutaneous primary afferent neurons to mechanical force or cold temperatures is unknown. Experiments in this proposal will use two new lines of transgenic mice where TRPA1 is selectively deleted from either keratinocytes or sensory neurons to determine the contribution of TRPA1 in either cell type to behavioral and primary afferent fiber responses to mechanical and cold stimuli. 2) Whether TRPA1 in sensory neurons or keratinocytes mediates the plasma membrane response to mechanical force or cold is not clear. Experiments in this application will use an innovative, quantitative focal mechanical stimulator during whole cell patch clamp recordings to identify the contribution of TRPA1 to mechanical currents in the membrane of sensory neurons and keratinocytes. 3) Mechanical hypersensitivity is one of the most frequent, devastating symptoms associated with neuropathic and inflammatory pain. The contribution of TRPA1 to sensitization of primary afferent fibers to mechanical and cold stimuli after nerve injury or inflammation is unknown. Experiments in this proposal will determine whether TRPA1 in either sensory neurons or keratinocytes mediates the mechanical or cold sensitization that occurs in a model of neuropathic pain, and a model of persistent peripheral inflammation. In this proposal, complimentary Specific Aims using cellular, teased fiber and behavioral assays will provide a multifaceted approach to identify functional roles of TRPA1 in sensory transduction in normal tissue, and during neuropathic and inflammatory pain.