前扣带回皮层后段深层（Cg2）谷氨酸能神经元在痛觉调制中的作用及环路机制研究

There are many challenges in the clinical management of pain, especially of neuropathic pain. More complete understanding of the structure and function of pain modulation system is of significance to the development of new analgesic targets and strategy. Anterior cingulate cortex (ACC) is activated at the presence of pain, but it’s roles in pain modulation are under debate, largely due to the heterogeneity of previous studies. Our pilot study revealed that selective activation of glutamatergic neurons in deeper layer (Cg2) of the caudal part of ACC (cACC) by optostimulation increased the nociceptive threshold to mechanical stimulation and inhibited the pain responses in the formalin test in mice. Of interest, this treatment seemed to attenuate sensory component of neuropathic pain following partial sciatic nerve ligation injury. Antegrade tracing by adeno-associated virus found that these neurons emit projections to certain subregions of zona incerta (ZI) and periaqueductal gray matters (PAG). In order to elucidate the roles of these neurons in pain modulation and the related neural circuitry, the present study plans to investigate the influence of selective manipulation of the activity of glutamatergic neurons in Cg2 of cACC on nociceptive pain and sensory and aversive components of neuropathic pain, by using optogenetics combining methods such as behavioral tests, electrophysiology, pharmacology and so on. Moreover, the functional relationship between the glutamatergic projections of these neurons and neurons in the ZI and PAG will be studied with multidisciplinary methodology, and the roles of these projections in pain modulation, including of nociceptive pain and neuropathic pain, will be examined systemically. This study will provide more knowledge about the pain modulation system under different painful conditions and will be helpful for the findings of new analgesic targets.

目前临床对疼痛，特别是对神经病理性疼痛的管理还存在很多挑战。充分认识痛觉调制系统的结构与功能有助于发现新的镇痛靶点。前扣带回皮层（ACC）在疼痛时被激活，但在痛觉调制中的作用尚未明确。课题组预实验发现，用光遗传学方法选择性激活ACC后段深层（Cg2）的谷氨酸能神经元能提高动物对机械刺激的痛觉感受阈值，减轻福尔马林引起的疼痛反应，对神经病理性疼痛的感觉成分也有抑制作用。病毒顺行追踪发现这些神经元投射到未定带（ZI）和中脑导水管中央灰质（PAG）的特定区域。本研究拟充分利用光遗传学技术，结合电生理学、药理学、组织化学、行为学等方法，研究和阐明选择性激活或抑制ACC后段Cg2谷氨酸能神经元对急性疼痛和神经病理性疼痛的感觉和情绪成分的调制作用，以及至ZI和PAG 的神经环路在其中的作用机制。本研究将为深入理解不同疼痛状态下ACC在痛觉调制中的作用和环路基础并发现新的药物镇痛靶点提供可靠的实验证据。

神经病理性疼痛是常见的一种慢性疾病。现有的神经病理性疼痛治疗药物只对部分患者有效，且常伴随难以耐受的副作用。深入解析神经病理性疼痛的发病机制，有助于发现有效的干预手段和药物作用靶点。神经损伤后多个脑区的结构和功能发生改变，但在神经病理性疼痛中的作用尚待研究。本研究利用多种技术手段，系统研究了中扣带皮层Cg2（midcingulate cortex area 2, MCC cg2）的谷氨酸能神经元（MCC cg2Glu）在神经病理性疼痛中的作用及环路机制。研究发现疼痛刺激可抑制大多数MCC cg2Glu，表明MCC cg2Glu与疼痛处理有关。用光遗传学方法特异性激活MCC Cg2Glu能抑制疼痛，光抑制则促进疼痛，提示MCC cg2Glu是内源性疼痛调制系统的组成部分。MCC cg2Glu到未定带（zona incerta, ZI）和背外侧PAG（DLPAG）有集中投射，光激活MCC cg2Glu投射到ZI而非DLPAG的纤维末梢具有镇痛作用，且该作用可以被提前在ZI局部注射谷氨酸能NMDA/AMPA受体混合拮抗剂逆转。MCC Cg2Glu与ZI 的γ-氨基丁酸能神经元（ZIGABA）形成单突触联系，且光激活MCC cg2Glu到ZI的投射末梢可以激活ZI的神经元。抑制ZIGABA能够拮抗光激活MCC cg2到ZI的谷氨酸能纤维末梢的镇痛作用。以上结果表明MCC cg2Glu的镇痛作用主要是通过激活ZIGABA实现的。神经损伤后MCC cg2Glu的神经元活性降低。尽管短暂激活MCC cg2到ZIGABA神经环路能够缓解与自发持续性疼痛相关的厌恶情绪，但只有连续激活这条神经环路才能减轻神经病理性异常疼痛，提示MCC cg2Glu的内源性镇痛作用减弱。相反，MCC cg1的谷氨酸能神经元具有易化疼痛的作用。神经损伤后神经元活性增强，只有连续抑制其活性才能缓解神经病理性异常疼痛。本研究发现MCC cg1Glu及其到ZI的神经环路参与内源性镇痛，其活性的下降参与了神经病理性疼痛的发生，连续激活MCC cg2Glu到ZIGABA的神经环路可恢复其内源性镇痛作用，减轻神经病理性疼痛。因此，用药物或其他手段选择性激活这条神经环路可以作为神经病理性疼痛的潜在治疗策略。