Effects Of Anshen Yizhi Method On Behavior And Aβ Expression In AD Zebrafish Model

Objective:Starting from the choroid of the self-development of traditional Chinese medicine theory,this paper combs the understanding of AD in ancient traditional Chinese medicine,and discusses the theoretical connotation of TCM in the prevention and treatment of AD by tranquilizing the mind and benefiting intelligence.Taking zebrafish as an animal model,Shenghui Decoction,the representative of Anshen Yizhi method,was selected for experimental research to explore the possible mechanism of its prevention and treatment of AD from the aspects of behavior,histopathology and molecular biology,in order to provide reference for the in-depth research and clinical practice of AD in TCM.Method:1.Theoretical research:consult and sort out relevant literature,summarize and analyze the theoretical exposition of ancient doctors on AD,focus on combing the etiology,pathogenesis and treatment methods of AD,and explore the rationality of the treatment of AD by Anshen Yizhi method combined with the treatment and prescription analysis of Anshen Yizhi method.2.Experimental study:eight month old wild-type AB zebrafish were randomly divided into Control group,model group,Donepezil group and Shenghui Decoction group.Except for the Control group,the other groups were soaked in Al Cl₃ for 30 days to establish AD zebrafish model.After the modeling,the Control group and model group were given conventional fish culture water,Donepezil group and Shenghui Decoction group were given Donepezil solution and Shenghui decoction solution respectively,Administration for 14 days.T-maze test was used to detect the learning and memory ability of zebrafi sh in each group,and to evaluate the effect of Anshen Yizhi method on the behavior of AD zebrafish model.The content of APP and Aβ_1-42in the brain tissue of zebrafish in each group was detected by ELISA,and the number of Neu N and GFAP positive cells in the brain of zebrafish in each group was observed by immunohistochemical staining to evaluate the effect of Anshen Yizhi method on the pathology of AD zebrafish model.Western blot was used to detect the protein expression levels of PI3K,Akt,p-Akt and m TOR in the brain tissue of zebrafish in each group,and to evaluate the effect of Anshen Yizhi method on the expression of genes related to PI3K/Akt/m TOR signal pathway in AD zebrafish model.Result:1.Results of Experiment 1 T-maze experiment:compared with the Control group,the latency of zebrafish entering the EC area in the model group increased（P<0.05）,and the cumulative residence time in the EC area decreased（P<0.05）.The difference was most significant on the 4th day（P<0.01）.2.Results of Experiment 2 T-maze experiment:compared with the model group,the latency of zebrafish entering EC area in Donepezil group and Shenghui Decoction group was significantly shorter（P<0.01）.From the perspective of exploration path,compared with the Control group,the exploration path of zebrafish in the model group in the T-maze is disordered and lack of purpose,so it is difficult to find the EC area quickly;compared with the model group,the exploration path of zebrafish in Donepezil group and SHD group is relatively simple and direct,and can enter the EC area quickly and accurately.3.Results of Experiment 3（1）ELISA detection:compared with the Control group,the content of APP in zebrafish brain tissue in the model group increased（P<0.05）,the content of Aβ_1-42 increased significantly（P<0.01）.Compared with the model group,the content of APP in zebrafish brain tissue in Donepezil group and Shenghui Decoction group decreased significantly（P<0.01）,the content of Aβ_1-42 decreased in varying degrees（P<0.05,P<0.01）.（2）Immunohistochemical staining:compared with the Control group,the number of Neu N and GFAP positive cells in zebrafish brain tissue in the model group decreased significantly（P<0.01）,and the number of Neu N and GFAP positive cells in zebrafish brain tissue in Donepezil group and Shenghui Decoction group increased（P<0.05）,the difference was statistically significant.4.Results of Experiment 4 Western blot detection:compared with the Control group,the expression levels of PI3K,Akt,p-Akt and m TOR proteins in the brain of zebrafish in the model group were significantly lower（P<0.01）.Compared with the model group,the expression levels of PI3K,Akt,p-Akt and m TOR protein in zebrafish brain tissue in Donepezil group and Shenghui Decoction group increased in varying degrees（P<0.05,P<0.01）.Conclusion:1.Aged and senescent,mental failure,and Shen apraxia is easy to trigger Alzheimer’s disease.The onset of of AD is closely related to"Shen"and"Zhi".Anshen Yizhi method is an important method for the prevention and treatment of AD.2.The AD zebrafish model was established by soaking adult zebrafish in aluminum trichloride solution,which is reproducible.3.Shenghui Decoction,the representative of Anshen Yizhi method,can improve the learning and memory ability and cognitive impairment of AD zebrafish model.4.Anshen Yizhi method can reduce the accumulation of abnormal Aβin the brain of AD zebrafish model,inhibit the toxic injury of Aβand promote the repair and regeneration of neurons.5.Anshen Yizhi method may affect the pathological changes of AD by activating PI3K/Akt/m TOR pathway,showing neuroprotective effect.