Study On Anti-Depression Effect Of Electroacupuncture On Hippocampal Synaptic Plasticity By Neurogenic Nitric Oxide Synthase Down-regulation

Objective:To explore the effects of electroacupuncture(EA)on synaptic plasticity and nNOS,then analyse the internal relation between these three factors.Methods:Experiment 1:Classifying 80 SD rats into 4 groups randomly based on their weights,namely normal control group,model control group,sham EA group and EA group.Except normal control group,using CUMS modeling to deal with the other groups.After making the model,the rats in the EA group were treated with EA at "Yintang"(GV29)and"Baihui"(GV20)with 2Hz continuous wave,once a day,15 minutes every time for consecutive 14 days.while the rats in sham EA group recover 14 days without any stimulus.With open-field test,western blot and transmission electron microscope,the differences between each groups in behavior,expression quantity of nNOS and neuronal synapse in hippocampal CA1 area were observed.Experiment 2:According to body weights,48 SD rats were randomly divided into a normal group,a model group,a sham-operation group and intracerebroventricular injection(surgery)groups at different concentrations.The model group was treated by chronic and unpredictable mild stress.The surgery groups received intracerebroventricular injection with 7-NI solutions at different concentrations,once/3 days and 3 times in total.The sham-operation group was injected with DMSO of the same volume.The rat behaviors were then subjected to the open field test(OFT).The hippocampal nNOS protein levels were detected by Western blotting.Experiment 3:Classifying male SD rats into 3 groups randomly based on their weights,namely normal control group,7-NI group and EA group.Then the rats in the 7-NI group were treated with intracerebroventricular 7-NI solution of 200nmol/0.5 μ l concentration,once in 3 days,3 times in total.The rats in the EA group were treated with EA at "Yintang"(EX-HN 3)and"Baihui"(GV 20)with 2 Hz continuous wave,once a day,15 minutes every time for consecutive 14 days.With western blot,REAL TIME-PCR and transmission electron microscope,the differences between each groups in expression quantity of nNOS,expression quantity of nNOS mRNA and neuronal synapse in hippocampal were observed.Results:Experiment 1:Compared with normal control group,after making the model,rats in the other groups show up obvious depression-like behaviors in open-field test.After 14 days of EA therapy,compared with model control group,each indicators of open-field test were significantly higher in the EA group(P<0.01).After 14 days of self-recovery process,compared with model control group,the moving distance were a bit higher in the sham EA group(P>0.05).It proves that EA therapy has the obvious recovery effect on the animals.It is unrelated to self-recovery process.Synaptic number statistics shows that,compared with normal control group,in hippocampal CA1 area of model control group and sham EA group,synaptic number decreased significantly(P<0.01).Compared with model control group,in hippocampal CA1 area of EA group,synaptic number increased significantly(P<0.01),without obvious difference compared with normal control group(P>0.05).The Result of the observation of transmission electron microscope shows that compared with normal control group,the neurons cells in hippocampal CA1 area in model control group and sham EA group showed damages such as pyknosis and mitochondria swelling.While recovery of the neurons cells in hippocampal CA1 area in EA group was significant.Western blot results show that compared with normal control group,the expression quantity of nNOS in rats’ hippocampal CA1 area in the model control group increased obviously(P<0.01).Compared with model control group,the expression quantity of nNOS in rats’ hippocampal CA1 area in the EA group decreased.The result has significant statistical significance(P<0.01).There is no difference between model control group and sham EA group(P>0.05).Experiment 2:Compared with the normal group,the total movement distances of model and 200nmol/0.5μl surgery group decreased significantly(P<0.05).Compared with the model group,such distances of sham-operation and surgery groups at other concentrations increased significantly(P<0.05).The normal group entered the central area significantly more times than model and 200nmol/0.5 μ l surgery group did(P<0.05),and the number of times entering the central area of the model group was significantly lower than that of the sham-operation group(P<0.05).Compared with the normal group,the durations of stay in the central area of model and 200nmol/0.5μl surgery group were shortened significantly(P<0.05).Western blotting showed that the hippocampal nNOS protein levels of model and 200nmol/0.5μl surgery group significantly exceeded that of the normal group(P<0.05).The levels of sham-operation and 50nmol/0.5μl surgery group,which were significantly lower than that of the model group(P<0.05),were similar to that of the normal group(P>0.05).Experiment 3:The Result of the observation of transmission electron microscope shows that compared with normal control group,the neurons cells in hippocampal CA1 area in 7-NI group showed damages such as pyknosis and mitochondria swelling.While recovery of the neurons cells in hippocampal CA1 area in EA group was significant.Synaptic number statistics shows that,compared with normal control group,in hippocampal CA1 area of 7-NI group,synaptic number decreased significantly(P<0.05).In hippocampal CA1 area of EA group,synaptic number increased significantly(P<0.05),without obvious difference compared with normal control group(P>0.05).Western blot results show that compared with normal control group,the expression quantity of nNOS in rats,hippocampal in the 7-NI group increased obviously(P<0.05).Compared with 7-NI group,the expression quantity of nNOS in rats’hippocampal in the EA group decreased.The result has significant statistical significance(P<0.01).REAL TIME-PCR results show that compared with normal group,the nNOS mRNA expression levels in 7-NI group were increased(P<0.05),whereas compared with7-NI group,the nNOS mRNA expression levels in EA group were decreased(P<0.05).Conclusion:EA can caused antidepressant-like effects.To cure depression through EA,we can use nNOS as mediation in order to influence the synaptic plasticity.