T型钙通道在大鼠脊髓背角II层神经元中的特异性表达和功能研究

The incidence of neuropathic pain is high and is hard to manage. It is necessary to unravel the underlying mechanism and find new treatment target for neuropathic pain. T-type calcium channel (Cav3), also named low-voltage-activated Ca2+ channel, plays an important role in modulation of neuronal excitability. Recent studies indicate that abnormal activation of T-type calcium channels are involved in the development and maintenance of neuropathic pain. However, the exact contribution of T-type calcium channels in spinal dorsal horn remains unclear. Substantia gelatinosa (SG) in superficial spinal dorsal horn is the primary integrate center of pain pathway. Our previous studies found that T-type calcium channels is not only development-dependent but also cell-type specific expressed in SG. Moreover, the larger the current induced by T-type calcium channel (IT), the faster the spike frequency. Furthermore, another low-voltage-activated HCN channel antagonist increased IT. Thus, we hypothesized that Cav channels have distinct distribution and function in SG neurons. Here, we first investigate the specific cellular distribution of Cav channels in SG. Then, we identify the current kinetics of different Cav subtypes and the effect on SG neuronal excitability. Finally, we study the expression and function of Cav channels in pathologic pain animal models. These observations may provide new ideas about the central mechanisms and new target for the treatment of neuropathic pain.

神经病理性痛发生率高且临床治疗颇为棘手，探索神经病理性痛的机制及寻找治疗的新靶点是亟待解决的现实问题。T型钙通道（Cav3）又称为低电压激活钙通道，对神经元兴奋性具有重要的作用。近年发现Cav3异常活化与神经病理性痛的发生有关。脊髓背角浅层的SG区是痛信号的初级整合中枢。我们预实验发现，SG区的Cav3具有发育和细胞特异性；而且，T型钙电流（IT）越大，SG神经元的动作电位放电频率越高；此外，另一种低电压激活的HCN通道的阻断剂可增大IT。据此我们提出Cav3亚型在脊髓背角浅层具有特异性分布和功能，并且受HCN通道调控的假说。围绕该假说，我们首先明确Cav3亚型在脊髓背角的表达。其次，进一步阐明不同Cav3亚型的电流动力学特性及对SG神经元兴奋性的作用机制。最后，研究Cav3在神经病理性痛动物模型中的作用机制。以上研究结果可为神经病理性痛的中枢机制提供新的研究思路和药物治疗的新靶标。

神经病理性疼痛由神经损伤引起，通常起源于初级传入纤维或脊髓背角浅层的神经环路，脊髓背角II层又称胶状质区（Substantia gelatinosa, SG），在痛信息的传入和整合中发挥重要的作用。T型钙通道（Cav3）又称为低电压激活钙通道，对神经元兴奋性具有重要的调控作用。本项目重点研究Cav3亚型在脊髓背角的特异性分布特点，对SG 神经元的膜特性、突触传递及动作电位的作用机制；并深入研究Cav3对神经病理性疼痛发生的影响。在本项目资助下，我们发现T型钙通道三种亚型Cav3.1、Cav3.2、Cav3.3在大鼠脊髓背角均有表达，且Cav3.2和Cav3.3的表达无显著差异，而Cav3.1的表达显著低于Cav3.2和Cav3.3。SG神经元中，44.5%（178/400）表达T型钙电流，幅值大小从6.1 pA到415.4 pA不等，平均幅值为84.8 ± 5.3 pA，大多数表达T型钙电流的神经元是Tonic放电的神经元（76.5%）和具有islet形态的神经元（41.2%）。部分坐骨神经结扎（PSNL）术后14天，大鼠SDH浅层Cav3.2 mRNA及蛋白的表达明显增加，而Cav3.1及Cav3.3的表达无明显变化。连续鞘内或间断腹腔注射Cav3通道阻滞剂可稳定而持续地缓解PSNL引起的机械痛敏和热痛敏，至少持续到术后14天。综上，本研究揭示了T型钙通道在大鼠脊髓背角SG神经元上的特异性分布及其对SG神经元功能的调控作用，神经病理性疼痛促进了SDH浅层Cav3.2的功能表达上调，这可能是PSNL诱发机械痛敏的潜在中枢机制，为脊髓Cav3.2通道作为疼痛的治疗靶点提供了新的理论依据。本项目资助下，共发表署项目号通讯作者论文14篇，其中SCI收录5篇，培养博士研究生3名，硕士生18名，青年教师1名。获江西省优秀硕士论文2人次。