BDNF和VEGF在慢性应激抑郁模型小鼠神经元损伤中的作用及其机制
发布时间：2018-07-29 15:23
【摘要】：研究目的:建立小鼠慢性不可预见性应激抑郁模型,研究抑郁模型小鼠脑内海马区(DG区、CA1区、CA3区)与前额叶皮层VEGF和BDNF的表达及其与抑郁症发生的关系,探究其在抑郁症发生中的作用及其相关分子机制。研究方法:随机将2月龄昆明系小白鼠分为对照组和应激组。对照组小鼠正常饲养,应激组小鼠采用7种不同的应激因子(禁水、禁食、高台、斜笼、足底电击、冰水游泳、通宵照明)21天,建立慢性不可预见性应激抑郁模型。实验一:采用旷场实验、悬尾实验、Morris水迷宫实验方法,检测小鼠行为和空间学习记忆能力各项指标的变化,通过免疫组化实验检测各组小鼠脑内海马(DG区、CA1区、CA3区)和前额叶皮层BDNF、VEGF及PI3K表达量的变化,以及通过HE染色,观察海马区和前额叶皮层神经元的形态变化。实验二:采用脑立体定位技术,小鼠双侧海马分别注射生理盐水和LY294002。C-ns组:对照组海马内注射生理盐水(NS),C-ly组:对照组海马内注射LY294002;S-ns组:应激组海马内注射生理盐水,S-ly组:应激组海马内注射LY294002。24小时后检测小鼠行为和空间学习记忆能力各项指标的变化,通过免疫组化实验检测各组小鼠脑内海马和前额叶皮层BDNF、VEGF及PI3K表达的变化,以及通过HE染色,观察海马区和前额叶皮层神经元的形态变化。研究结果:实验一:1.旷场实验:与对照组相比,应激组小鼠的爬行格数、修饰次数及直立次数均显著减少,中央格停留时间显著增加。2.悬尾实验:与对照组相比,应激组小鼠的第一次静止不动时间(潜伏期)显著减少,后4分钟累计不动时间显著增加。3.Morris水迷宫实验:对照组和应激组小鼠随着训练次数的增加其逃避潜伏期和游泳总路程均缩短,对照组逃避潜伏期和游泳总路程均显著低于应激组;空间搜索实验中应激组小鼠在目标象限的停留时间显著少于对照组。4.免疫组化实验,与对照组相比,应激组小鼠DG区、CA1区、CA3区及前额叶皮层的BDNF和VEGF表达均显著降低,PI3K的表达在DG区和前额叶皮层显著降低。表明,慢性应激可引起小鼠海马和前额叶皮层BDNF、VEGF和PI3K的表达降低。实验二:1.旷场实验:相比于C-ns组,C-ly组小鼠的爬行格数、修饰次数及直立次数均显著减少,S-ns组小鼠的爬行格数、修饰次数显著减少;与S-ns组相比,S-ly组小鼠中央格停留时间、爬行格数、修饰次数均显著减少。2.悬尾实验:与C-ns组相比,C-ly组小鼠的第一次静止不动时间显著减少,后4分钟累计不动时间显著增加,S-ns组小鼠后4分钟累计不动时间显著增加;与S-ns组相比,S-ly组小鼠后4分钟累计不动时间显著增加。3.Morris水迷宫实验:随着训练次数的增加,C-ns组和S-ns组小鼠定位航行实验的逃避潜伏期和游泳总路程变化显著,C-ly组和S-ly组没有显著变化。组间对比发现,与C-ns组相比,C-ly组、S-ns组小鼠的逃避潜伏期和总路程均显著增高;S-ly组小鼠的逃避潜伏期和游泳总路程均显著高于S-ns组。空间搜索实验中,与C-ns组相比,C-ly组和S-ns组小鼠在目标象限的停留时间显著降低;S-ly组小鼠在目标象限的停留时间显著低于S-ns组。4.免疫组化实验:与C-ns组相比,C-ly组小鼠海马的DG区、CA1区和CA3区内的BDNF、VEGF表达均显著下降,DG区和前额叶皮层内的PI3K表达显著下降;与S-ns组相比,S-ly组小鼠海马DG区、CA1区、CA3区和前额叶皮层内的BDNF、PI3K表达均显著下降,而VEGF在海马的DG区、CA1区和CA3区表达显著下降。表明,慢性应激后双侧海马注射PI3K-Akt信号通路阻断剂LY294002后可导致BDNF、VEGF和PI3K的表达显著降低。研究结论:1.慢性应激可引起小鼠明显的焦虑、抑郁样行为,空间学习记忆能力减退;海马内注射LY294002后小鼠的行为和学习记忆功能损伤更严重。2.慢性应激后小鼠海马和前额叶皮层BDNF、VEGF和PI3K的表达降低;海马内注射LY294002后小鼠BDNF、VEGF和PI3K的表达降低更显著。3.海马和前脑皮层BDNF和VEGF表达的下调与慢性应激所致小鼠的抑郁密切相关,且可能通过PI3K/Akt信号通路参与抑郁症的发生。
[Abstract]:Objective: to establish a model of chronic unpredictable stress depression in mice, to study the expression of VEGF and BDNF in the hippocampus (DG region, CA1 region, CA3 area) and prefrontal cortex in the brain of the depressive model mice and their relationship with the occurrence of depression, and to explore its role in the occurrence of depression and its related molecular mechanism. The research method: the 2 month old Kunming system was randomly selected. The mice in the control group were divided into the control group and the stress group. The mice in the control group were reared normally. The mice in the stress group adopted 7 different stress factors (water prohibition, fasting, high platform, oblique cage, foot shock, ice water swimming, all night lighting) to establish a chronic unpredictable stress and depression model. A test of open field experiment, tail suspension experiment, and Morris water maze experiment was used. The changes in the behavior of mice and the ability of spatial learning and memory were detected. The changes in the expression of BDNF, VEGF and PI3K in the hippocampus (DG area, CA1 area, CA3 area) and prefrontal cortex were detected by immunohistochemistry, and the morphological changes of neurons in the hippocampus and prefrontal cortex were observed by HE staining. Experiment two: the use of brain Stereotactic technique, the mice bilateral hippocampus was injected with saline and LY294002.C-ns group respectively: the control group was injected with saline (NS) in the hippocampus, group C-ly: the control group was injected with LY294002 in the hippocampus, S-ns group: the stress group was injected with saline in the hippocampus, group S-ly: the behavior and spatial learning and memory of the mice were detected after the hippocampus injection LY294002.24 hours in the stress group. Changes in various indexes, the changes in the expression of BDNF, VEGF and PI3K in the hippocampus and prefrontal cortex of the mice were detected by immunohistochemistry, and the morphological changes in the hippocampal and prefrontal cortex neurons were observed by HE staining. The results were as follows: Experiment 1: 1. open field experiment: the creeping number of mice in the stress group, compared with the control group, was compared with the control group. The number of modification and erect times were significantly reduced, and the residence time of the central lattice was significantly increased by the.2. suspension experiment. Compared