Mechanisms in primary nociceptors that drive ongoing activity and ongoing pain

项目摘要 该项目的长期目标是发现新的，高度针对性的方法来治疗正在进行的 通过定义驱动这种疼痛的初级伤害感受器中的持续活动（OA）的关键机制来研究疼痛。最近 研究发现OA在可能的伤害感受器中自发产生，并与持续的疼痛有关 脊髓损伤（SCI）后的神经功能损伤与所有三种电生理学改变有关，原则上， 促进OA。这些是静息膜电位（RMP）的去极化， 动作电位（AP），以及大的、瞬时的、去极化自发波动频率增加 （DSF）。两种与炎症相关的外源性介质，5-羟色胺（5-HT）和辣椒素（模拟 内源性TRPV 1激活剂）也促进OA，在很大程度上通过增强DSF。事实上， 大型DSFs的潜在机制。三个具体目标将测试有关DSF生成的假设， 增强，采用全细胞膜片记录，通过Ca 2+释放刺激，药理学和 转基因方法、体内记录和行为测试。目标1将定义离子电导和电池 信号传导（Ca 2+和cAMP）对大DSFs急性产生的贡献，利用了 5-HT，毛喉素和辣椒素快速刺激大的DSF，使用幼稚大鼠和转基因小鼠。的 重点将包括HCN通道、T型Ca 2+通道和Nav1.8通道。将特别注意 TRPC 4/5通道，对OA很重要，具有独特的特性， 大的DSFs。目标2将定义促进大DSF产生的离子电导和细胞信号， 慢性SCI和亚急性外周炎症模型（后爪注射完全弗氏佐剂 - CFA）。在目标1中发现的对大型DSFs重要的通道将被测试改变的贡献， 每个模型中的表达式。预计促进OA的改变在这些模型中是共享的（因此， 潜在地驱动多种形式的持续疼痛）。目标3将检验联合干预的预测 选择性地阻断大的DSF和提高AP阈值将减少持续的疼痛。新型镇痛 将测试一种策略，该策略将阻止大DSF生成的药物（TRPC 4/5阻滞剂）与 选择性提高伤害感受器AP阈值的药物（Nav1.8阻滞剂）。该组合应 有效地抑制伤害感受器OA和随之而来的持续疼痛，剂量低于观察任何 对任何一种药物单独给药的持续疼痛的影响。该预测将在记录的C-纤维OA上进行体内测试。 从麻醉大鼠的背根和持续疼痛的SCI大鼠和大鼠和小鼠CFA模型。这 有针对性的方法可以为治疗严重持续疼痛的新疗法奠定基础， 副作用，并提供阿片类药物的替代品及其伴随的风险。

Project Summary The long-term objective of this project is to discover novel, highly targeted approaches for treating ongoing pain by defining critical mechanisms of ongoing activity (OA) in primary nociceptors that drive this pain. Recent discoveries revealed that the OA generated spontaneously in probable nociceptors and linked to ongoing pain after spinal cord injury (SCI) is associated with all three electrophysiological alterations that, in principle, can promote OA. These are depolarization of resting membrane potential (RMP), reduced voltage threshold for action potentials (APs), and increased frequency of large, transient, depolarizing spontaneous fluctuations (DSFs). Two extrinsic mediators related to inflammation, serotonin (5-HT) and capsaicin (mimicking endogenous TRPV1 activators), also promote OA, in large part by enhancing DSFs. Virtually nothing is known about mechanisms underlying large DSFs. Three specific aims will test hypotheses about DSF generation and potentiation, employing whole cell patch recording, stimulation by Ca2+ uncaging, pharmacological and transgenic approaches, in vivo recording, and behavioral tests. Aim 1 will define ion conductance and cell signaling (Ca2+ and cAMP) contributions to the acute generation of large DSFs, taking advantage of the ability of 5-HT, forskolin, and capsaicin to rapidly stimulate large DSFs, using naïve rats and transgenic mice. The focus will include HCN channels, T-type Ca2+ channels, and Nav1.8 channels. Special attention will be paid to TRPC4/5 channels, which are important for OA and have unusual properties that account for unique features of large DSFs. Aim 2 will define ion conductances and cell signals that promote large DSF generation in chronic SCI and in a subacute peripheral inflammation model (hindpaw injection of complete Freund's adjuvant - CFA). The channels found in Aim 1 to be important for large DSFs will be tested for altered contributions and expression in each model. Alterations promoting OA are predicted to be shared in these models (and thus to potentially drive many forms of ongoing pain). Aim 3 will test the prediction that combined interventions selectively blocking large DSFs and elevating AP threshold will reduce ongoing pain. A novel analgesic strategy will be tested, which combines a drug that prevents large DSF generation (a TRPC4/5 blocker) with a drug that selectively elevates AP threshold in nociceptors (a Nav1.8 blocker). The combination should efficiently suppress nociceptor OA and consequent ongoing pain at doses lower than required to observe any effect on ongoing pain from either drug alone. This prediction will be tested in vivo both on C-fiber OA recorded from dorsal roots of anesthetized rats and on ongoing pain in SCI rats and in rat and mouse CFA models. This targeted approach could lay the foundation for new treatments for severe ongoing pain that have relatively few side effects and provide an alternative to opioids, with their attendant risks.