基于NMDAR1/cGMP通路探究“三法三穴”对轻度慢性压迫性神经损伤大鼠脊髓背角的镇痛启动机制  

Exploring the analgesic initiation mechanism of "three-manipulations and three-acupoints" on the spinal dorsal horn of rats with minor chronic constriction injury based on the NMDAR1/cGMP pathway

作　　者：杨震杰 萨出拉 于天源[1] 陈金平 张润龙 张英琦 张汉钰 孙佳伟 刘家玥 YANG Zhenjie;SA Chula;YU Tianyuan;CHEN Jinping;ZHANG Runlong;ZHANG Yingqi;ZHANG Hanyu;SUN Jiawei;LIU Jiayue(School of Acupuncture-Moxibustion and Tuina,Beijing University of Chinese Medicine,Beijing 102488,China)

机构地区：[1]北京中医药大学针灸推拿学院,北京102488

出　　处：《北京中医药大学学报》2024年第7期1017-1024,共8页Journal of Beijing University of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(No.82274675,No.82074573);北京市自然科学基金项目(No.7232278)。

摘　　要：目的 探究“三法三穴”推拿手法对轻度慢性压迫性神经损伤(minor CCI)模型大鼠的镇痛启动机制。方法 35只SD大鼠按照随机数字表法分为5组,即正常组、假手术组、模型组、推拿组、推拿+MK-801组。模型组、推拿组和推拿+MK-801组结扎右侧坐骨神经干建立minor CCI大鼠模型;假手术组仅暴露右侧坐骨神经,不予结扎;正常组不予任何操作。正常组不予任何干预措施;模型组、假手术组造模后第7天予抓握束缚9 min;推拿组造模后第7天进行1次三法(点法、拨法、揉法)三穴(右侧“殷门”“承山”“阳陵泉”)干预,每法每穴干预1 min,共9 min;推拿+MK-801组造模后第5~7天鞘内注射MK-801,每次6μg(10μL),每日1次,最后一次鞘内注射30 min后再进行推拿干预,推拿具体操作同推拿组。造模前、造模后、干预后检测各组大鼠冷敏阈值(CST)和机械缩足反射阈值(MWT);干预后,免疫组织化学法检测腰4~6节段脊髓背角中环磷酸鸟苷(cGMP)蛋白阳性表达,蛋白质印迹法检测腰4~6节段脊髓背角中N-甲基-D-天冬氨酸受体1(NMDAR1)、神经源型一氧化氮合酶(nNOS)、可溶性鸟苷酸环化酶β(sGCβ)、蛋白激酶G1(PKG1)蛋白表达,实时荧光PCR法检测腰4~6节段脊髓背角中NMDAR1、nNOS、sGCβ、cGMP、PKG1 mRNA表达。结果 与正常组、假手术组比较,造模后模型组、推拿组和推拿+MK-801组CST升高,MWT降低(均P<0.05);干预后模型组腰4~6节段脊髓背角中cGMP蛋白阳性表达增多,NMDAR1、nNOS、sGCβ、PKG1蛋白表达增多,NMDAR1、nNOS、sGCβ、cGMP、PKG1 mRNA表达增多(均P<0.05)。与模型组比较,干预后推拿组和推拿+MK-801组CST降低,MWT升高(均P<0.05);推拿组、推拿+MK-801组腰4~6节段脊髓背角中cGMP蛋白阳性表达减少,NMDAR1、nNOS、sGCβ、PKG1蛋白表达减少,NMDAR1、nNOS、sGCβ、cGMP、PKG1 mRNA表达减少(均P<0.05)。结论 推拿干预1次后即可有效改善周围神经病理性疼痛引起的温度觉�Objective To explore the analgesic initiation mechanism of "three-manipulations and three-acupoints" of tuina on minor chronic constriction injury(minor CCI) model rats.Methods According to the random number table method,35 SD rats were randomly divided into five groups:normal group,sham group,model group,tuina group,and tuina + MK-801 group.The model group,tuina group,and tuina + MK-801 group were subjected to ligation of the right sciatic nerve trunk to establish a minor CCI rat model.The sham group was only exposed to the right sciatic nerve without ligation,and the normal group was not subjected to any operation.The normal group was not subjected to any intervention measures.On the seventh day after modeling,the model group and the sham group underwent 9 minutes of grasping restraint,while the tuina group underwent one intervention of three-manipulations(point method,dialing method,and kneading method) and three-acupoints(right "Yinmen"(BL37),"Chengshan"(BL57),and "Yanglingquan"(GB34)acupoints) with each manipulation and acupoint intervention for 1 minute for a total of 9 minutes.The tuina+MK-801 group received intrathecal injection of MK-801 from the fifth to seventh days after modeling,with a dose of 6 μg(10 μL) per day,tuina intervention was performed 30 minutes after the last intrathecal injection,and the specific operation of tuina was the same as that of the tuina group.Before modeling,after modeling,and after intervention,each group of rats was subjected to cold sensitivity threshold(CST) and mechanical withdrawal threshold(MWT) testing.After intervention,immunohistochemistry was used to detect the positive expression of cyclic guanosine monophosphate(cGMP) in the spinal dorsal horn(SDH) at L4-6 segments;protein expressions of N-methyl-D-aspartate receptor 1(NMDAR1),neurogenic nitric oxide synthase(nNOS),soluble guanylyl cyclase β(sGCβ),and protein kinase G1(PKG1) in SDH at L4-6 segments were detected by Western blotting;mRNA expressions of NMDAR1,nNOS,sGCβ,cGMP,and PKG1 in SDH at L4-6 segments we

关 键 词：推拿 神经病理性疼痛 N-甲基-D-天冬氨酸受体1 脊髓背角 镇痛 大鼠 

分 类 号：R244.1[医药卫生—针灸推拿学]