嘌呤信号介导艾灸镇痛穴位局部作用机制

Purinergic signaling has being a new field and hot spot involved in acupuncture-induced analgesia. In previous study we have found that ATP other than adenosine which was reported in Nature Neuroscience (2010) increased at the acupoint site Zusanli during moxibustion-induced analgesia. Based on this new finding, we proposes a novel hypothesis that ATP and its purinergic receptors at the acupoint location would be likely to involve in moxibustion-induced analgesia which will different from that in acupuncture-induced analgesia. Guided by this hypothesis, we will plan to construct inflammation and neuropathic pain mouse model and then give them intervention with moxibustion, acupuncture, agonist and antagonists of purinergic receptors, etc. In the proposed study, micodialysis, high performance liquid chromatography, immunofluorescence, western blot, and real-time PCR, etc will be applied to monitor and analyze the dynamic changes of ATP, ADP,AMP, adenosine and P1, P2 receptors at moxibustion site and reveal the rules of their dynamic changes. After this systematic and comprehensive study, we hope to determine in the acupoint location which purines out of ATP, ADP, AMP and adenosine are involved in acupuncture and moxibustion-induced analgesia. and explore purinergic mechanisms underlying Moxibustion and acupuncture-induced analgesia. In the end, it also will provide more scientific evidences for explaining that what is the difference between moxibustion and acupuncture analgesia by means of modern science and technology.

嘌呤信号是近年来阐释针灸镇痛原理新的方向和热点。在前期研究中我们发现：艾灸镇痛穴位局部嘌呤信号以ATP升高为主，而非2010年Nature neuroscience 报道的针刺镇痛穴位局部以腺苷升高为主。基于这一新的发现，我们提出"艾灸镇痛穴位局部嘌呤信号参与机制和针刺镇痛不同，主要以ATP为主的嘌呤信号介导"研究假说。围绕这一新的发现和工作假说，我们拟建立炎症痛和神经病理痛模型，结合穴位局部微透析、高效液相色谱、受体激动和阻断、免疫荧光、Western Blot、荧光定量PCR等方法技术，全面解析艾灸镇痛穴位局部嘌呤信号物质（ATP、ADP、AMP和腺苷）动态变化特征和相应的嘌呤信号受体（P1、P2）变化规律；并比较分析穴位局部嘌呤信号在针刺、艾灸镇痛中的差异；为回答穴位局部嘌呤信号在艾灸镇痛中的作用及"针所不为、灸之所宜"理论观点提供科学依据。

嘌呤信号是近年来阐释针灸镇痛原理新的方向和热点。项目组首先运用福尔马林疼痛模型、完全弗氏佐剂（CFA）注射诱导形成的炎症痛模型和神经病理痛模型明确艾灸镇痛效应的基础上，重点以CFA诱导形成的炎症痛动物模型，给予“足三里”艾灸干预，并结合穴位局部微透析、高效液相色谱、受体激动和阻断等方法技术，解析介导艾灸镇痛穴位局部嘌呤信号物质和嘌呤信号受体的动态变化特征；比较分析穴位局部嘌呤信号在针刺、艾灸镇痛中的差异。研究结果表明:（1）在艾灸镇痛过程中，穴位局部嘌呤信号ATP升高；穴位局部给予ATPase促进ATP分解后，艾灸镇痛效应减弱；抑制穴位局部ATP分解，艾灸镇痛作用加强。表明穴位局部嘌呤信号ATP是艾灸镇痛穴位局部机制的重要因素。（2）穴位局部给予嘌呤信号P2X3受体激动剂后，艾灸镇痛作用加强；给予嘌呤信号P2X3、P2X7受体拮抗剂后，艾灸镇痛作用减弱。表明嘌呤信号P2X3和P2X7受体主要是介导艾灸热传导后镇痛效应，而非直接镇痛效应。（3）穴位局部给予嘌呤信号A1受体激动剂或A2a受体拮抗剂后，艾灸镇痛作用加强；给予嘌呤信号A1受体拮抗剂或A2a受体激动剂后，艾灸镇痛作用减弱。表明穴位局部嘌呤信号P1受体的作用不仅介导针刺镇痛，也是艾灸镇痛穴位局部机制的一个重要环节。（4）生理状态下，穴位局部ATP、ADO对针刺、艾灸的反应体现为峰值时间差异：针刺捻转在30min显著升高，维持至60min；艾灸有延后效应，在60min显著升高。病理状态下，穴位局部ATP、ADO对针刺、艾灸的反应体现为升高幅度差异：针刺、艾灸都能使CFA诱导的炎症痛模型大鼠“足三里”穴位局部ATP 、ADO在30min显著升高，但针刺维持至60min，而且针刺捻转比艾灸以及不捻转升高穴位局部ATP 、ADO水平更加显著。研究结果从穴位局部嘌呤信号反应差异角度为“针所不为、灸之所宜”理论观点提供了部分科学依据。