| Objectives The depression model of rats was established by chronic unpredictable mild stress(CUMS) in order to observe the change of the expression level of microtubuleassociated protein p MAP-2 and p-Tau in the hippocampus of rats with depression, and further discuss the relationship and significance between microtubule-associated protein in the hippocampus and depression.Methods Eighty adult healthy male SD rats(SPF grade) with the weight of 180~220 g per rat were randomly divided into a control group and a model group. CUMS was given the model group to establish a model. 11 kinds of stress methods were applied, including wet pad for 24 h, behavioral restrictions for 24 h, squirrel cage tilted 45° for 24 h, 4 ℃ icewater swimming for 5 min, water deprivation for 24 h, fasting deprivation for 24 h, 42℃hot water swimming for 5 min, ears shock for 5 s twice(0.8 m A), tail clamp for 1 min,squirrel cage shake for 15 min, continuous illumination for 24 h. The above-mentioned 11 kinds of stress methods were distributed randomly. One kind of stress stimulation was given the rats in the model group every day and the same kind of stress stimulation was not applied continuously. The above procedures were carried out twice. The rats in the control group were left untreated. After the stress stimulation, the rats were put to death respectively after 1d, 7d, 14 d and 28 d. The execution time in the model group was the same with that in the control group. The sucrose preference test, an open field test and Morris water maze were conducted to detect the ethology. The morphological change of hippocampal neurons was carried out by Nissl staining. Transmission electron spectroscopy was used to observe the cell ultrastructure of the hippocampal neurons,especially the structural change of microtubule. The location distribution of p MAP-2 and p-Tau in the hippocampus was observed using double-labelling immunofluorescence.The Western blotting method is used to detect the expression level of the p MAP-2 and p-Tau of the hippocampus. The SPSS17.0 statistical software was used to analyze the obtained data. The difference has a statistical meaning when P is less than 0.05.Results In the sucrose preference test, there is no obvious change in ordinary water consumption for the model group and the control group. The sucrose consumption and the percentage of sucrose preference in model group are lower than those in control group,which indicates that the difference has a statistical meaning. In open field test, the total walking distance, central activity time, up-right times, and grooming behavior in model group are lower than those in control group(P<0.01). In the Morris water maze, theaverage escape latent period in model group is higher than that in control group(P<0.01).In the Nissl staining, the experimental results indicate that the density of hippocampus cells in the control group is large and pyramidal cell layer is thick. In addition, the pyramidal cells are arranged closely, full, and raised obviously. On the contrary, the density of hippocampus cells in the model group is small; the pyramidal cell layer is thinning; the intercellular space is big; the pyramidal cells are arranged loosely, shrank,and do not rise obviously. The results of Transmission electron spectroscopy indicate that the cell nuclear membrane of hippocampus neurons in rats in the control group was integrated, both chromatin structure and ultrastructure are normal. However, hippocampal neuronal nucleolus in rats in the model group happened to shrink, the heterochromatin in the nucleolus increased, small and irregular clot appeared, the edge gathered, the rough endoplasmic reticulum swelled, mitochondria crest disappeared, swelled, and even existed in the cavitation, organelles reduced, the structure was damaged, and different sizes of vacuoles were formed in the cytoplasm. According to the results of the doublelabelling immunofluorescence, the green fluorescence belongs to p MAP-2 motivated by Alexa Fluor 488 and the red fluorescence belongs to p-Tau motivated by Alexa Fluor 594.p MAP-2 and p-Tau proteins express simultaneously in the intracytoplasm of visible part neuron, indicating that there is some co-localization of p MAP-2 and p-Tau proteins. In the results of Western blotting, The expression level of p MAP-2 protein in the hippocampus of rats after CUMS in model group is lower than that in control group at 1d(P<0.01), starts to rise at 7d(P<0.01), obviously higher than that in control group at 14d(P<0.01), and still a little higher than that in control group at 28d(P<0.05).Conclusions 1 The depression model of animals was successfully established. 2 The hippocampal neuron in the depression model changed. 3 The expression level of p MAP-2protein in the depression model of hippocampus of rats decreases firstly and then increases, which might affect the stability of microtubule and destroy the steady state of cytoskeleton, thus affecting the structural change of hippocampal neuron. |
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