Effects Of Exercise And Metformin On SIRT1 And Synaptic Plasticity In Chronically Stressed Mice

Background:According to the WHO,in 2016,350 million people of all ages in the world suffered from depression,and the number of depressive patients increased by about 18.4%between 2005 and 2015.It is estimated that by 2020,depression is expected to become the second largest health burden.It is found that the commonly used antidepressant drugs can increase the risk of diabetes,so it is urgent to develop antidepressant drugs or other effective interventions and treatments for metabolic diseases.In 2015,Nature reported that the Flint research team through the analysis of the gene sequence of patients with Major depressive disorder,found two depression related mutation site,one located near the SIRT1 gene,which was first found in depression related genes.This important discovery provides an important basis for the research of SIRT1 as the target of antidepressant drugs.Regular exercise can significantly improve depression-like behaviors,however,the underlying mechanism is not clear.So it is worth studying whether exercise can regulate the expression of SIRT1 and influence the synaptic plasticity.As the first-line drug for the treatment of diabetes,metformin plays a wide range of physiological effects in cancer,cognitive memory and anti-aging.However,the role is not clear in depression.Objective:The current study aims to investigate the expression of SIRT1 and synaptic plasticity associated proteins in hippocampus and prefrontal cortex of chronic unpredictable mild stress(CUMS)mouse model.On this basis,we will further explore the exercise and metformin regulating effect and molecular mechanism of depression-like behaviors.Methods:A total of 50 healthy male C57BL/6 mice,5 weeks old,weighing 18-20g,were randomly divided into five groups:Con group:normal control group;CUMS group:the experimental mice experienced seven weeks of chronic unpredictable mild stress;CUMS+Swim group:fourth weeks of CUMS began to swimming exercise;CUMS+Saline group:fourth weeks of CUMS began to intraperitoneal inject saline;CUMS+Met group:fourth weeks of CUMS began to intraperitoneal inject metformin(Met).After 7 weeks of experimental treatment,behavioral tests were performed.Mice were sacrificed and the hippocampus and prefrontal cortex were rapidly and carefully extracted.Real-time PCR were used to examine the mRNA levels of SIRT1,miRNA-124,miRNA-134,miRNA-138 in hippocampus and prefrontal cortex of mice.Western Blotting was used to examine the protein levels of SIRT1,CREB,BDNF,synaptophysin(SYN)and GAP-43 in hippocampus and prefrontal cortex.Results:(1)Compared to Con group,body weight of mice in CUMS group is significantly lower(P<0.01),sugar consumption decreased significantly(P<0.05),immobility time in forced swimming test and the tail suspension test significantly increased(P<0.05),the number of poking and reaing in open field test reduced significantly(P<0.01).But distance in center had no significant change.Compared to the CUMS group,the body weight of CUMS+Swim mice showed no significant difference in the 0～4th weeks and were significantly decreased in 5～7th weeks(P<0.01),sucrose preference had no significant difference,immobility time in forced swimming test and the tail suspension test was significantly decreased(P<0.05),the number of poking and distance in center in open field test had a very significant increase(P<0.01),and significantly increased the number of rearing(P<0.05).Compared with CUMS+Saline group,CUMS+Met group of mice weight,sucrose preference,immobility time in tail suspension test,the number of rearing and distance in center in open field test was no significant difference,immobility time in forced swimming test was significantly decreased(P<0.05),the number of poking in open field test increased significantly(P<0.05).(2)Compared with Con group,the mRNA expression of SIRT1 in hippocampus and prefrontal cortex of CUMS group was significantly lower(P<0.05),but there was no significant change of SIRT1 protein expression of CUMS group.Compared with the CUMS group,the mRNA expression of SIRT1 in hippocampus and prefrontal cortex of CUMS+Swim group significantly increased(P<0.05),the protein expression of SIRT1 in hippocampus of CUMS+Swim group increased significantly(P<0.05),no differences in prefrontal cortex.Compared with CUMS+Saline group,the mRNA expression of SIRT1 inhippocampus and prefrontal cortex of CUMS+Met group had no significant difference,the protein expression of SIRT1 in hippocampus was very significantly increased(P<0.001),significantly increased in the prefrontal cortex(P<0.05).(3)Compared to Con group,CUMS exposure significantly decreased the mRNA levels of miRNA-138 and miRNA-134 in hippocampus,as well as the protein level of CREB and BDNF(P<0.05);the mRNA levels of miRNA-124,miRNA-134 and miRNA-138 had no significant changes in the prefrontal cortex,as well as the protein level of CREB and BDNF.Compared to CUMS group,Swimming exercise significantly increased the mRNA levels of miRNA-124 in hippocampus(P<0.05),very significantly increased the mRNA levels of miRNA-134 and miRNA-138 in hippocampus(P<0.01);Swimming exercise significantly decreased the mRNA levels of miRNA-124 and miRNA-134 in prefrontal cortex(P<0.05),the mRNA levels of miRNA-138 in prefrontal cortex did not change significantly.Compared to CUMS group,Swimming exercise significantly increased the protein levels of BDNF in hippocampus(P<0.05),but no significant difference of CREB;the protein levels of CREB and BDNF in prefrontal cortex were not significantly difference.Compared to CUMS+Saline group,Metformin did not significantly change the mRNA levels of miRNA-124,miRNA-134 and miRNA-138,as well as the protein level of CREB and BDNF in hippocampus and prefrontal cortex.(4)Compared to Con group,CUMS exposure significantly decreased the protein levels of synaptophysin and GAP-43 in hippocampus(P<0.05),but no significant changes in prefrontal cortex.Compared to CUMS group,Swimming exercise significantly increased the protein levels of synaptophysin and GAP-43 in hippocampus(P<0.05),but no significant changes in the prefrontal cortex.Compared to CUMS+Saline group,Metformin significantly increased the protein levels of synaptophysin and GAP-43(P<0.05),but no significant changes in the prefrontal cortex.(5)The correlation analysis showed that immobility time in forced swimming test was inversely correlated with the mRNA levels of SIRT1 in hippocampus;immobility time in tail suspension test was inversely correlated with the mRNA levels of SIRT1 in hippocampus;the number of rearing in open field test was positively correlated with the mRNA levels of SIRT1 in hippocampus.The protein levels of SIRT1 was positively correlated with the protein levels of synaptophysin in hippocampus,and the protein levels of SIRT1 was positively correlated with the protein levels of GAP-43 in hippocampus.Conclusion:(1)Swimming and metformin can significantly improve depression-like behaviors of chronic unpredictable mild stress mice.(2)Stress-induced depression-like behaviors is related to the decreased hippocampal SIRT1 expression,and both swimming and metformin ameliorated depression-like behaviors are associated with increased hippocampal SIRT1 expression of chronic stress mice.(3)Chronic stress damage of hippocampal synaptic plasticity,may be associated with the dysfunction of hippocampal miRNAs;swimming exercise improve hippocampal synaptic plasticity may associated with reversing the dysfunction of hippocampal miRNAs,and metformin improve hippocampal synaptic plasticity may be independent with hippocampal miRNAs.