电针对神经病理性痛大鼠脊髓背角细胞内钙离子浓度以及钙-钙调素依赖性蛋白激酶Ⅱ的影响  

Effect of Electroacupuncture on Intracellular Calcium Ion Concentration and Expression of Calcium/Calmodulin Dependent Protein Kinase Ⅱ in Lumbar Spinal Cord in Rats with Neuropathic Pain

作　　者：吴辛甜[1] 梁伟 闫丽萍[3] 王玲玲[3] 马骋[3] 

机构地区：[1]南京中医药大学附属常州市中医医院颈腰痛中心,江苏常州213003 [2]江苏凯基生物技术股份有限公司,南京210012 [3]南京中医药大学江苏省针灸学重点实验室,南京210029

出　　处：《针刺研究》2017年第6期496-501,共6页Acupuncture Research

基　　金：国家自然科学基金面上项目(No.81173329)

摘　　要：目的:通过观察坐骨神经分支选择性损伤(SNI)大鼠脊髓背角神经细胞内钙离子([Ca2+]i)浓度以及钙-钙调素依赖性蛋白激酶Ⅱ(CaMKⅡ)的变化和电针干预对其的影响,探讨电针干预神经病理性痛的脊髓机制。方法:SD大鼠随机分为假手术组、模型组、电针组、N-甲基-D-天冬氨酸受体拮抗剂(AP-5)组和一氧化氮合酶抑制剂(L-NAME)组,每组22只。采用SNI法制备神经病理性痛模型,假手术组仅分离不结扎。电针组电针大鼠损伤侧"委中""环跳"穴30min,1次/d,AP-5组予AP-5 0.7mg·kg-1·d-1腹腔注射,L-NAME组予L-NAME 60mg·kg-1·d-1腹腔注射,连续7d。造模前及SNI后10d、16d分别测定机械痛阈。激光共聚焦显微镜技术观察脊髓背角[Ca2+]i的荧光强度;Western blot和免疫组化法测定脊髓CaMKⅡ的表达。结果:与假手术组比较,SNI后10d各组机械痛阈值均降低(P<0.01);与模型组比较,电针组、AP-5组和L-NAME组SNI后16d时机械痛阈值均升高(P<0.01,P<0.05)。与假手术组比较,模型组脊髓背角[Ca2+]i浓度与CaMKⅡ表达均升高(P<0.01,P<0.05);与模型组比较,电针组、AP-5组和L-NAME组脊髓背角[Ca2+]i浓度均被逆转(P<0.05,P<0.01),而CaMKⅡ的表达仅电针组被逆转(P<0.05)。结论:电针减轻大鼠神经病理性痛的机制之一,可能与其有效下调脊髓背角[Ca2+]i浓度及CaMKⅡ的表达,继而抑制其一系列后效应有关。Objective To observe the changes of intracellular calcium （EOa2＋ Ⅱ） concentration and expression of calcium/ calmodulin dependent protein kinase Ⅱ （CaMK Ⅱ） in spinal dorsal horn neurons of spared nerve injury （SNI） rats, so as to explore its mechanisms underlying improvement of neuropathic pain. Methods One hundred and ten SD rats were randomly divided into 5 groups： sham control, model, EA, AP-5 and L-NAME groups. The sham group underwent only a simple separation of the sciatic nerve but without ligation and abscission. The neuropathic pain model was established by abscission of the right tibial and common peroneal nerve. EA （2 Hz, 1 -3 mA） was applied to right ＂Weizhong＂ （BL 40） and ＂Huantiao＂ （GB 30） for 30 min, once a day for 7 days, starting from day 11 after surgery. For rats of the AP-5 and L-NAME groups, AP-5 （a competitive antagonist for NMDA receptor, 0.7 mg· kg-1 · d-1） and L-NAME （a non-selective antagonist for nitric oxide synthase [NOS], 60 mg · kg-1· d-1） were respectively administrated by intraperitoneal injection, once daily for 7 days. The mechanical pain threshold was measured, and the calcium fluorescence intensity （shown by Fluo-3/AM calcium fluorescence indicator） of the superficial layer of the lumbar spinal cord （L 4- L 6） was measured by immunohistochemical staining and the expression of spinal cord （L 4 - L 6） CaMK Ⅱ protein was detected by Western blot （WB）. Results After modeling, the mechanical pain threshold was significantly decreased on day 10 and 16 after operation in comparison with the sham operation group and baseline data of pre-operation in each group （P〈0.01）, and remarkably increased in the EA, AP-5 and L-NAME groups relevant to the model group on day 16 （P〈0.01, P〈0.05）, while the effect of EA was significantly superior to that of AP-5 and L-NAME groups （P〈0.05）, suggesting a reduction of EA analgesia after administration of AP-5 and L-NAME. The concentration of intracellular [Ca2�

关 键 词：电针 神经病理性痛 脊髓 细胞内钙离子 钙-钙调素蛋白激酶Ⅱ 激光共聚焦显微镜 

分 类 号：R245.97[医药卫生—针灸推拿学]