电针“水沟”对大鼠急性脑梗死后远隔部位继发性损伤的影响

Effect of Electro-acupuncture “Shuigou(DU26)” on Secondary Injury to the Distal Compartment after Acute Cerebral Infarction in Rats

【目的】探讨电针水沟穴对急性脑梗死大鼠的治疗作用及机制。【方法】将60只SD大鼠随机分为假手术组、大脑中动脉闭塞(MCAO)组、MCAO+皮层扩散性抑制(CSD)组和电针组,每组再分为造模后6 h、24 h、72 h 3个亚组,每亚组5只。采用改良Zea Longa再灌注法制备大鼠MCAO模型,再采用机械针刺的方式诱导产生单波CSD构建MCAO伴随CSD模型。造模成功后,电针组电针水沟穴,每次30 min。干预结束后,采用Zea Longa评分对大鼠神经行为学进行评估,末端脱氧核苷酸转移酶介导的dUTP缺口末端标记(TUNEL)染色法观察海马远隔CA1区神经细胞凋亡情况,免疫荧光标记法观测海马远隔CA1区单羧酸转运体1(MCT1)/胶质纤维酸性蛋白(GFAP)共表达情况,定磷法检测海马远隔CA1区Na^(+)-K^(+)-ATP酶活性。【结果】(1)与MCAO+CSD组比较,电针组6、24、72 h时相神经功能缺损评分明显降低(P<0.05)。(2)与假手术组比较,其余3组各时相海马远隔CA1区神经细胞凋亡指数均显著升高(P<0.05);与MCAO组比较,MCAO+CSD组各时相神经细胞凋亡指数明显升高(P<0.05);与MCAO+CSD组比较,电针组各时相神经细胞凋亡指数均明显降低(P<0.05)。(3)与假手术组比较,MCAO组、MCAO+CSD组各时相Na^(+)-K^(+)-ATP酶活性均显著降低(P<0.05);与MCAO组比较,MCAO+CSD组6 h、24 h时相Na^(+)-K^(+)-ATP酶活性显著降低(P<0.05);与MCAO+CSD组比较,电针组6 h、24 h时相Na^(+)-K^(+)-ATP酶活性均显著提高(P<0.05)。(4)与假手术组比较,MCAO+CSD组各时相MCT1/GFAP共表达量显著升高(P<0.05),MCAO组、电针组在24 h时相显著升高(P<0.05);与MCAO组比较,MCAO+CSD组各个时相MCT1/GFAP共表达量显著升高(P<0.05),电针组在24 h时相MCT1/GFAP共表达量显著降低(P<0.05);与MCAO+CSD组比较,电针组各个时相MCT1/GFAP共表达量显著降低(P<0.05)。【结论】电针水沟穴可有效改善大鼠急性脑梗死后远隔部位继发性损伤,其机制可能与提高海马远隔区Na^(+)-K^(+)-ATP酶活性,抑制MCT1、GFAP星形胶质细胞内正向表达的作用有关。

Objective To investigate the therapeutic effect and mechanism of electro-acupuncture at Shuigou(DU26)acupoint in rats with acute cerebral infarction.Methods Sixty SD rats were randomly divided into the sham-operation group,the middle cerebral artery occlusion(MCAO)group,MCAO+cortical spreading depression(CSD)group and electro-acupuncture group,and each group was further divided into 3 subgroups of 5 rats:6 h,24 h and 72 h.A modified Zea Longa reperfusion method was used to prepare the rat MCAO model,and then mechanical acupuncture was used to induce a single wave of CSD to construct the MCAO with CSD model.After successful modeling,the electro-acupuncture group applied electro-acupuncture at the Shuigou acupoint for 30 minutes each time.At the end of the intervention,the neurobehaviour of the rats was assessed using the Zea Longa scoring,terminal deoxynucleotide transferase-mediated dUTP Nick end labeling(TUNEL)staining method was used to observe the apoptosis of neurons in the distant CA1 region of hippocampus,the co-expression of monocarboxylic acid transporter 1(MCT1)/glial fibrillary acidic protein(GFAP)in the distant CA1 region of hippocampus was observed by immunofluorescence labeling method,and the activity of Na^(+)-K^(+)-ATPase in distant CA1 region of hippocampus was detected by fixed phosphorus method.Results(1)Compared with the MCAO+CSD group,the neurofunction deficit score was significantly decreased in the electro-acupuncture group at 6-,24-,72-hour time phases(P<0.05).(2)Compared with the sham-operation group,the apoptotic index of neuronal cells in the distal CA1 region of the hippocampus was significantly increased in the remaining three groups at all time phases(P<0.05);compared with the MCAO group,the apoptosis index of neural cells in each phase of MCAO+CSD group was significantly increased(P<0.05),compared with the MCAO+CSD group,the apoptosis index of neural cells in each phase of the electro-acupuncture group was significantly decreased(P<0.05).(3)Compared with the sham-operation group,the activity of Na^(+)-K^(+)-ATPase was significantly decreased in the MCAO group and MCAO+CSD group at all time phases(P<0.05);compared with the MCAO group,the activity of Na^(+)-K^(+)-ATPase was significantly decreased in the MCAO+CSD group at the 6-,24-hour time phases(P<0.05);compared with the MCAO+CSD group,the activity of Na^(+)-K^(+)-ATPase was significantly increased in the electro-acupuncture group at the 6-,24-hour time phases(P<0.05).(4)Compared with the sham-operation group,MCT1/GFAP co-expression was significantly increased in the MCAO+CSD group at all time phases(P<0.05),and significantly increased in the MCAO group and electro-acupuncture group at the 24-hour time phase(P<0.05);compared with the MCAO group,the co-expression of MCT1/GFAP at all time phase in MCAO+CSD group was significantly increased(P<0.05),and the co-expression of MCT1/GFAP at 24-hour time phase in electro-acupuncture group was significantly decreased(P<0.05);compared with the MCAO+CSD group,the coexpression of MCT1/GFAP in electro-acupuncture group was significantly decreased at all time phase(P<0.05).Conclusion Electro-acupuncture at Shuigou acupoint has effectively improved the secondary injury in the distal compartment after acute cerebral infarction in rats,and the mechanism may be related to the effect of increasing Na^(+)-K^(+)-ATPase activity in the hippocampal distal compartment and inhibiting positive expression in GFAP and MCT1 astrocytes.