缝隙连接蛋白Cx36介导钙通道α2δ1亚基参与三叉神经病理痛机制研究

三叉神经损伤所致疼痛剧烈，易扩散。在损伤和邻近未损伤神经支配区域可分别检测到原发痛和继发痛。钙通道α2δ1亚基（Cavα2δ1）可能参与三叉神经病理痛的发生，机制尚不明确。神经元特异表达的缝隙连接蛋白connexin 36（Cx36），介导神经元间快速信息传递，可能是Cavα2δ1参与三叉神经病理痛产生及扩散的下游信号分子。Cx36在三叉神经节广泛表达，功能知之甚少。本项目采用三叉神经分支皮下注射病毒干扰或过表达目的蛋白、行为学测试、电生理（钙成像、膜片钳、dual patch）、组织透明化联合免疫组化、免疫电镜及western blot等多种技术手段，验证以下假说：Cavα2δ1以PKC和CaMKⅡ依赖的方式调控Cx36表达和功能，Ca2+由损伤神经支配的神经元向邻近神经元扩散，激活Ras/ERK/CREB信号通路，致疼痛产生及扩散。旨在为三叉神经病理痛的治疗提供新思路和新靶点。

Orofacial pain is a debilitating disorder that diminishes the quality of life and seriously inhibits the health of patients. Trigeminal neuropathic pain in the injured area (primary hyperalgesia) also spreads to wide orofacial areas (secondary hyperalgesia) and is a serious clinical problem. Chronic constriction injury of the infraorbital nerve (CCI-ION) caused orofacial hypersensitivity that correlated with voltage-gated calcium channel α2δ1 subunit (Cavα2δ1) up-regulation in trigeminal ganglion (TG) neurons and the subnucleus caudalis (Vc). Blocking Caα2δ1 with antagonists led to a reversal of orofacial hypersensitivity, supporting an important role of Caα2δ1 in orofacial pain processing. However, the corresponding mechanisms are still unclear..Connexin 36 (Cx36), a gap junction protein specifically expressed by neurons, rapidly transmits information between neurons, possibly functions as the downstream molecule of Cavα2δ1 contributing to the induction and spreading of trigeminal neuropathic pain. Cx36 is widely expressed in trigeminal ganglion neurons and its function is poorly understood. We proposed the following hypothesis: Cavα2δ1 modulates the expression and function of Cx36 in a PKC and CaMKII dependent manner, which results to the transmission of Ca2+ rapidly from the neuron inervated by the injured nerve to adjacent neurons, and activates the downstream Ras/ERK/CREB signaling pathway. All of above leads to the sensitization and plastic changes of TG neurons, which underlies the induction of primary and secondary trigeminal neuropathic pain..To test this hypothesis, the present study intervene the function of Cavα2δ1 and the downstream molecules by specifically knocking down their expression through subcutaneous injection of virus in the injured area and uninjured area. Also some other methods including pain behavior testing, electrophysiological recording, transparent tissue technique combined with immunohistochemistry staining, Western blot, immune electron microscopy were employed. The present study aims to solve the following problem: Whether Cavα2δ1 mediates the induction and spreading of trigeminal neuropathic pain by modulating the expression and function of Cx36 and the activation of the downstream Ras/ERK/CREB signaling pathway? The present study aims to elucidate the role of Cavα2δ1 and downstream Cx36 in the development of trigeminal neuropathic pain and provide new sight for the treatment.