Effects of electroacupuncture of different frequencies on free radicals in hippocampus of mice with vascular dementia

Objective:To observe the effects of electroacupuncture (EA) of three different frequencies (2 Hz,80 Hz and 2 Hz/80 Hz) on the free radicals in hippocampus of vascular dementia (VD) model mice.Methods:A total of 100 Kunming mice were randomly divided into a sham operation group,a model group,a 2 Hz EA group,an 80 Hz EA group and a 2 Hz/80 Hz EA group,with 20 mice in each group.The ischemia-reperfusion VD model was established by repeated blockade of bilateral common carotid arteries.Mice in EA groups began EA treatment on the 4th day after the operation.Baihui (GV 20),Dazhui (GV 14),Geshu (BL 17) and Zusanli (ST 36) were punctured and then connected to EA instrument,with different waves of 2 Hz,80 Hz or 2 Hz/80 Hz (10 min/time) applied accordingly,once a day.During the jumping stand experiment,the learning performance,memory performance and hippocampal calcitonin gene-related peptide (CGRP),nitric oxide synthase (NOS),malondialdehyde (MDA),changes in superoxide dismutase (SOD) and true choline esterase (TChE) were observed.In hippocampus,the CGRP level was determined by radioimmunoassay;the MDA level was determined by thiobarbituric acid colorimetric method;the activities of NOS and TChE were determined by spectrophotometry;the activity of SOD was determined by xanthine oxidase method.Results:Compared with the sham operation group,the performances of learning and memory decreased significantly in the model group (P＜0.01);in hippocampus,the CGRP level decreased,the MDA level increased,the activities of NOS and TChE increased,and the activity of SOD decreased in the model group.Compared with the model group,the learning and memory performances of the EA groups were significantly improved (P＜0.05 or P＜0.01);in hippocampus,the CGRP level increased,the MDA level decreased,the NOS and TChE activities decreased,and the SOD activity increased (P＜0.05 or P＜0.01).Among EA groups,the 2 Hz/80 Hz EA group was superior to the 2 Hz EA group and the 80 Hz EA group (P＜0.05 or P＜0.01).Conclusion:EA can improve the cognitive impairment of mice with ischemia-reperfusion VD.The mechanism may be related to the improvement of cerebral blood circulation,regulation of the central neurotransmitters,fighting lipid peroxidation and promoting nerve cell repair.The therapeutic effects of EA with different frequencies were different,and the intervention effect by EA at 2 Hz/80Hz is the most significant.

电针对血管性痴呆大鼠海马区c-Jun氨基末端激酶信号通路的影响

目的:观察电针干预对血管性痴呆(VD)大鼠海马组织c-Jun氨基末端激酶(JNK)信号通路相关蛋白表达的影响,探讨电针治疗VD的作用机制。方法:雄性SD大鼠随机分为假手术组、模型组、电针组,每组10只。采用反复阻断双侧颈总动脉法制备VD大鼠模型。电针组电针"大椎""百会"及双侧"后三里""膈俞"穴,10min/次,1次/d,连续治疗14d。Morris水迷宫检测大鼠行为学变化,尼氏染色法观察海马神经元病理改变,原位末端转移酶标记(TUNEL)法检测海马神经元凋亡,Western blot法检测JNK、磷酸化JNK(p-JNK)、半胱氨酸蛋白酶-8(Caspase-8)、半胱氨酸蛋白酶-3(Caspase-3)蛋白表达。结果:与假手术组比较,模型组大鼠Morris水迷宫实验逃避潜伏期明显延长,穿越原平台次数明显减少(P<0.01),海马神经元损伤加重,存活神经元明显减少(P<0.01),凋亡神经元增多,细胞凋亡指数明显升高(P<0.01),海马组织JNK、p-JNK、Caspase-8、Caspase-3蛋白表达均明显升高(P<0.01)。与模型组比较,电针组大鼠Morris水迷宫逃避潜伏期明显缩短,穿越原平台次数明显增多(P<0.01),海马神经元损伤减轻,存活神经元明显增多(P<0.01),凋亡神经元减少,细胞凋亡指数明显降低(P<0.01),海马组织JNK、p-JNK、Caspase-8、Caspase-3表达均显著降低(P<0.01)。结论:电针可提高VD大鼠学习记忆能力,其作用机制可能与电针调控JNK信号通路相关蛋白表达,抑制神经元凋亡有关。

Effects of electroacupuncture with different frequencies on hippocampal neuronal apoptosis and JNK signaling pathway in rats with vascular dementia

Objective:To observe the effects of electroacupuncture(EA)with three frequencies(100 Hz,2 Hz,and 2 Hz/100 Hz)on the apoptosis of neurons and c-Jun N-terminal kinase(JNK)signaling pathway in the hippocampus of rats with vascular dementia(VD),and explore the mechanism of EA intervention for VD.Methods:Fifty male Sprague-Dawley rats were randomly divided into a model group,a sham operation group,a 100 Hz EA group,a 2 Hz EA group,and a 2 Hz/100 Hz EA group,with ten rats in each group.The VD model rats were established by repeated ischemia-reperfusion of bilateral common carotid arteries.The rats in the EA groups received EA intervention at Baihui(GV20),Dazhui(GV14),Geshu(BL17)and Zusanli(ST36),once a day for 14 d.Afterward,Morris water maze was used to examine the learning and memory performances of the rats in each group,hematoxylin-eosin staining to observe the histomorphological changes in the hippocampal CA1 region,terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling to test the apoptosis of neurons in the hippocampal CA1 region,and Western blot to detect the protein expression levels of JNK,phosphorylated JNK(p-JNK),Caspase-8,and Caspase-3 in the hippocampus tissue.Results:Compared with the sham operation group,the escape latency of the model group in water maze test was prolonged;the number of crossing the original platform was decreased(P<0.01);the hippocampal neurons were severely damaged and the number of surviving neurons was decreased(P<0.01),whereas the number of apoptotic neurons was increased(P<0.01);the protein expression levels of JNK,p-JNK,Caspase-8,and Caspase-3 in the hippocampus were significantly increased(P<0.01).Compared with the model group,the escape latency of each EA group was significantly shortened;the number of crossing the original platform was significantly increased(P<0.01);the damage of hippocampal neurons was alleviated,the number of surviving neurons was increased(P<0.01),and the number of apoptotic neurons was decreased(P<0.01);the protein expression levels of JNK,p-JNK,Caspase-8,and Caspase-3 in the hippocampus were decreased(P<0.01).The results in the 2 Hz EA group and the 2 Hz/100 Hz EA group were superior to those in the 100 Hz EA group.Conclusion:EA with the three frequencies(100 Hz,2 Hz,and 2 Hz/100 Hz)can improve the learning and memory performances in VD rats subjected to ischemia-reperfusion,its mechanism may be related to the inhibition of neuronal apoptosis and the regulation of the related protein expression of JNK signaling pathway,and the intervention effects of EA with 2 Hz and 2 Hz/100 Hz are more significant.