ACC-vlPAG抑制性突触联系参与重复电针镇痛的机制研究

Acupuncture has become the replacement therapy for the treatment of neuropathic pain. Periaqueductal gray (PAG), which involved in acupuncture analgesia, is the core of descending inhibitory system. Anterior cingulate cortex (ACC) is the starting point of the descending inhibitory system, and establishes inhibitory synaptic connections with the ventrolateral region of PAG (vlPAG). Our early studies results have confirmed that repeated electroacupuncture (EA) could decrease the frequency and amplitude of mIPSC, and the amplitude of IPSC which were both mediated by GABAAR in vlPAG significantly. It was suggested that repeated EA could decrease the inhibitory synaptic function of ACC-vlPAG pathway. Adenylate cyclase 1 (AC1) participated in central sensitization of ACC. The applicant's studies also found that the excitabilities of ACC layer V neurons were downregulated obviously by repeated EA, and the cumulative analgesic effect of repeated EA was also inhibited by the AC1 agonist. Accordingly, we propose a hypothesis that ACC AC1 activity was reduced by repeated EA, which resulting in the inhibitory synaptic function of ACC-vlPAG pathway decrease, and enhancing the activity of the descending inhibitory system, leading to the analgesic effect. Optogenetics, electrophysiological methods and molecular biology technology were used to investigate the effect of repeated EA on ACC neuron excitability and inhibitory synaptic transmission of the ACC-vlPAG pathway. The new central mechanism of repeated EA on neuropathic pain will be elucidated, which has important theoretical significance and clinical significance.

针刺已成为治疗神经病理性痛的替代疗法，导水管周围灰质(PAG)是下行抑制系统的核心，参与针刺镇痛。前扣带回(ACC)是下行抑制系统的起点，与PAG腹外侧区（vlPAG）形成抑制性突触联系。申请者预实验证实重复电针显著降低vlPAG GABAAR介导的mIPSC频率和幅度及IPSC幅度，提示重复电针减弱ACC-vlPAG抑制性突触联系。腺苷酸环化酶1 (AC1)参与ACC中枢敏化，申请者证实重复电针降低ACC神经元兴奋性，其镇痛效应能被AC1激动剂抑制。据此我们提出假说：重复电针通过降低ACC AC1活性，导致ACC-vlPAG抑制性突触联系功能减弱，增强下行抑制系统活性，发挥镇痛作用。本研究拟采用光遗传学和电生理等功能和分子生物学技术探讨重复电针对ACC-vlPAG通路上ACC神经元兴奋性和vlPAG抑制性传递效率的影响，将阐明重复电针抑制神经病理性痛的新机制，有重要的理论意义和临床意义。

项目的背景.针刺具有镇痛效果好，易于实施，不良反应少等优点，人们将其视为神经病理性疼痛（NP）治疗的替代疗法。单次针刺的镇痛作用具有衰减性和延迟性特征。这些单次针刺的镇痛特点远不能满足人们治疗NP的初衷。文献报道将单次EA变为重复EA后，发现针刺镇痛作用加强，而且出现累积镇痛效应，但是重复针刺产生累积镇痛效应的中枢机制尚不明确。PAG是下行抑制系统的核心，前扣带回（anterior cingulate cortex, ACC）是下行抑制系统的起点，PAG腹外侧区（vlPAG）与ACC形成突触联系，调控下行抑制系统。预实验发现重复EA能明显抑制ACC V层锥体神经元的兴奋性，并且其累积镇痛效应也被AC1的兴奋剂阻断。提示重复EA通过降低ACC锥体神经元的兴奋性可能导致ACC-vlPAG的突触联系减弱。根据前期研究和预实验提出：重复电针通过降低ACC活性，导致ACC-vlPAG突触联系功能减弱，发挥镇痛作用。..主要研究内容.① ACC区域AC1对REA产生的累积镇痛的影响.② REA产生的累积镇痛时ACC AMPAR表达变化.③ REA产生的累积镇痛时ACC-vlPAG突触功能改变..重要结果.在CCI模型上，对ST36和GB34穴位重复电针刺激，表现累积镇痛效应。在累积镇痛的动物模型上发现ACC-vlPAG通路上ACC锥体神经元的兴奋性降低，LTP幅度降低；vlPAG抑制性突触传递功能减弱。这些改变与REA后ACC AMPAR受体表达水平降低有关。..关键数据. CCI模型，对ST36和GB34穴位重复电针刺激显著提高损伤侧机械缩足阈值（1.24±0.53 g 提高至6.78±0.47 g），明显降低ACC-vlPAG通路上ACC锥体神经元的兴奋性（基强度：221±17.5 pA增大至265±16.28 pA；阈值：-32±1.79 mV增大至-35±1.6 mV）和LTP幅值（1.96±0.12减小至1.50±0.11），影响vlPAG抑制性突触传递（mIPSC幅值从10.5±2.3 pA减小至6.7±1.86 pA）。..科学意义.阐明重复电针产生累积镇痛效应的中枢机制，既重复EA减弱ACC-vlPAG通路中ACC投射神经元兴奋性和突触可塑性变化和抑制vlPAG的抑制性传递介导重复电针的累积镇痛效应。