Influences Of Exercise And Stress On Hippocampal Plasticity And Anxiety-Like Behavior In Adult Mice

Background:Anxiety disorder is a kind of mental illness to serious risk to human health. To explore the developmental mechanism of anxiety disorder, and seek effective therapeutic methods has become an urgent problem to be solved. Animal model can simulate the human emotional disorder well, and animal model of anxiety is the animal in a series of stressful environments, resulting in some obstacles of animal behavior. Among them, the hippocampus is one of the limbic system and closely related to emotional reaction. In particular, adult neurogenesis is a process that new granule cells can be continuously generated in the granule cell layer of the dentate gyrus, and these new neurons migrate, differentiate, and mature into newly integrated, functioning cells. Research shows that stress inhibits adult neurogenesis in the hippocampus, while exercise not only could promote production of newborn neurons in the hippocampus, but also has anxiolytic action. Therefore, adult hippocampal neurogenesis might be the central nerval mechanism of stress-induced anxiety-like behavior and its related cognitive impairment, as well as the key regulator in the anxiolytic effect of exercise.Objectives:To comprehensively and systematically analysze the influences of exercise and stress on hippocampal plasticity and anxiety-like behavior in adult mice, in order to further aware and understand the close connection between "physical exercise and mental health ".Methods:Experiment (1):10weeks-old male C57B1/6J mice were randomly divided into eight groups:the biochemical control (C1) group, the biochemical stress (S1) group, the biochemical exercise (R1) group, the biochemical stress-exercise (S-R1) group, the behaviral control (C2) group, the behavioral stress (S2) group, the behavioral exercise (R2) group, the behavioral stress-exercise (S-R2) group,10mice for each group. C1and C2mice did not give any interventions such as stress or exercise, R1and R2mice housed with running wheel equipment, S1and S2mice received chronic restraint stress, S-R1and S-R2mice housed with running wheel equipment, as well as chronic restraint stress. All mice were injected bromodeoxyuridine (BrdU) for10days to label newborn neurons in the hippocampus. The behavioral groups (C2, S2, R2, S-R2) mice were used to do the open-field test, and the biochemical groups (C1, S1, R1, S-R1) mice were used to make freezing brain slice. Then, the number of new neurons was counted by BrdU immunohistochemistry.Experiment (2):Experiment animals were grouped and injected BrdU as experiment (1). The behavioral groups (C2, S2, R2, S-R2) mice were used to test with the Morris water maze, and the biochemical groups (C1, S1, R1, S-R1) mice were used to make the freezed brain slice. Then, the number of new neurons was counted by BrdU immunohistochemistry, the volume of the dentate gyrus for each group was detemiined by DAPI staining, and the percentage of BrdU+cells colabeling with NeuN to determine a neuronal phenotype.Experiment (3):Experiment animals were grouped as experiment (1). The behavioral groups (C2, S2, R2, S-R2) mice were used to do the elevated-plus maze test after the fear conditioning, and the biochemical groups (C1, S1, R1, S-R1) mice were used to determinate the level of BDNF protein by westernblot.Experiment (4):30male C57B1/6J and NSE-noggin mice for each. Among them,12C57B1/6J mice (C1) and12NSE-noggin mice (N1) were used to test with mouse touchscreen. And18C57B1/6J mice (C1) and18NSE-noggin mice (N1) were used to do the Morris water maze and the object novel recognition tests. C1and N1mice were injected BrdU for10days to label new neurons in the hippocampus, then were used to make freezed brain slice after the behavioral test. Then, the number of newborn neurons was counted by using BrdU immunohistochemistry.Results:Experiment (1):The weight of mice was very significantly reduced (p<0.01) after4weeks voluntary wheel running and3weeks chronic restraint stress together. Compared to the control group, the average running distance of the stress-exercise mice was significantly down-regulated (p<0.05) during the first and second week. Compared to the control mice, the exercise mice and the stress-exercise mice were significantly different such as the time in the center, the ratio of distance travelled in the center to distance travelled in the periphery, the number travelled in the center to distance travelled in the periphery, and the distance travelled in the center during the open-field test, respectively. However, only the exercise mice showed significantly more (p<0.05) BrdU+-labelling cells than the control.Experiment (2):The latency in the place navigation test of the exercise mice and the stress-exercise mice was significantly reduced (p<0.05) than the control. During the spatial probe test (24h) after the place navigation test, the time in targeted quadrant with platform of running mice was significantly enhanced (p<0.05). Compared with the control and the stress group, only5weeks running mice had significantly more (p<0.05) BrdU+cells. Meanwhile, BrdU+cells colabeling with NeuN were upregulated significantly. However, the volume of the dentate gyrus for each group of animals was not different.Experiment (3):In the contextual fear conditioning, the percentage of freezing time of stress mice and the stress-exercise mice was significantly reduced (p<0.05) compared with the control; however, there is no difference among each group about the percentage of freezing time in the cued fear conditioning test. There is significant difference (p<0.05) between the stress-exercise mice and the running mice in the time in closed arm and open arm during the elevated-plus maze test, and the stress-exercise mice had significantly more number of entrance (p<0.05) to open arm than the exercise. Although there is no difference in hippocampal BDNF protein levels among each group, the stress mice showed strong trends toward an increase (p=0.070).Experiment (4):NSE-noggin transgenic mice showed few numbers of reversals (p<0.01) and more trials to criterion (p<0.05) compared to the control. However, there is no difference in BrdU+cells between two groups. In addition, there is very significant difference (p<0.01) in the latency in the place navigation test, the number of swimming through the platform corresponding to the position in the spatial probe test (24h), and the time in targeted quadrant with platform. And there is significant difference (p<0.05) in similar object exploration time in the object recognition test, and there is no difference in preference index.Conclusions:Experiment (1):Voluntary running promotes adult neurogenesis in the hippocampus and alleviates the anxiety-like behavior in mice. Chronic restraint stress reduces the running distance of mice as well as inhibits exercise-induced neurogenesis, and has no effect on the anxiolytic action of exercise, which suggested that exercise-enhanced neurogenesis in the hippocampus might be one of the possible mechanisms of the anxiolytic effect of exercise, there is also other mechanisms regulating the anxiolytic effect of running. Experiment (2):Running increases the number of newborn neurons in the hippocampus, and improves the ability of spatial memory in mice. Stress inhibits neurogenesis as well as impairs spatial memory in mice, which suggested that running-induced adult neurogenesis may be the key regulator of exercise-improved spatial cognitive function.Experiment (3):Stress impairs contextual memory in adult mice, and increase hippocampal BDNF protein. In the elevated-plus maze test after fear conditioning, the stress-exercise mice showed significantly lower level of anxiety-like behavior than the running mice, which suggested that chronic restraint stress protected the anxiolytic effect of exercise from footshock during fear conditioning training.Experiment (4):Compared to the control mice, the ability of small spatial pattern separation in NSE-noggin transgenic mice was reduced, and there is no difference between two groups in the number of new neurons of mice. In addition, the performance of Morris water maze was worsened in NSE-noggin mice, but there is no difference between two groups in the visible platform test, which suggested the capacity of spatial cognition in NSE-noggin transgenic mice was impaired. However, the results of the object recognition test suggested that NSE-noggin transgenic mice has strong tendency to explore or close new object.