| Alzheimer’s disease(AD)is an age-related neurodegenerative disease characterized by progressive memory and cognitive dysfunction.Its pathogenesis is unclear,in which "Aβcascade theory" has been accepted by most scholars.Synaptic plasticity is not only the molecular basis of learning and memory,but also the molecular mechanism of signal transduction of nervous system diseases.The long-term potentiation(LTP)is an objective electrophysiological index of neuronal physiological activity in the process of synaptic plasticity and learning and memory formation.LTP refers to the long-term enhancement of synaptic transmission after the tetanic stimulation or other special stimulation.It has been clarified that small molecule soluble Aβ oligomers have inhibited the formation of animal memory and LTP production before the formation of senile plaque.Kai Xin San(KXS)was first appeared in<Qianjin Yaofang>by Sun Simiao in Tang dynasty.KXS is composed of Ginseng Radix et Rhizoma(Ren-Shen,Root and rhizome of Panax ginseng C.A.Mey.),Polygalae Radix(Yuan-Zhi,root of Polygala tenuifolia Wild.),Acori Tatarinowii Rhizoma(Shi-Chang-Pu,rhizome of Acorus tatarinowii Schott),and Poria(Fu-Ling,sclerotium of Poria cocos(Schw.)Wolf).which have a wide range of pharmacological effects particularly in promoting intelligence,anti-aging and anti-oxidantion.Studies have shown that the role of KXS in promoting intelligence may be depend on the direct effects in the the physiological process of central learning and memory,especially on the synaptic transmission process related to learning and memory of hippocampus region.However,its exact mechanism of action and active ingredients are not entirely clear.Polygalae Radix is commonly used in ancient prescriptions which has a significant role in promoting intelligence.The active ingredient study of KXS showed that the drug-like effective compounds of KXS are mainly from Polygalae Radix,and the compounds absorbed into the blood are Polygala oligosaccharide ester in the majority.Polygala oligosaccharide esters also have the neuroprotective effects.However,it is still not clear whether the main role of the Poly gala oligosaccharide ester and Polygalae Radix in KXS.The aim of this study was to investigate the effect and mechanism of KXS on synergistic plasticity of APP/PS1 double transgenic mice,and further analysize the role of Polygalae Radix and Polygala oligosaccharide ester in KXS.1 Effects of KXS,KXS without Polygalae Radix and Polygala oligosaccharide esters on long-term potentiation in APP/PS double transgenic miceObjective:To investigate the effect of KX on LTP in APP/PS1 double transgenic mice and anylaysize the role of Polygala and Polygala oligosaccharides ester in KXS.Methods:3-month-old male APP/PS double transgenic mice were randomly divided into the model control group and Donepezil 0.75 mg·kg-1·d-1,KXS 3.0 g·kg-1·d-1、1.5 g·kg-1·d-1,KXS without Polygala 2.6 g·kg-1·d-1、1.3 g·kg-1·d-1,polygala oligosaccharide esters 0.1 g·kg-1·d-1、0.05 g·kg-1·d-1 groups.Each administration group APP/PS double transgenic mice were orally administered for 3 months,respectively.C57/BL6J mice were used as the normal control group.Mice were positioned on a stereotaxic frame after being anesthetized.A bipolar stimulating electrode and the recording electrode was positioned in the precise positioning points(PP-DG),then LTP were induced and recoreded by adjusting the stimulus intensity.Results:LTP tests showed that the fEPSPs slopes were significantly decreased in APP/PS1mice compared with control group mice(P<0.01).KXS、KXS without Polygalae Radix and polygala oligosaccharide esters and Donepezil treatment significantly enhanced the fEPSPs slopes in the APP/PS double transgenic mice(P<0.05).And the effect of KXS is the most obvious.Conclusion:The result showed that KXS can increase the fEPSPs slopes in the APP/PS double transgenic mice,suggesting that KXS has the enhancement on synaptic plasticity.Polygalae Radix has the weak effect on LTP,and polygala oligosaccharide esters are involved in the effect of KXS on LTP,but maybe not the mainly active ingrediects or synergistic reaction with other compounds of KXS.2 Effects of KXS,KXS without Polygalae Radix and Polygala oligosaccharide esters on expression of Synaptic Plasticity-related Protein in APP/PS1 Double Transgenic MiceObjective:To investigate the mechanisms of KXS on the synaptic plasiticity through detecting the synaptic plasticity-related proteins expression in the hippocampus of APP/PS double transgenic mice,and to explore the role of Polygalae Radix and Polygala oligosaccharide esters in the effect of KXS on the synaptic plasiticity.Methods:According to the above-mentioned grouping and administration methods,the mice were anesthetized and fixed with 4%paraformaldehyde by heart,and then the brain was taken and fixed in 4%paraformaldehyde,embedded and cut into slicies.Immunohistochemical method was used to detect the expression of synaptic related protein PSD-95 in brain of APP/PS double transgenic mice.After LTP determination,the mice were decapited,the hippocampus were separated and then frozen at-80℃.the expression of P-AKT,AKT,AMPAR,P-CaMKⅡ,CaMKⅡ,P-MAPK and MAPK in hippocampus of APP/PS double transgenic mice by Western-blot.Results:Immunohistochemical results showed the average optical density of