Effect Of Adipokine ZAG On Depression-like Behavior And Hippocampal Oxidative Stress In Fasting Mice

The hippocampus is an important part of the brain of higher mammals.Its functions involve the regulation of learning and memory,emotion and cognition.Hippocampal oxidative stress is closely related to the occurrence of epilepsy,depression and mania.Studies have shown that mitochondrial dysfunction and neuronal damage caused by increased oxidative stress in hippocampus is one of the causes of depression.Reducing neuronal oxidative damage and mitochondrial dysfunction may become a new strategy for the treatment of depression.Zinc α2 glycoprotein(ZAG)is an adipokine distributed in multiple tissues.Studies have shown that leptin and adiponectin in the brain can not only regulate neuronal proliferation,survival and synaptic plasticity,but can regulate cell metabolism and inhibit inflammation.Leptin and adiponectin can be used as diagnostic markers in neurodegenerative diseases.ZAG can promote lipid metabolism and glucose homeostasis,regulating insulin sensitivity and inflammation also has something to do with ZAG.The relationship between ZAG and hippocampal oxidative stress and whether it has a protective effect on neurons is still unknown.Short-term acute fasting and deprivation of cultured cells can change their metabolic state and oxidative stress levels.We have studied the effect of ZAG on anxiety and depression-like behavior in mice by fasting treatment in vivo,and explored the role of ZAG in oxidative stress in hippocampal neurons through an in vitro serum/glucose deprivation model.1 The distribution of ZAG in brain tissue and the effect of fasting for 24h on the energy supply of ZAG knockout miceIn this experiment,the distribution of ZAG in mouse brain tissue and its expression in cortex,hippocampus,and hypothalamus were studied.At the same time,we explored the effect of fasting for 24h on ZAG knockout mice.The experiment used 6-8 week male mice,and were divided into ZAG knockout(KO,n=12)group and wild type(WT,n=12)group according to the genotype of the mouse.First,we detected the distribution and expression of ZAG in the brain of wild-type mice by immunohistochemistry and Western Blotting.We found that ZAG is distributed in cortex,hippocampus,and hypothalamus and the expression level in hypothalamus and hippocampus is significantly higher than that in cortex.After normal feeding for 24h,there was no significant difference in the weight of ZAG knockout mice in the control group.After 24 hours of fasting,the body weight of ZAG knockout mice increased significantly(P<0.05).Serum biochemical results showed that:under normal feeding,the serum cholesterol concentration of ZAG knockout mice was significantly lower than that of the control group(P<0.01),and the concentration of triglyceride was significantly higher than that of the control group(P<0.05);fasting for 24h,ZAG The serum cholesterol,triglyceride and free fatty acid concentrations of knockout mice decreased significantly(P<0.01),and the glucose concentration was significantly higher than that of the control group(P<0.05).Fasting significantly increased serum ZAG protein levels(P<0.05),and extremely significantly increased hippocampal ZAG protein levels(P<0.01).The above results indicate that ZAG is distributed in the cortex,hippocampus,and hypothalamus.Fasting significantly increased serum and hippocampal ZAG protein levels.2 Effects of fasting for 24h on anxiety and depression-like behaviors in ZAG knockout miceIn this experiment,6-8-week male ZAG knockout and wild-type mice were used,which were divided into KO group and WT group according to genotype.From six weeks of age,the mice were successively subjected to the tail-hanging experiment,forced swimming experiment,open field experiment,and elevated cross maze experiment.They were fasted for 24 hours before the experiments and recovered for three days after the end of a behavioral experiment.All behavior experiments were resumed for one week after the end of the experiments,and samples were taken after fasting.The results showed:Fasting for 24 hours,compared with the WT group,the KO group increased the immobility time in the tail suspension test and the forced swimming test,indicating that the KO group had increased depression-like behavior(P<0.05);compared with the WT group,the KO group In the elevated cross maze experiment,the number of entering the open arms decreased(P<0.05),indicating increased anxiety-like behavior.Compared with the WT group,the KO group had increased ROS in the hippocampus(P<0.01),and the protein levels of Nrf2 and HO-1 decreased significantly(P<0.05),indicating that the oxidative stress in the hippocampus increased.The above results indicate that the anxiety and depression-like behavior of mice in the KO group increased after 24 hours of fasting;meanwhile,ROS and oxidative stress increased in the hippocampus.3 Effect of overexpression of ZAG on serum or glucose deprivation of HT-22 cellsIn this experiment,hippocampal nerve cells(HT-22)were treated with serum or glucose deprivation for 8 hours as a model,and the role of ZAG in oxidative stress of hippocampal neurons was studied by overexpression.The results showed that in the serum deprivation test,ZAG overexpression increased cell activity(P<0.05),reduced neuronal death(P<0.05);reduced cellular ROS(P<0.05)and relieved mitochondrial membrane potential decline(P<0.05),Increase the intracellular ATP concentration(P<0.05);significantly increase the cellular mitochondrial encoding genes COX1,ATP6,ATP8(P<0.05),significantly increase the expression levels and antioxidant of COX3,ND1,ND3,ND4,ND5(P<0.01)Regulate gene(Nrf2,CAT,HO-1,NQOl)expression levels(P<0.05);activate β3-AR/PKA/CREB pathway.In the glucose deprivation test,overexpression of ZAG alleviated intracellular ROS accumulation(P<0.01)and mitochondrial membrane potential decline(P<0.05);significantly increased the gene expression level of mitochondrial encoding genes ATP6,ND5,ND6(P<0.05),significant Increase oxidoreductase-1 NQO-1 gene expression level(P<0.05)and antioxidant regulatory proteins Nrf2,KEAP1,NQO1 levels(P<0.05).The above results indicate that ZAG overexpression in hippocampal neurons reduces neuronal oxidative stress and mitochondrial damage.Increasing neuronal activity during serum deprivation and reducing neuronal death are also its functions.