| Objective:The aim of this study was to investigate the effect of electroacupuncture on pattern separation mediated by newborn immature granule cells in the hippocampal dentate gyrus in a mouse model of Alzheimer’s disease(AD)and its possible mechanism.Methods:According to the results of gene identification,3-month-old 5×FAD-positive mice were randomly divided into model group,electroacupuncture group,model+Wnt5a interference virus group,electroacupuncture+Wnt5a interference virus group,electroacupuncture+empty virus group.Negative mice in the same litter were used as the wild group,with 20 mice in each group.Electroacupuncture at Baihui and Shenting points of mice,sparse-dense wave,frequency1/20 Hz,once a day,30 minutes each time,5 days a week,intervention for 4 weeks;The mice in the wild group,model group and model+Wnt5a interference virus group were grasped and fixed under the same conditions without any treatment.Since electroacupuncture,each mouse received Brd U intraperitoneal injection twice every other day at a dose of 50 mg/kg.Before and after the intervention,the open field test was used to evaluate the exploration ability,and the object location discrimination test to evaluate the spatial discrimination memory.The touch screen behavioral test to evaluate the pattern separation ability of the mice after the intervention.Immunohistochemistry was used to detect the deposition level of Aβplaques in the bilateral hippocampal DG region after the intervention;Immunofluorescence was used to detect the number of Ca R~+/Brd U~+cells in the DG region of the bilateral hippocampus after the intervention.Western blotting was used to detect the expression levels of Ca R and Prox1 genes in the hippocampus after the intervention.Western blotting was used to detect the expression levels of Wnt5a,DVL2,Ca MKII and p-Ca MKII in the Wnt pathway in the hippocampus after the intervention.Results:Effects of electroacupuncture at Baihui and Shenting points on cognitive behavior:In the object location discrimination test,before the intervention,compared with the wild group,the position discrimination index of the remaining five groups of mice decreased(P<0.01),and there was no statistical difference in the total distance(P>0.05).After the intervention,compared with the wild group,the discrimination index of the model group decreased(P<0.001);Compared with the model group,the recognition index of the electroacupuncture group increased(P<0.05),and the discrimination index of the model+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the discrimination index of the electroacupuncture+Wnt5a interference virus group was decreased(P<0.05),but there was no difference in the electroacupuncture+empty virus group(P>0.05),and there was no statistical difference in the total distance(P>0.05).It was found that there was no statistical difference in the total distance,the distance in the central area,and the time in the central area of the mice in each group before and after the intervention in the open field test(P>0.05).Effects of electroacupuncture at Baihui and Shenting points on the Trial-Unique Nonmatching-to-Location of the touch screen test:The results of the showed that:after the intervention,under the difficulty of large separation interval S2,compared with the wild group,the correct rate of the model group decreased(P<0.001);Compared with the model group,the correct rate of the electroacupuncture group increased(P<0.01),the correct rate of model+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the correct rate of the electroacupuncture+Wnt5a interference virus group decreased(P<0.05),but there was no difference in the electroacupuncture+empty virus group(P>0.05).Under the difficulty of small separation interval S1:the correct rate of the wild group was higher than that of the model group(P<0.05),and there was no difference between the other groups(P>0.05).Effects of electroacupuncture at Baihui and Shenting points on the pathological manifestations of Aβin the DG:After intervention,compared with the wild group,the deposition of Aβin the model group was significantly increased(P<0.001);Compared with the model group,the number of plaques in the electroacupuncture group decreased(P<0.001),the number of plaques in the model+Wnt5a interference virus group did not increase significantly(P>0.05);Compared with the electroacupuncture group,the number of plaques in the electroacupuncture+Wnt5a interference virus group had no significant change(P>0.05),and there was no difference in the number of plaques in the electroacupuncture+empty virus group(P>0.05).Effects of electroacupuncture at Baihui and Shenting points on the regeneration of immature granule cells in the DG:After intervention,immunofluorescence detection showed that compared with the wild group,the number of Ca R~+/Brd U~+cells in the model group decreased(P<0.001);The number of Ca R~+/Brd U~+in the acupuncture group increased(P<0.05),and the number of Ca R~+/Brd U~+in the model+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the number of Ca R~+/Brd U~+in the electroacupuncture+Wnt5a interference virus group decreased(P<0.05),but there was no difference in the electroacupuncture+empty virus group(P>0.05);Western blot showed that compared with the wild group,the expression of Ca R and Prox1 in the model group decreased(P<0.001);Compared with the model group,the expression of Prox1 in the electroacupuncture group increased(P<0.05),the expression of Ca R in the electroacupuncture group increased(P<0.01);The expression of Ca R and Prox1 in mode+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the expression of Ca R and Prox1 in the electroacupuncture+Wnt5a interference virus group decreased(P<0.05),but there was no difference in the electroacupuncture+empty virus group(P>0.05);The effect of electroacupuncture at Baihui and Shenting points on the expression of Wnt pathway-related proteins:After intervention,Western blott showed that:compared with the wild group,the expression of Wnt5a in the model group decreased(P<0.001);Compared with the model group,the expression of Wnt5a in the electroacupuncture group increased(P<0.05),and the expression level of Wnt5a in the model+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the expression level of Wnt5a of the electroacupuncture+Wnt5a interference virus decreased(P<0.05),and there was no difference in the electroacupuncture+empty virus group(P>0.05);In terms of the expression level of receptor DVL2,compared with the wild group,the expression of DVL2 in the model group decreased(P<0.01);Compared with the model group,the expression level of DVL2 in the electroacupuncture group increased(P<0.01),and the expression level of the model+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the expression level of DVL2 of the electroacupuncture+Wnt5a interference virus decreased(P<0.05),but there was no difference in the electroacupuncture+empty virus group(P>0.05);There was no statistical difference in the expression of Ca MKII among the groups(P>0.05);Compared with the wild group,the expression of p-Ca MKII in the model group decreased(P<0.001).The expression level of p-Ca MKII increased(P<0.01),and the expression level of model+Wnt5a interference virus group decreased(P<0.05);Compared with the electroacupuncture group,the expression level of p-Ca MKII in the electroacupuncture+Wnt5a interference virus group decreased(P<0.05),but there was no difference in the electroacupuncture+empty virus group(P>0.05);Conclusion:Electroacupuncture at Baihui and Shenting points can promote the pattern separation mediated by the neurogenesis of immature granule cells in the hippocampal DG area by regulating the Wnt signaling pathway,thereby improving the memory discrimination ability of AD model mice. |
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