Effects Of Shenguiren Mixture On The Morphology,Number Of Frontal Lobe Neurons And The Expression Of GABA Signal Molecules In Rats With Postpartum Depression

Objective:This experiment is based on the recognized rat model of postpartum depression,with the traditional Chinese medicine compound Shenguiren mixture for intervention.Observe the behavioral changes of experimental rats,the damage of rat frontal lobe tissues,the apoptosis of rat frontal lobe nerve cells and the expression ofγ-aminobutyric acid（GABA）related molecules in the frontal lobe of rats.Explore the mechanism of Shenguiren mixture in the treatment of postpartum depression,and provide a basis for its clinical application.Methods:62 SD rats were selected for the animal experiment,and then divided into 5groups（blank group,sham operation group,model group,E₂group and Shenguiren group）according to their body weight,and tested each group of rats in the open field experiment（open filed test,OFT）whether the scores are on the same baseline.The rats in the blank group were not operated on.Rats in the sham operation group had dorsal incisions on both sides,and the ovaries were not removed after the ovaries were explored,and the incisions were directly sterilized and sutured.The other three groups of rats underwent ovariectomy via both sides of the back first,followed by 5consecutive days after the operation for vaginal cytology smears of rats,and rejected the unqualified rats with estrous cycle changes,and then the vaginal smears for qualified animals After the Hormone simulated pregnancy（HSP）was performed,the model was created by the withdrawal method.After the completion of the model building,corresponding drug interventions were given respectively.OFT,Sucrose preference test（SPT）and Forced swimming test（FST）were performed on the 7th and14th day of drug intervention to observe the behavioral changes of experimental rats.After the drug intervention,6 rats were randomly selected to infuse the brains with paraformaldehyde,and the obtained brain tissues were respectively used for hematoxylin-eosin（hematoxylin-eosin,HE）staining to examine the morphology of the frontal lobe neurons of the rats.In situ terminal transferase labeling（Terminal deoxynucleotidyl transferase-mediated d UTP nick-end labeling,TUNEL）method was used to detect neuronal cell apoptosis in the frontal lobe of rats.The brains of the remaining rats in each group were taken on ice,and the obtained brain tissues were determined by Western Blot（WB）for the content of GABA-related molecules in the frontal lobe of rats.The final result is quantified by Image J software,and the data obtained is statistically analyzed and graphically drawn by prism software and SPSS software.One-way analysis of variance（One-way Analysis of Variance,one-way ANOVA）is used for the comparison of multi-sample means,and the least significant difference method（Learned-Significant Difference,LSD）is used for multiple samples if the data is tested for homogeneity of variance.For pairwise comparisons,non-parametric rank sum tests are used for uneven variances.The significance level isα=0.05,and P<0.05 is statistically significant.Result:1 Model copy results1.1 Behavioral results:Compared with the blank control group after modeling,the OFT horizontal score,vertical score and total score of each modeling group（model group,E₂group and Shenguiren group）were significantly reduced,and the difference was statistically significant.（P<0.05）,there was no statistical difference in the sham operation group,the result suggested that the model was successful.1.2 HE results:Compared with the blank control group,5 samples of the frontal lobe neurons in the model group showed slight（+）degeneration and necrosis,while the sham operation group had no obvious pathological changes.It suggests that the animals in the model group have obvious brain tissue damage.1.3 TUNEL results:Compared with the blank control group,the neuronal apoptosis percentage in the frontal lobe tissue of the model group was significantly higher（P<0.05）,while the sham operation group had no statistical difference.It suggested that the animals in the model group had obvious brain tissue damage.1.4 WB results:Compared with the blank control group,the expression of GABAG2protein in the frontal lobe of the model group was significantly increased（P<0.05）;the expression of GABAG2 protein in the frontal lobe of the model group was significantly increased（P<0.05）;The expression of GABAα1 protein in the frontal lobe of rats in the group was significantly increased（P<0.05）.2 Results of drug intervention2.1 OFT results2.1.1 Level score:Compared with the model group,there was no difference in the level score of each group after one week of administration;compared with the model group after two weeks of administration,the level score of the Shenguiren group was significantly higher（P<0.05）,and There was no statistical difference between the blank group,the sham operation group and the E₂group.2.1.2 Vertical score:Compared with the model group after one week of administration,the blank group,E₂group and Shenguiren group had significantly higher vertical scores（P<0.05）,while there was no statistical difference in the sham operation group;Compared with the model group after one week,the blank group and the Shenguiren group had significantly higher scores in the vertical group（P<0.05）,while there was no statistical difference between the sham operation group and the E₂group.2.1.3 Total score:One week after administration,compared with the model group,the total score of each group has no statistical difference;two weeks after administration,compared with the model group,the level score of Shenguiren group was significantly higher（P<0.05）,While there was no statistical difference between the blank group,the sham operation group and the E₂group.2.2 SPT results:Compared with the model group after one week of administration,the sugar and water consumption of the blank group was significantly higher（P<0.05）,but there was no statistical difference between the sham operation group,the E₂group and the Shenguiren group;Compared with the model group,the consumption of sugar and water in the blank group,sham operation group,E₂group and Shenguiren group was significantly higher（P<0.05）.The consumption of sugar and water in the Shenguiren group after two weeks of administration was significantly higher than that of one week of administration（P<0.05）,and there was no statistical difference in the consumption of sugar and water in the other groups after two weeks of administration compared with that of one week of administration.2.3 FST results:Compared with the model group after one week of administration,the forced swimming immobility time of the blank group,E₂group and Shenguiren group was significantly lower（P<0.05）,and there was no statistical difference in the sham operation group.Compared with the model group after two weeks of administration,the time of forced swimming immobility was significantly lower in the blank group,sham operation group,E₂group and Shenguiren group（P<0.05）.2.4 HE staining results:Compared with mild（+）degeneration and necrosis of some neurons in the 5 samples of the model group,the frontal lobe neurons of the E2 group and the Shenguiren group only showed mild pyknosis,and the staining was deepened,suggesting that the E2 group and the Shenguiren group Group intervention measures have a good effect on improving the brain tissue damage of model animals.2.5 TUNEL results:Compared with the model group after the drug intervention,the percentage of apoptosis in the frontal lobe of the rats in the blank group,sham operation group,E₂group and Shenguiren group was significantly lower,which was statistically significant（P<0.05）.Suggesting that the treatment of Shenguiren group and E₂group can significantly improve cell apoptosis in the frontal lobe tissue of model rats.2.6 WB results:The protein quantitative results are:（1）Compared with the model group,the GABAG2 protein expression of rats in the blank group,sham operation group,E₂group and Shenguiren group was significantly reduced,and the difference was statistically significant（P<0.05）;（2）Compared with the model group,the GABA receptor-related protein（GABARAP）expression in the blank group,the Shenguiren group and the E₂group was significantly reduced,and the difference was statistically significant（P<0.05）.The GABARAP protein expression in the sham operation group was There was no difference with the model group;（3）Compared with the model group,the expression of GABAα1 protein in the blank group,the Shenguiren group and the E₂group was significantly reduced,and the difference was statistically significant（P<0.05）,while the GABAα1 protein in the sham operation group The expression is no different from the model group.Conclusion:（1）Shenguiren mixture can improve the postpartum depression behavior of rats,and its effect is more obvious after two weeks of administration;（2）Shenguiren mixture can improve the number and morphology of rat frontal lobe neurons,and reduce neuronal cell apoptosis;（3）Shenguiren mixture may improve postpartum depression by regulating the expression of synaptic GABA-related molecules.