DRG神经元胞体N-型钙通道上调引起神经病理性疼痛的机制

It is well known that N-type calcium channel (Cav2.2) is densely expressed in the afferent terminals of DRG, regulates the release of various neurotransmitters, and plays a key role in sensory transmission, especially pain transmission. We recently found that IL-1beta upregulates Cav2.2 in the uninjured DRG neurons after peripheral nerve injury caused hyperalgesia, but the mechanism is not clear. The preliminary experiment of this project found that: the uninjured and injured DRGs subthreshold membrane potential oscillation increased significantly, and Cav2.2 blocker selectively inhibited the uninjured DRG subthreshold membrane potential oscillation; p-STAT3 increased the expression of uninjured DRG, intrathecal injection of JAK2/STAT3 signaling pathway inhibitors alleviated hyperalgesia. These results suggest that Cav2.2 expressed in DRG neurons is contributed to the increase of excitability, and IL-1beta regulates the expression of Cav2.2 through JAK2/STAT3 signaling pathway. In this project, by in vivo and in vitro method, we will explore: 1) To clarify the mechanism of Cav2.2 participating in subthreshold membrane potential oscillation and excitability increase of neurons; 2) To explore the mechanism of IL-1beta regulating Cav2.2 expression through JAK2/STAT3 signaling pathway. This project will reveal the role of Cav2.2 expressed in DRG neurons and the mechanisms involved in neuropathic pain, providing potential targets for clinical treatment.

众所周知N-型钙通道（Cav2.2）密集表达在DRG传入末梢调控神经递质释放，在感觉尤其是痛觉传递起关键作用。我们最近发现外周神经损伤后IL-1β上调未损伤DRG神经元Cav2.2引起痛觉过敏，但机制不清楚。预实验发现：未损伤和损伤DRG阈下膜电位震荡明显增加，使用Cav2.2阻断剂选择性的抑制未损伤DRG阈下膜电位震荡；p-STAT3在未损伤DRG表达上调，鞘内注射JAK2/STAT3信号通路的抑制剂缓解痛觉过敏。提示Cav2.2参与神经元兴奋性的增加，IL-1β通过JAK2/STAT3信号通路调控Cav2.2表达。本项目拟通过体内和体外实验：1)明确Cav2.2参与阈下膜电位震荡和引起神经元兴奋性增加的机制；2）探讨IL-1β通过JAK2/STAT3信号通路调控Cav2.2表达的机制。本项目将揭示在DRG神经元上表达的Cav2.2的作用和参与神经病理性疼痛的机制为临床治疗提供潜在靶点。

外周神经损伤经常引起神经病理性疼痛，通常在病灶去除或者损伤痊愈之后疼痛依旧存在，在临床上严重影响患者生活质量。N-型钙通道（Cav2.2）密集分布于DRG及脊髓背角的神经元上，在感觉尤其是痛觉传递起关键作用。在正常情况下体内的致炎细胞因子和抗炎细胞因子是处于一种动态稳定的状态，但是神经损伤之后这种动态稳定被打破，上调表达的致炎细胞因子在神经病理性疼痛的产生和维持中起了重要的作用。我们前期研究发现外周神经损伤后IL-1β上调未损伤DRG神经元上表达的Cav2.2引起痛觉过敏，但机制不清楚。本研究项目的实验结果发现，L5-SNL术后P-STAT3在未损伤L4 DRG显著上调表达而在损伤的L5 DRG下调表达。我们先给予DRG局部注射IL-1β的抑制剂Diacerein或P-STAT3的抑制剂S3I-20然后行L5-SNL术，发现明显减弱了P-STAT3在未损伤L4 DRG的上调表达并且缓解了机械痛觉异常。体外培养DRG神经元加入rrIL-1β也可以上调表达P-STAT3，这和在体动物实验结果相一致。此外，膜片钳实验记录显示，未损伤L4 DRG和损伤L5 DRG神经元中斜坡电流引起的动作电位的潜伏期明显短于假手术组而动作电位的频率明显高于假手术组，其中未损伤的DRG的电生理数据变化更明显。使用特异性Cav2.2阻断剂ω-CgTx可以显著增加未损伤的L4 DRG神经元动作电位的潜伏期，但对损伤的L5 DRG没有影响。以上结果提示我们外周神经损伤之后IL-1β/STAT3通路参与上调Cav2.2的表达，通过缩短神经元产生动作电位的潜伏期增加了神经元的兴奋性，有助于神经病理性疼痛的发生。