Role of trpv4 in trigeminally mediated nociception

三叉神经介导的疼痛障碍在分子水平上还没有完全被理解。色氨酸(V)离子 通道被认为在三叉神经感觉转导中起作用。TRPV4是多模的 例如由机械、热和紧张性刺激激活的，并且TRPV4基因的产物已经被 在三叉神经节感觉神经元及其外周投射中检测到，f/PYF‘’小鼠 由PI产生的，对于有害的机械刺激和紧张性刺激具有更高的阈值。关键人物 TRPV4*三叉神经感觉神经元是否以及如何转导这种刺激仍未解决 这会引发伤害性行为。因此，这项提议的目标是研究分子和细胞 FRPV4介导三叉神经伤害性渗透和机械刺激的传导机制 通过开发一种新的小鼠模型来研究神经元。需要检验的假设是TRPV4基因 三叉神经感觉神经元在伤害性反应中的表达是神经感觉转导的关键 刺激物。具体地说，(1)TRPV4~*~小鼠三叉神经细胞的原代培养将通过 膜片钳和钙离子成像对张力和机械刺激的反应，受激活的 蛋白酶激活受体2(PAR-2)，一种生理上相关的前痛觉通路，以及它们的 电压门控钠通道(LNA)和辣椒素反应的“调谐”；(2)与AIM(1)相同，但 用于从感觉神经元特异的、可诱导的TRPV4‘~小鼠中分离感觉神经元；(3)感觉神经元-- 将产生特定的、可诱导的TRPV4^~小鼠，以测试这些小鼠对有害渗透刺激的反应 以及三叉神经外周辣椒素刺激的渗透调节，由PAR-2的激活调节。为 针对(1)-(2)，原代培养的神经元将被基因编码的荧光报告基因标记， 在细菌转基因小鼠染色体系中表达。对于AIM(3)，小鼠将受到 脑干三叉神经脊束核对脑电刺激反应的行为学和c-fos研究 刺激。因此，三叉神经伤害性刺激的感觉转导和传递依赖于 TRPV4将在细胞水平上被解构，阐明三叉神经感觉的相对贡献 神经元。此外，还将阐明TRPV4如何在辣椒素反应和调谐中发挥作用。 Ina-.以及PAR-2对TRPV4的调控作用。因此，我们的新老鼠模型将帮助我们更好地理解 三叉神经节神经元对伤害性刺激的反应及TRPV4基因在其中的作用 过程，希望为三叉神经介导的疼痛的治疗开辟新的途径。

Trigeminally-mediated pain disorders are incompletely understood at the molecular level. TRP(V) ion channels have been implicated to function in trigeminal sensory transduction. TRPV4 is multimodally activated, e.g. by mechanical, thermal and tonicity stimuli, and products of the trpv4 gene have been detected in trigeminal ganglion sensory neurons and their peripheral projections, f/pyf'' mice, previously generated by the PI, have an elevated threshold for noxious mechanical and tonicity stimuli. The critical question remains unresolved whether and howTRPV4* trigeminal sensory neurons transduce such stimuli that elicit nociceptive behavior. Thus, the objective of this proposal is to investigate molecular and cellular mechanisms of frpv4-mediated transduction of noxious osmotic and mechanical stimuli in trigeminal sensory neurons by development of a novel mouse model. The hypothesis to be tested is whether trpv4 gene expression in trigeminal sensory neurons is critical for neurosensory transduction in response to noxious stimuli. Specifically, (1) primary cultures of trigeminal neurons from trpv4~*~ mice will be investigated by patch-clamp and Ca*+ imaging in response to tonicity and mechanical stimuli, modulated by activation of proteinase-activated receptor 2 (PAR-2), a physiologically relevant pro-algesic pathway, and for their "tonicity-tuning" of voltage-gated sodium channels (lNa) and capsaicin-response; (2) same as in Aim (1),but for dissociated sensory neurons from sensory-neuron-specific, inducible trpv4'~ mice; (3) sensdry-neuron- specific, inducible trpv4^~ mice will be generated to test the response of these mice to noxious osmotic stimuli and osmotic modulation of capsaicin stimuli of trigeminal peripherals, modulated by activation of PAR-2. For Aims (1)-(2), primary cultured neurons will be labeled by a genetically-encoded fluorescent reporter gene, expressed in a bacterialTartificial chromosometransgenic line of mice. For Aim (3), mice will be subjected to nocifensive behavioral assays and c-FOS studies of the brain stem trigeminal spinal nucleus in response to stimulation. Thus, the dependence of sensory transduction and transmission of noxious trigeminal stimuli on trpv4 will be deconstructed at the cellular level, clarifying the relative contribution of the trigeminal sensory neuron. Moreover, light will be shed on how trpv4 functions in "tonicity-tuning" of the capsaicin response and of iNa-. and on modulation of TRPV4 by PAR-2. Thus, our new mouse model will help us better understand how trigeminal ganglion neurons respond to noxious stimuli and which role the trpv4 gene plays in this process, hopefully opening up new avenues for treatment of trigeminally-mediated pain.