Effect And Mechanism Of Modified Suanzaoren Decoction On Learning And Memory Of Alzheimer’s Disease Model Rats

Objective: To observe the effect of modified suanzaoren decoction on cognitive and memory function in Alzheimer’s disease model rats and explore its possible mechanism.Methods: 72 male SD rats were randomly divided into Normal group,AD model group,Estazolam group,modified Suanzaoren decoction group(low-dose group(JWSZRT-L),medium-dose group(JWSZRT-M),high-dose group(JWSZRT-H))by random number table method,with 12 rats in each group.Rat models of Alzheimer’s disease were prepared by D-galactose and Aβ1-42.Rats were intraperitoneally injected with 5% D-galactose at A dose of 100mg/kg/ D for 42 consecutive days.On the 21 st day of the experiment,the rats were injected with Aβ1-42 in the right hippocampus,but the normal group was not treated.Modified Suanzaren Decoction low-dose,medium-dose and high-dose groups were given intragastric administration of 5.94g/kg/d,11.88g/kg/d and 23.76g/kg/d,respectively,from the beginning of modeling,and model group was given intragastric administration of 0.9% sodium chloride for 42 consecutive days.The esazolam group was given 0.1mg/kg/d by gavage on the 21 st day of modeling,i.e.,the 2nd day after injection of Aβ1-42,for 21 consecutive days.No medication was given to the normal group.Morris water maze test was used to evaluate the cognitive and memory ability of rats,and the autonomous activity of rats was detected by the multi-functional automatic activity recorder.The body temperature of the rats was measured at 6:00,12:00,18:00 and24:00.Take the rat’s serum,cerebrospinal fluid,hippocampus and hypothalamus samples,ELISA method to detect the rat serum 5-HT,5-HIAA,IL-1 beta,TNF alpha,cerebrospinal fluid(MT),Real time PCR detection-Bmal1,Clock mRNA expression in hypothalamus,Western blot detection the expression of rat hippocampal CX3CL1,immunohistochemical detection of the expression of rat hippocampal tissue Tauser199,MTR,HE dyeing observation pathological changes of hippocampus in rats.Results:1.Changes in spontaneous activity and body temperature rhythm of rats Compared with Normal,the number of spontaneous activity of AD model rats increased(P < 0.01);Compared with AD model,the autonomous activity times of Estazolam,JWSZRT-L,JWSZRT-M and JWSZRT-H rats were significantly decreased(P < 0.01),and there was no significant difference between each treatment group.After modeling,the body temperature of AD model rats changed greatly at different time points,and the rhythm was disturbed,and the body temperature increased significantly at 12:00(P < 0.01).All treatment groups maintained normal body temperature rhythm,with the lowest body temperature at 12:00 and the highest body temperature at 24:00.2.Effects of modified Suanzaoren Decoction on learning and memory function of AD model rats2.1 Positioning navigation experiment:With the increase of training times,the onstage latency of rats in each group was gradually shortened;Compared with Normal,the incubation period of AD model was significantly prolonged(P < 0.05,P <0.01).Compared with AD model,the onstage latency of rats in each treatment group was shortened from the 3rd day of training,and Estazolam and JWSZRT-M were more significant(P < 0.01).On the 6th day,the latency of JWSZRT-M was the shortest among the groups with different doses of modified Suanzaoren Decoction(P< 0.05).The distance to the stage of rats in each group also showed a shortening trend.Compared with Normal,the distance before AD model was significantly increased(P < 0.05,P < 0.01);From day 3,the distance before the launch of Estazolam and JWSZRT-M was significantly shorter than that of AD model(P <0.05).2.2 Space exploration experiment:Compared with AD model group,the number of crossing platform and the residence time in target quadrant of other groups were increased,and the number of crossing platform of Normal and JWSZRT-M was significantly increased(P < 0.05),and the residence time in target quadrant of Normal,JWSZRT-M and Estazolam was significantly increased(P < 0.05).2.3 Histopathological changesHE staining results of hippocampal CA1 region of rats showed that compared with the AD model,the cell structure of Estazolam,JWSZRT-L,JWSZRT-M and JWSZRT-H groups was clear,with relatively regular arrangement,and fewer nuclear swellings and fragmentation,among which Estazolam and JWSZRT-M were more similar to the normal group.Compared with Normal,the number of Tauser199 positive cells in hippocampal CA1 region of rats in AD model group was significantly increased,while the number of Tauser199 positive cells in all treatment groups was lower than that in AD model group,and the expression of Estazolam and JWSZRT-M were the least.3.Effects of Modified Suanzaoren Decoction on 5-HT,5-HIAA,MT and MTR of AD model ratsThe serum concentrations of 5-HT and 5-HIAA in AD model rats were significantly lower than those in Normal rats(P < 0.01).The serum 5-HT content of Estazolam and JWSZRT-M was significantly higher than that of AD model(P < 0.01).The serum5-HIAA content of JWSZRT-L,JWSZRT-M,Estazolam and JWSZRT-H was higher than that of AD model(P < 0.01,P < 0.05).Compared with AD mode,the MT concentrations of JWSZRT-L,JWSZRT-M,JWSZRT-H and Estazolam were all increased(P < 0.01).Among them,JWSZRT-M was most significantly up-regulated(P< 0.05).Immunohistochemical results showed that compared with Normal,MTR expression in hippocampal CA1 region of AD model group was significantly decreased.Compared with the AD model,the expression of MTR in each treatment group was significantly increased,especially JWSZRT-M.4.Effects of modified Suanzaoren Decoction on inflammation in AD model rats The expression of CX3CL1 in the hippocampus of AD model rats was significantly lower than that of Normal rats(P < 0.01).Compared with AD model,the expression of CX3CL1 in JWSZRT-L,JWSZRT-M,JWSZRT-H and Estazolam groups was significantly up-regulated(P < 0.01).Compared with Normal,the serum IL-1β and TNF-α of AD model rats were significantly up-regulated(P < 0.01),while the serum IL-1β and TNF-α of the treated group were significantly down-regulated(P < 0.01).The down-regulation of serum IL-1β and TNF-α of JWSZRT-H in the modified Suanzaren Decoction group was the least.5.Effect of Modified Suanzaoren Decoction on the clock controlling gene in hypothalamus of AD model ratsCompared with Normal,the mRNA expressions of BMAL1 and Clock in AD model rats were significantly down-regulated(P < 0.01,P < 0.05).Compared with AD model,BMAL1 mRNA and Clock mRNA expressions in each administration group were significantly up-regulated(P < 0.01,P < 0.05).The up-regulation of BMAL1 mRNA and Clock mRNA in j WSZRT-H group was small(P < 0.01).Conclusion:1.Modified Suanzaoren Decoction can improve the cognitive and memory function of AD rats.2.Modified Suanzaoren Decoction may improve the cognitive and memory function of AD rats by improving sleep,up-regulating BMAL1 mRNA and Clock mRNA expressions,promoting the secretion of MT and 5-HT,and increasing the expression of MTR;3.Modified Suanzaoren Decoction may increase the expression of CX3CL1 in the hippocampus of AD model rats,reduce the levels of IL-1β and TNF-α,reduce the inflammatory response,and thus reduce the phosphorylation of Tau protein.4.Modified Suanzaoren Decoction and estazolam both have positive effects on the improvement of sleep disorders and cognitive function in AD rats,suggesting that the use of sleep improving drugs to regulate sleep may have a certain preventive effect on the progression of AD disease.