PSD-95 positive cells of CA1 area and cerebral cortex in APP/PS double transgenic mice was obviously less than that in control group,with statistical significance(P<0.05).Donepezil increased the average optical density of PSD-95 positive cells in the CAI area(P<0.05),KXS 1.5 g·kg-1·d-1 increased the average optical density of PSD-95 positive cells in the CA1 and cerebral cortex(P<0.05).KXS 3 g·kg-1·d-1 increased the average optical density of PSD-95 positive cells in the cerebral cortex(P<0.05).KXS without Polygala 2.6 g·kg-1·d-1 of increased the average optical density of PSD-95 positive cells in the cerebral cortex(P<0.05).KXS without Polygala 1.3 g·kg-1·d-1 and increased the average optical density of PSD-95 positive cells in the CA1(P<0.05).Polygala oligosaccharide esters increased the average optical density of PSD-95 positive cells in the CA1 and cerebral cortex(P<0.05).The expression of AMPAR in the hippocampus of APP/PS double transgenic mice was obviously less than that in control group,with statistical significance(P<0.05).KXS 3 g·kg-1·d-1 increased the expression of AMPAR in the hippocampus of APP/PS double transgenic mice(P<0.05).KXS without Polygala Radix、Polygala oligosaccharide esters can increase the expression of AMPAR in the hippocampus of APP/PS double transgenic mice,but there are no obvious statistical significance.Compared with control group,the expression of P-CaMKII in the hippocampus of APP/PS double transgenic mice was obviously decreaed,with statistical significance(P<0.05).KXS and Donepezil increased the amount of P-CaMKII in the hippocampus of APP/PS double transgenic mice(P<0.05).KXS without Polygalae Radix and Polygala oligosaccharide esters have the increasing trend on the expression of P-CaMKII in the hippocampus of APP/PS double transgenic mice.Compared with control group,the expression of AKT in the hippocampus of APP/PS double transgenic mice was obviously decreased(P<0.05).KXS、KXS without Polygalae Radix、Polygala oligosaccharide esters could significantly increase the expression of AKT in the hippocampus of APP/PS double transgenic mice(P<0.05).Conclusion:KXS has the enhancement of synaptic plasticity,which maybe related with the up-regulation of the expression of plasticity-related proteins(AMPAR,P-P-CaMKII and AKT)of APP/PS1 transgenic mice.The effect of KXS without Polygalae Radix on the aboved protein expression is weaker than that of KXS,and the Polygala oligosaccharide esters has the significant effect on the expression of AKT and PSD-95,suggesting Polygala oligosaccharide esters are the active indigrents in KXS,and Polygalae Radix is involed in the effects of KXS on these protein expression.3 Effects of KXS,KXS without Polygalae Radix and Polygala oligosaccharide esters on expression of the mRNA of Syn and GAP-43 in APP/PS1 Double Transgenic MiceObjective:To investigate the effects of KXS on the expression of the mRNA of Syn and GAP-43 in the hipocampus of APP/PS double transgenic mice,and further analysize the role of Polygalae Radix and Polygala oligosaccharide esters in the effect of KXS on the synaptic plasiticity.Method:After LTP determination,the mice were decapited,the hippocampus were separated and then frozen at-80℃.RT-PCR was used to detecte the expression of synaptic protein Syn and GAP-43 mRNA in brain of APP/PS double transgenic mice.Results:The results showed that there was no significant difference in the expression amount of Syn mRNA between the control group and the APP/PS1 model group.KXS,KXS without Polygalae Radix and Polygala oligosaccharides ester can increasing the expression of Syn mRNA in the brain of APP/PS1 model group,with statistical significance(P<0.05).There is no significant difference between KXS and KXS without Polygala.Compared with the control group,the expression of GAP-43mRNA in brain of APP/PS1 mice was decreased,but there was no significant difference.There is no significant difference on the the expression of GAP-43 mRNA in the hippocampus of APP/PS1 double transgenic mice treated with between KXS、KXS without Polygalae Radix and Polygala oligosaccharides ester compared with APP/PS1 model group.Conclusion:KXS can promote the expression of Syn mRNA in APP/PS double transgenic mice,its effect is no more than KXS without Polygalae Radix,and Polygala oligosaccharides esters can promote the expression of Syn mRNA.These results suggested that Polygala and Polygala oligosaccharides ester are involed in KXS enhanceing the expression of Syn,which may be not the most important indigrent,but a synergistic reaction with other compounds of KXS.In summary,KXS can enhance the synaptic plasticity of APP/PS1 double transgenic mice which is associated with regulation of PSD-95,AMPAR,P-CaMKII,and AKT protein expression and Syn mRNA expression in hippocampus of APP/PS1 mice.Compared with KXS,the effects of KXS without Polygalae Radix are less than that of KXS,but there are no statistic significance.Polygala oligosaccharides esters showed the enhancment of LTP and are involved in regulating the expression of proteins above.These resultes indicated that Polygalae Radix and Polygala oligosaccharides esters are involed in KXS’s effect on the synaptic plasticity of APP/PS1 mice.They maybe exert effects through synergies with other components in KXS. |
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