| ObjectiveGinseng, the root of Panax ginseng C.A. Meyer (Araliaceae), is one of the most famous and valuable traditional herbal medicine. Its tranquilizing function was recorded in Sheng Nong’s Herbal Classic. Ginseng total saponins (GTS) are the principal bioactive ingredients behind claims of ginseng efficacy. Recently, ginseng and ginsenosides are reported to have antidepressant or anti-stress effects. In the present study, the depression model is induced by repeated corticosterone injections, the antidepressant efficacy of GTS is investigated basing on neuroplasticity, and the underlying mechanism is explored with respect to the CREB-BDNF signaling pathway.MethodsExperiment1:Preparation of higher-purity GTSGTS were efficiently purified by dynamic anion-cation exchange following removal of hydrophilic impurities by macroporous resin D101.Experiment2:Effects of stress on depression-like behavior and hippocampal structural plasticityMice were assigned to6groups which received repeated injections of vehicle or20mg/kg/d of corticosterone for1,3or5weeks. The groups were named Control1w, Corticosterone1w, Control3w, Corticosterone3w, Control5w, Corticosterone5w. Body weights, serum corticosterone levels, depression-like behavior and hippocampal structural plasticity were assessed. Forced swimming test (FST) and tail suspension test (TST) were used to indicate depression-like behavior. Hippocampal protein levels of neurofilament light chain (NF-L), synaptophysin (SYP) and brain-derived neurotrophic factor (BDNF) were assessed by Western blot to indicate structural plasticity.Experiment3:Effects of GTS on depression-like behavior and hippocampal structural plasticity in the mouse depression modelGrouping:the control group, the corticosterone (CORT) group, the corticosterone+10mg kg-1d-1fluoxetine (CORT+FLU) group, the corticosterone+50mg kg-1d-1GTS (CORT+ GTSH) group, the corticosterone+25mg kg-1d-1GTS (CORT+GTSM) group, the corticosterone+12.5mg kg-1d-1GTS (CORT+GTSL) group. FST and the TST were performed on day20and day21, respectively. On day22,6hours after the last injection of corticosterone, venous blood samples were collected for assessing serum corticosterone before mice were sacrificed. Hippocampal mRNA and protein levels of NF-L and BDNF were assessed by Real-time PCR and Western blot. Immunohistochemisty staining was employed to indicate BDNF expression in CA1region of hippocampus.Experiment4:Effects of GTS on depression-like behavior and hippocampal structural plasticity in normal miceGrouping:the control group,10mg kg-1d-1fluoxetine (FLU) group,50mg kg-1d-1GTS (GTSH) group,25mg kg-1d-1GTS (GTSM) group,12.5mg kg-1d-1GTS (GTSL) group. FST and the TST were performed on day20and day21, respectively. Mice were sacrificed on day22for assessing hippocampal protein levels of NF-L and BDNF by Western blot.Experiment5:Antidepressant mechanism of GTS with respect to the cAMP response element binding protein (CREB)-BDNF signaling pathwayThe grouping method is the same as Experiment3. Mice were sacrificed on day22for assessing hippocampal protein levels of CREB, p-CREB (Ser133), glycogen synthase kinase3β (GSK-3β), p-GSK-3β (Ser9) and phosphor-protein kinase A Catalytic Subunit (p-PKA C)(Thr197) by Western blot.ResultsExperiment1:The estimated contents of total saponins were107%according to the colorimetric method. The total amount of representative ginsenosides Re, Rd, Rgl, and compound K was approximately22%based on ultrahigh-performance liquid chromatography-charged aerosol detector quantitative analysis.Experiment2:Compared with the respective contol groups, all corticosterone groups had lower body weights (p<0.01) and higher serum corticosterone levels (p<0.01); the immobility time was prolonged in Corticosterone3w and Corticosterone5w (p<0.05,p<0.05, respectively) during FST, and in Corticosterone1w, Corticosterone3w and Corticosterone5w (p<0.05,p<0.05,p< 0.05, respectively) during TST; hippocampal protein levels of NF-L and BDNF were higher (p<0.05,p<0.05, respectively) in Corticosterone1w; hippocampal protein levels of NF-L, SYP and BDNF were lower in Corticosterone3w (p<0.05, p<0.05, p<0.01, respectively) and Corticosterone5w (p<0.05,p<0.05,p<0.05, respectively).Experiment3:The serum corticosterone level increased significantly in the CORT group, as compared with the control group (p<0.01). Compared with the CORT group, all three doses of GTS had no significant effect. The immobility time prolonged during FST and TST in the CORT as compared with the control group (p<0.01). The forementioned immobility time was almost reversed by simultaneous treatment with GTSH (p<0.05,p<0.01, respectively) or GTSM (p<0.01,p<0.01, respectively). GTSH and GTSM reversed the decreased levels of BDNF mRNA (p<0.01,p<0.05, respectively), BDNF protein (p<0.01,p<0.05, respectively), NF-L mRNA (p<0.01, p<0.05, respectively), and NF-L protein (p<0.01,p<0.05, respectively) induced by repeated corticosterone injections. GTSL also reversed the corticosterone-induced lower level of BDNF mRNA (p<0.05). GTSH and GTSM reversed the corticosterone-induced lower expression of BDNF in CA1region (p<0.05).Experiment4:The immobility time was prolonged in GTSM during FST (p<0.05), and in GTSH and GTSM during TST (p<0.05, p<0.01, respectively). Hippocampal protein levels of NF-L and BDNF in GTSH, GTSM or GTSL differed insignificantly compared with the control group.Experiment5:Hippocampal protein levels of CREB and p-PKA C in GTSH, GTSM or GTSL differed insignificantly compared with the control group. GTSH, GTSM and GTSL reversed the decreased hippocampal protein levels of p-CREB (p<0.01,p<0.01,p<0.05, respectively) and p-GSK-3p (p<0.01,p<0.01, p<0.05, respectively) induced by repeated corticosterone injections, but had no significant effect on the corticosterone-induced higher GSK-3β level.ConclusionThe structural plasticity of the adult hippocampus is critical for the action of antidepressants and the underlying pathophysiology of depression. In the corticosterone-induced mouse depression model, certain doses of GTS exhibit antidepressant-like activities by reversing the decrease of some plasticity-related proteins and activating the CREB-BDNF signaling pathway in hippocampus. The promotion of GSK-3β inhibitory phosphorylation which activates the CREB-BDNF signaling pathway, may account for the antidepressant-like activity of GTS. |
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