Lipid kinase regulation of pain signaling and sensitization

项目总结 治疗慢性疼痛需要新的方法，特别是在现有的止痛药已经 严重的副作用，在治疗炎症性疼痛和神经病理性疼痛时并不总是有效的-这两种 人类最常见的慢性疼痛。炎症和神经损伤导致释放一种 复杂的化学混合物，通过分子上不同的伤害性(产生疼痛)受体发出信号。 这些受体的激活增加了伤害性背根神经节的兴奋性和敏感性。 和三叉神经节神经元。不幸的是，到目前为止，阻止单个伤害性感受器的努力取得了进展 未能找到治疗慢性疼痛的有效方法。在这里，我们提出了一种创新的方法来减少 疼痛过敏，绕过了这个与受体多样性相关的长期问题。我们的 方法是基于选择性地降低脂质第二信使磷脂酰肌醇4，5- 背根神经节神经元中的二磷酸(PIP2)。大多数伤害性感受器需要PIP2来启动下游 发信号。此外，许多检测伤害性刺激的Trp通道和调节膜的离子通道 兴奋性需要PIP2才能活动。因此，PIP2处于不同受体离子的关键汇聚点 促进和维持慢性疼痛的通道和信号通路。在对小鼠的初步研究中，我们 发现了一种在背根节神经元中产生至少50%PIP2的脂蛋白激酶。此外，停用这一功能 脂激酶可显著降低对炎性介质的伤害敏感度。基于我们的 初步数据显示，我们推测这种脂激酶通过PIP2依赖的机制来调节 背根神经节神经元的伤害性感受器信号与体内痛敏作用。为了检验这一假设，我们 威尔：1.用小鼠评价该脂蛋白在体内调节伤害性敏感化的程度 急性、慢性和自发性疼痛模型，包括炎症性疼痛模型和神经病理性疼痛模型。 我们将使用一种创新的遗传方法来降低激酶的活性。这种方法选择性地减少了 背根神经节中PIP2的浓度，但不影响处理疼痛信号的其他组织中的PIP2浓度。 我们将通过生化救援实验进一步评估PIP2的依赖性。2.评估以下方面的程度 这种激酶通过不同的伤害性感受器调节信号，包括G蛋白偶联 受体，一种酪氨酸激酶受体，以及检测有害刺激的Trp通道。3.利用新的 条件性基因敲除小鼠仅在成人感觉神经元中诱导删除该激酶并评估 该激酶调节炎症性疼痛和神经病理性疼痛的启动和维持的程度。 我们将是第一个严格研究这种激酶在慢性疼痛中的重要性的人。我们的 初步数据表明，这种脂激酶是疼痛信号和敏化的主要调节器。

PROJECT SUMMARY New approaches for treating chronic pain are needed, particularly since existing analgesics have serious side effects and are not always effective at treating inflammatory pain and neuropathic pain-the two most common forms of chronic pain in humans. Inflammation and nerve injury lead to the release of a complex mix of chemicals that signal through molecularly diverse pronociceptive (pain-producing) receptors. Activation of these receptors increases the excitability and sensitivity of nociceptive dorsal root ganglia (DRG) and trigeminal ganglia neurons. Unfortunately, efforts to block individual pronociceptive receptors have so far failed to produce effective treatments for chronic pain. Here, we propose an innovative approach to reduce pain hypersensitivity that bypasses this long-standing problem associated with receptor diversity. Our approach is based on selectively reducing the level of the lipid second messenger phosphatidylinositol 4,5- bisphosphate (PIP2) in DRG neurons. Most pronociceptive receptors require PIP2 to initiate downstream signaling. Moreover, many TRP channels that detect noxious stimuli and ion channels that regulate membrane excitability require PIP2 for activity. PIP2 thus sits at a key convergence point for diverse receptors, ion channels and signaling pathways that promote and maintain chronic pain. In preliminary studies with mice, we identified a lipid kinase that generates at least 50% of all PIP2 in DRG neurons. Moreover, inactivation of this lipid kinase profoundly reduced nociceptive sensitization in response to an inflammatory agent. Based on our preliminary data, we hypothesize that this lipid kinase acts through PIP2 dependent mechanisms to regulate pronociceptive receptor signaling in DRG neurons and pain sensitization in vivo. To test this hypothesis we will: 1. Evaluate the extent to which this lipid kinase regulates nociceptive sensitization in vivo, using mouse models of acute, chronic and spontaneous pain, including models of inflammatory pain and neuropathic pain. We will use an innovative genetic approach to knock-down kinase activity. This approach selectively reduces PIP2 concentration in DRG but does not affect PIP2 concentration in other tissues that process pain signals. We will further evaluate PIP2-dependence using biochemical rescue experiments. 2. Evaluate the extent to which this kinase regulates signaling through diverse pronociceptive receptors, including G protein-coupled receptors, a tyrosine kinase receptor, and TRP channels that detect noxious stimuli. 3. Utilize a new conditional knockout mouse to inducibly delete this kinase only in sensory neurons of adults and to evaluate the extent to which this kinase regulates initiation and maintenance of inflammatory pain and neuropathic pain. We will be the first to rigorously study the importance of this kinase in the setting of chronic pain. Our preliminary data suggest this lipid kinase is a master regulator of pain signaling and sensitization.