| Objective: In this experiment,the behavioral indexes of Tongdu Anshen fang for depression model rats were observed,and the CyclicAdenosine Monophosphate（cAMP）,Protein kinase A（PKA）,cAMP response Element binding protein（CREB）protein expression level,to explore the anti-depression mechanism of Tongdu Anshen Fang.Methods:Forty SPF male SD rats were randomly divided into 5groups: blank control group,model control group,fluoxetine group,Xiaoyao San group and Tongdu Anshen Fang group,with 8 rats in each group.Chronic mild and unpredictable stimulation combined with solitary cultivation were used to make model rats with depression for 21 consecutive days.Body mass index,sugar water preference test and open field test were observed before and after modeling and treatment to test the success of modeling.After successful modeling,drug intervention was carried out,and 7-day gavage treatment was carried out.Normal control group and depression model control group were given equal volume normal saline,and fluoxetine group,Xiaoyao San group and Tongdu Anshen Fang group were given corresponding drugs.After treatment,the protein expression levels of cAMP,PKA and CREB in hippocampus were detected by Western blot（WB）,and the content of CREB in hippocampus was detected by immunohistochemistry.Statistical analysis was performed on the experimental data,which was expressed as mean±standard deviation（X±S）.One-way ANOVA was used for comparison of multiple means,and P<0.05 was statistically significant.Results:1.Behavioral test results:According to the weight change results,after modeling,the weight growth rate of model control group,Xiaoyao San group,Tongdu Anshen Fang group and fluoxetine group was slower than that of blank control group,and the differences were statistically significant compared with blank control group（P<0.01）.After gavage treatment,compared with the model control group,the body weight of the fluoxetine group increased significantly（P< 0.01）,but there was no statistical significance between Xiaoyao San group and Tongdu Anshen Fang group.The results of sugar water preference experiment showed that there was no significant difference in sugar water preference rate of rats before modeling.After modeling,compared with blank control group,sugar water preference rate of model control group,Xiaoyao San group,Tongdu Anshen Fang and fluoxetine group was significantly decreased（P < 0.01）;After administration,compared with model control group,the preference rate of sugar water in Tongdu Anshen Fang group was significantly increased（P<0.05）.Compared with blank control group,the total open field distance of rats was significantly decreased and the rest time of rats was significantly increased after modeling（P < 0.01 or P < 0.05）.After drug administration,compared with model control group,the total open field distance of rats in all treatment groups was significantly increased（P < 0.01）,and the difference was statistically significant.2.Experimental test index results:（1）Results of WB method: After treatment,compared with blank control group,the expression levels of cAMP,PKA and CREB protein in model control group were significantly decreased（P<0.01）;compared with model control group,the expression levels of cAMP protein in Xiaoyao San group,Tongdu Anshen Fang group and fluoxetine group were significantly increased（P < 0.05 or P <0.01）;The protein expression levels of PKA and CREB in Tongdu Anshen Fang group and fluoxetine group were significantly increased（P<0.05）;Compared with Xiaoyao San group,cAMP and CREB protein expression levels in Tongdu Anshen Fang group were significantly increased（P<0.01）;Compared with fluoxetine group,cAMP protein expression level in Tongdu Anshen Fang group was significantly increased（P<0.01）.（2）Immunohistochemical test results:Compared with blank control group,the positive expression of CREB in hippocampus CA1 and CA3 of model control group was significantly decreased（P<0.01）;Compared with model control group,the positive expression of CREB in hippocampus CA1 region in fluoxetine group was significantly increased（P<0.01）,the positive expression of CREB in hippocampus CA1 and CA3 region in Tongdu Anshen Fang group was significantly increased（P<0.05）,while there was no statistical significance in Xiaoyao San group.Conclusion:Tongdu Anshen Fang can improve the behavioral performance and symptoms of depression in model rats.It can increase the contents of cAMP,PKA and CREB proteins in the hippocampus of depressed model rats,and improve the depressive state of rats by enhancing the effect of cAMP-PKA-CREB signaling pathway,suggesting that cAMP signaling pathway is one of the targets of Tongdu Anshen Fang. |
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