Spectroscopic analyses of TRPV1 during gating

瞬时受体电位香草酸1（TRPV 1）是细胞内必需的多模式离子通道， 检测有害刺激的潜在机制。TRPV 1被热、质子、辣椒素激活， 动物毒素，并由促痛觉炎症剂调节（例如，缓激肽、生物活性脂质）产生 对组织损伤的反应。我们的长期目标是阐明多模式离子通道在感觉中的作用 神经元兴奋和它们促进炎性疼痛的机制。我们的理由是， 拟议的研究是，TRPV 1质子和热依赖性门控的更深入的机制理解 将极大地促进改善TRPV 1介导的炎性疼痛的策略的发展，而不 扰乱了正常的感官生理虽然TRPV 1的功能和结构表征揭示了 辣椒素和毒素激活的机制，这两个主要的内源性 激活剂、质子和热、触发门控仍然在很大程度上未知。此外，细胞内TRPV 1 C 末端是调节刺激敏感性的关键调节位点。然而，任何潜在的变构相互作用 与质膜的区域或假定的接触仍有待探索。我们认为， 需要光谱方法来完全确定负责的变构构象变化 TRPV 1激活和描绘C-末端/膜相互作用。为此，我们将 电生理分析以及电子顺磁共振光谱实验， 封闭和开放的国家。利用这些数据，我们将描述TRPV 1经历的构象变化 在质子和热依赖门控期间。我们将追求两个具体目标：1）确定动态 TRPV 1在质子和热活化过程中的构象重排，以及2）探索相互作用 TRPV 1 C末端结构域和质膜之间。所提出的研究是有意义的 因为预期其在治疗与广泛的疼痛相关的疼痛中具有广泛的转化重要性， 一系列的病理生理条件。

The transient receptor potential vanilloid 1 (TRPV1) is a polymodal ion channel essential to the cellular mechanism underlying the detection of noxious stimuli. TRPV1 is activated by heat, protons, capsaicin, and animal toxins, and is modulated by proalgesic inflammatory agents (e.g., bradykinin, bioactive lipids) produced in response to tissue injury. Our long-term goal is to delineate the roles of polymodal ion channels in sensory neuron excitation and the mechanisms by which they contribute to inflammatory pain. The rationale for our proposed research is that a deeper mechanistic understanding of TRPV1 proton- and heat-dependent gating would greatly facilitate the development of strategies to ameliorate TRPV1-mediated inflammatory pain, without disrupting normal sensory physiology. While functional and structural characterization of TRPV1 have shed light on the mechanisms of capsaicin and toxin activation, the processes whereby the two main endogenous activators, protons and heat, trigger gating remain largely unknown. Moreover, the intracellular TRPV1 C terminus is a key regulatory site for regulating stimulus sensitivity. However, any potential allosteric interacting regions or putative contacts with the plasma membrane have yet remain to be explored. It is our contention that spectroscopic approaches are needed to fully define the allosteric conformational changes responsible for TRPV1 activation and to depict the C-terminal/membrane interaction. To this end, we will carry out electrophysiological analyses together with electron paramagnetic resonance spectroscopy experiments in both closed and open states. With these data, we will depict the conformational changes that TRPV1 undergoes during proton- and heat-dependent gating. We will pursue two Specific Aims: 1) Determine the dynamic conformational rearrangements of TRPV1 during proton and heat activation, and 2) Explore the interaction between the TRPV1 C-terminal domain and the plasma membrane. The proposed research is significant because it is expected to have broad translational importance in the treatment of pain associated with a wide range of pathophysiological conditions.