| Objective Prepare an animal model of depression is prepared with chronic unpredicted mild stress(CUMS).We will investigate the effects of fluoxetine on Notch signaling pathway and relevant behavioral changes in rats model of depression through detecting the expressions of Notch signaling pathway-related proteins MAP-2,Caspase-3,Notch-1,and Jagged-1.Methods 1.Animal model and grouping: 120 healthy male SPF SD rats(150g-170g)wererandomly divided into control group,depression model group(depression group),and drug intervention group(intervention group),40 rats in each group.CUMS method was employed to established a rat model of depression,and rats in the control group were grasped once at the same time as the model group,and no other operations needed.The experimental rats were sacrificed at 1 day,7 days,14 days and 28 days after the drug intervention treatment.2.Test methods: 1 Morphological changes of the brain tissue from the experimental rats were observed using HE staining method;2 The localization and expression of MAP-2,Caspase-3,Notch-1 and Jagged-1 proteins were measured with immunohistochemistry method;3 The protein quantification of MAP-2,Caspase-3,Notch-1,Jagged-1 determined with Western blotting method;4 The changes of rats’ preference for sugar water was detected with sugar water preference test;5 The water contentof the brain tissue in rats were measured with dry-wet proportion method;6 The changes of motor activity and anxiety behavior of depressed rats wasdetected with open field test;7Spatialcognitive ability of experimental rats was measured with water maze test.3.Statistical analysis: Image-Pro Plus 6.0 image analysis software was used to conduct semi-quantitative analysis.expressed as integrated optical density value(IOD value)For protein quantification,using Image J analysis processing software to perform semi-quantitative analysis on the bands reacted by the protein to obtain the optical density value(OD value).the OD ratio of the target protein to the internal reference(β-actin)as basic control.Excel was used to maintain the experimental data,and the data was represented by mean ± standard deviation(x ± s),SPSS 16.0 software was employed to analyse the data.Comparisons more than 3 groups were conducted by one-way analysis ovariance followed by the post-hoc test.The comparison between the two groups was performed by t test.P values less than 0.05 were considered to indicate a statistically significant difference between 2 groupsResults 1.HE staining result: the neuronal cells of the rats in the depression group showed obvious pathological changes in the cell body,the cytoplasmic staining of the cells was significantly weakened,the nucleus of the cells could be seen with deep staining,and necrotic neuronal cells exist;while,the pathological changes of the cells was significantly improved in the intervention group;2.Immunohistochemical results:MAP-2,Caspase-3,Notch-1,and Jagged-1 were localized in cytoplasm,while MAP-2,Caspase-3,Notch-1,and Jagged-1 were only weakly expressed in the control group,with light brownish yellow color.Compared with the control group,the positive expression in the depression model group was enhanced at each time point,and the immune reactivity was stronger(P<0.05).Compared with the depression model group,the corresponding positive expression in the drug intervention group showed a weakening trend(P<0.05),but there is no change in the overall expression trend;3.Western blotting results: The results showed that the MAP-2,Caspase-3,Notch-1 and Jagged-1 proteins in the control group were only weakly expressed,and compared with the control group,the expression of MAP-2,Caspase-3,Notch-1 and Jagged-1 in the depression group was significantly increased(P<0.05).Compared with the depression group,the protein expression intensity of the drug intervention group was significantly decreased(P<0.05),but still higher than that of the control group;4.Sugar water preference test: There was no significant change in the consumption of ordinary water between the normal control group and the depression model group.Compared with normal control group,the sugar water consumption and the sugar water preference percentage of the experimental rats in the depression model group were lower,and the difference has statistics significance(P<0.05).Compared with depression model group,the consumption of sugar water and the sugar water preference percentage in the drug intervention group were significantly improved(P<0.05);5.Brain tissue water content test: water contentof the brain tissue of depressed rats in the control group was relatively stable at each time point.The water content of the brain tissue of the rats in the depression group also increased significantly over time(P<0.05).Compared with the depression group,the brain water content of the intervention group showed a clear downward trend,but still higher than that of the control group(P<0.05);6.Open field test result: the total walking distance,the activity duration in the central grid,the number of standing up and the number of modification behaviors of the experimental rats in the depression model group were less than those in the normal control group(P<0.05);compared with the depression group,the data of intervention group had significant improved(P<0.05);7.Water maze test result:Rat’s exercise ability will be affected after depression,so the experimental operation of each group is scheduled to be carried out on the 6th,7th,8th and 9th days after the depression model.According to test result,the time spent on searching for safe islands in the depressed group is much higher,and the difference has statistics significance(P<0.05);afterdrug treatment,the time spent on searching for the safety platform on the8 th and 9th day was less than that in the depression group,and the difference has statistics significance(P<0.05).Conclusion After successful preparation of a rat model of depression,the expression of Notch signaling pathway-related proteins was detected after fluoxetine intervention,indicating that the Notch signaling pathway was related to the pathogenesis of depression in rats,the cognitive function and behaviors of depressed rats might be effectively improved.Figure[17];Table[6];Reference [134]... |
