| ObjectivesIn this research,the chronic unpredictable mild stress(CUMS)model was conducted to observe the effect of the ?-asarone on the synaptic plasticity related factors GAP-43,SYN and PSD-95 expressions and reveal its antidepressed mechanism,which would provide a new insight into the development of new antidepressants.MethodsThe clean-grade male SD rats of 3 to 4 months old were allowed to acclimate for a week.By open-field test,100 SD rats were randomly assigned into control group,model group,the fluoxetine hydrochloride group(10mg/kg/d),the low dosage ?-asarone group(25mg/kg/d)and high dosage ?-asarone group(50mg/kg/d).Except the control group,the rats in other groups were fed separately.And then,in the 0 d,28 d and 29d before the model-building,Open Field test,sugar water preference test(SPT)and forced swimming test(FST)were performed respectively to evaluate the effect of CUMS and ?-asarone.The Nissl staining was used to investiagate the changes of the Nissl bodies in each groups;HE staining was conducted to observe the changes of cell morphology;Immuno-fluorescence technique was performed to measure the GAP-43,SYN and PSD-95 protein expressions;And Western Blot and RT-PCR were adopted to observe the levels of GAP 43,SYN,PSD-95 protein and mRN A.Results1.The behavioral study showed that when compared with control group,the horizontal and vertical activity and the preference of sugar water of the model group were reduced,and the immobile time in the forced swimming was extended(P<0.05)after a period of 28d CUMS.But the low dosage ?-asarone(25mg/kg/d),high dosage ?-asarone(50mg/kg/d)and fluoxetine hydrochloride(10mg/kg/d)showed a significant increase in the preference of sugar water and the scores of locomotor activity,but shortened the time of immobility(P<0.05).2.Nissl staining showed that,observe the dyeing under optical microscope,the blue nissl bodies in hippocampal CA3 neurons of model group were decresed and cell morphology was changed,including the edema of axon,vacuoles in the neuronal cytoplasm and nucleolar pycnosis,which showed that after CUMS,the hippocampus neurons in model rats were damaged by some traumatic changes like degeneration.In ?-asarone group(25mg/kg/d)and ?-asarone group(50mg/kg/d),the neuron morphology in hippocampal CA3 area was normal relatively and the nissl bodies were more than that in model group,which was similar with the effect of positive control drug fluoxetine hydrochloride(lOmg/kg/d).3.The HE staining results showed that,the number of neurons decreased significantly in hippocampal CA3 area of model group,and the arrangement was disordered,cell morphology was loose and poor.Some neurons were swell,some became atrophy and degenerate,simultaneously pycnosis existed in nucleus,vacuoles in loose cytoplasm and the number of dendrites were decreased.In low dosage?-asarone group,high dosage ?-asarone group and the fluoxetine hydrochloride group,the neuron morphology in hippocampal CA3 area were normal relatively,the cell numbers were more than that in model group,and the cell arrangement was relatively neat,structures were integral,and the dendritic number was increased.4.Immunofluorescence technique was conducted to detect the expressions of GAP-43,SYN and PSD-95 of each group in rats hippocampal CA3 area.The result found that the GAP-43-positive cells numbers in model group was more than in other groups with statistical significance(P<0.05),which showed the rats' hippocampal neurons were damaged in model group,and low dosage and high dosage ?-asarone could reduce the expression of GAP-43(P<0.05),which was similar with the treatment of fluoxetine hydrochloride group(10mg/kg/d).When compared with control group,the number of SYN and PSD-95 positive cells in rats hippocampal in CA3 area of model group were reduced(P<0.05),but this effect was reversed by the treatment of the low dosage ?-asarone,the high dosage ?-asarone group and fluoxetine hydrochloride(P<0.05).5.Western blot and RT-PCR methods were used to investigate the protein and mRNA levels of GAP-43,SYN,PSD-95.According to the result,the expression of GAP-43 was up-regulated in model group(P<0.05),but the treatments of low dosage ?-asarone(25mg/kg/d)and high dosage ?-asarone(50mg/kg/d)blocked this up-regulation(P<0.05),which was similar with the treatment of fluoxetine hydrochloride.The expressions of SYN,PSD-95 were down-regulated in model group,but up-regulated by the treatments of low dosage?-asarone(25mg/kg/d),high dosage ?-asarone(50mg/kg/d)and fluoxetine hydrochloride(P<0.05).Conclusion1.?-asarone could obviously improve the depressive behavior of CUMS rats.2.?-asarone exhibited protective effect on structure and function of synaptic plasticity in hippocampal neurons of CUMS.3.The mechanism of ?-asarone on antidepressant effects partly related with the regulation of the stability of the synaptic plasticity. |
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