The Experimental Study On The Influence Of Different Frequency Of Electroacupuncture On Glu-NMDAR Signaling Pathway Of Hippocampus In Vascular Dementia Rats

Objective: To observe the intervention effect of electropuncture（2/100 Hz,2Hz,and 100 Hz variable frequency）on behavior,brain tissue pat hological morphology of hippocampal CA1 area,hippocampus Glu,Ca²⁺content,expression of NMDAR protein of vascular dementia（Vascular Dementia,VD）rats,discuss therole of VD Mechanism,and provide theVD electroacupuncture parameters to clinical treatment of acupuncture.Methods: Screened normal rats with learning and memory function by Morris water maze,they were randomly divided into the sham operation group and the model group.The bilateral common carotid artery repeatedly by intraperitoneal injection of ischeia-reperfusion plus sodium nitrate tome,method of preparation of VD model separation control rats common carotid artery,not injection,sodium nitrate,don,t block the blood flow.Blocking carotid artery after 1min to observe rat is reflection and iris changes to ensure that the brain ischemia.Postoperative survival model rats were randomly divided into the model group,the 2/100 Hz electric acupuncture group,the 2Hz electric acupuncture group,and the 100 Hz electric acupuncture group.They were treated on the 3d.The housanli（double）,geshu（double）,dazhui,baihui,were choosed on the electropuncture group,acupuncted with filifo,connected with the electric acupuncture apparatus instrument,respectively with frequency conversion of 2/100 times/second,2 times/second of low frequency and 100 times/s of high frequency,10 minutes every time and one time a day,continuous treatment of 15 days.Control group and model group are in the same position fixed 10 minutes,one time a day,continuous 15 days.On the 17 day,we examined jumping stand experiment.Then,The mice were beheaded,the right hippocampus was make into 10% homogenate,conventional paraffin embedding,sectioning undertest.The homogenate was centrifugated for 5 min at 4℃,12000r/min,the supernatant was extracted.Then,we testing the expression of NMDAR by Western Blot.The rats were beheaded and the left brains were taken out on the ice plate,and the left side of the braintissue was taken and fixated in 4% formal dehyde.The tissue was stained by HE（Hematoxylin and Eosin）.We observed morphologic changes of hippocampus CA1 region under light microscope.Results:1 The effect of electropuncture with different frequency on ability of learning and memory of VD ratsThe results of jumping stand learning ability experiment show: compared with the sham operation group,the reaction time（s）of model group rats is significantly prolong and error times increased significantly,with significant statistical differences（P<0.01）.The three kinds of frequency electric acupuncture can shorten the reaction time（s）and to decrease the error times（P<0.01 or P<0.05）.There were no significant difference among 2/100 Hz group,2Hz group,100 Hz group（P>0.05）.The results of jumping stand memory ability experiment show: compared with the sham operation group,the latency（s）of model group rats is significantly shorten and error times increased significantly,with significant statistical differences（P<0.01）.The three kinds of frequency electric acupuncture can prolong the reaction time（s）and to decrease the error times（P<0.01 or P<0.05）.There were no significant difference among 2/100 Hz group,2Hz group,100 Hz group（P>0.05）.2 The effect of electropuncture with different frequency pathomorph ology on Hippocampus CA1 region of VD ratsCompared with sham-operated rats,the number of pyramidal cells in the model group hippocampus significantly reduced,appear edema,cell arrangement scattered,loosely arranged,cell shape irregularly,some nuclei deeply stained,condensation,nucleolar fragmentation,even nucleolus disappears.Pyramidal cells in the hippocampus compared with model group,three EA group rats significantly improved,the loss of pyramidal cells reduced,edema reduced,cells arranged in neat,tight,nearly circular or oval cells,nucleolus clearly.3 The effect of electropuncture with different frequency on the content of Glu in hippocampus CA1 regionof VD ratsCompared to the sham operation group,Glu content in hippocampus of model group were significantly increased,there are significant differences（P<0.01）.Three frequencies electroacupuncture can reduce Glu content in hippocampal tissue,compared with the model group were significantly different（P<0.05 or P<0.01）,but compared between any two of each EA group were not statistically different（P>0.05）.4 The effect of electropuncture with different frequency on the content of Ca²⁺ in hippocampus CA1 region of VD ratsCompared to the sham operation group,Ca²⁺ content in hippocampus of model group were significantly increased,there are significant differences（P<0.01）.Three frequencies electroacupuncture can reduce Ca²⁺ content in hippocampal tissue,compared with the model group were significantly different（P<0.05 or P<0.01）,but compared between any two of each EA group were not statistically different（P> 0.05）.5 The effect of electropuncture with different frequency on the expr essions of NMDAR in hippocampus CA1 regionof VD ratsCompared to the sham group,NMDAR protein in hippocampus of rats in model group was significantly increased,a significant difference（P<0.01）.Three frequencies electroacupuncture can reduce NMDAR protein expression,compared with the model group were statistically significant differences（P<0.01）,but there was no difference between EA group（P>0.05）.Conclusion:In this study,2Hz,100Hz,2/100 Hz three frequencies electroacupuncture intervention can improve ischemic injury in hippocampus of VD model rats,improve learning and memory ability of the model rats.Its mechanism may be through reducing content of Glu in hippocampus,inhibition of NMADR receptor expression,adjusting the Glu-NMADR signal transduction system,reducing the content of Ca²⁺ in hippocampus,thereby reducing the hippocampus neural cells of VD model rats of excited toxicity,fighting the nerve cell damage,improving the function of the central nervous system.But three treatment effects of different frequencies of electro-acupuncture showed no significant differences.