with the control group, the first static time (incubation period) of the mice in the stress group decreased significantly, and the cumulative time of the latter 4 minutes increased significantly in the.3.Morris water maze test: the control group and the stress group increased with the increasing number of training times. The escape latency and the total swimming course were shortened, the escape latency and the total swimming course of the control group were significantly lower than those in the stress group; the time of the stress group in the target quadrant of the stress group was significantly less than the.4. immunization test in the control group. Compared with the control group, the mice in the stress group, the CA1 area, the CA3 region and the BDN of the prefrontal cortex were BDN. The expression of F and VEGF decreased significantly, and the expression of PI3K decreased significantly in the DG and prefrontal cortex. It showed that chronic stress could cause the expression of BDNF, VEGF and PI3K in the hippocampus and prefrontal cortex of mice. Experiment two: 1. open field experiment: the number of creeping, the number of modification and the erect times of the C-ly group were significantly decreased compared to the C-ns group, and the S-ns mice were significantly reduced. The number of crawling lattices and the number of modification decreased significantly. Compared with the S-ns group, the residence time of the central lattice, the number of creeping and the number of modifications in the S-ly group reduced the.2. suspension test significantly: compared with the C-ns group, the first time of static immobility in the C-ly group was significantly reduced, the cumulative time for the latter 4 minutes increased significantly, and the cumulative effect of the S-ns group was not moved after 4 minutes. Compared with the S-ns group, the cumulative time of 4 minutes after the S-ly group was significantly increased by the.3.Morris water maze experiment. As the number of training times increased, the escape latency and the total swimming distance of the C-ns and S-ns mice were significantly changed, and there was no significant change in the C-ly and S-ly groups. The escape latency and total route of mice in group C-ly and group S-ns were significantly higher than that in group S-ly. The escape latency and total swimming course of mice in group S-ly were significantly higher than that in group S-ns. Compared with the C-ns group, the stay time of the C-ly and S-ns mice in the target quadrant was significantly lower than that in the C-ns group; the time for the S-ly mice to stay in the target quadrant was significant. Compared with group.4., the expression of VEGF in DG area, CA1 area and CA3 area in group C-ly was significantly decreased, and PI3K expression in the DG area and prefrontal cortex decreased significantly compared with the group C-ns, and the expression in the hippocampus of the hippocampus, the region of the hippocampus, the frontal cortex and the cortex of the prefrontal cortex decreased significantly compared with those in the S-ns group. The expression of VEGF in the hippocampal DG region, CA1 area and CA3 region decreased significantly. The results showed that the expression of BDNF, VEGF and PI3K decreased significantly after the injection of PI3K-Akt signaling blocking agent LY294002 after chronic stress. The conclusion: 1. chronic stress can cause obvious anxiety, depressive behavior, spatial learning and memory impairment in mice; hippocampus After intramuscular injection of LY294002, the behavior and learning and memory function of mice were more severe. The expression of BDNF, VEGF and PI3K in the hippocampus and prefrontal cortex of mice decreased after.2. chronic stress, and the expression of BDNF, VEGF and PI3K decreased more significantly in the hippocampus and the lower.3. hippocampus and the downregulation of BDNF and VEGF expressions in the hippocampus and the prefrontal cortex and the decrease of chronic stress and chronic stress. Depression is closely related to mice, and may participate in the occurrence of depression through the PI3K/Akt signaling pathway.
【学位授予单位】：曲阜师范大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R749.4;R-332
本文编号：2153110
[Abstract]:Objective: to establish a model of chronic unpredictable stress depression in mice, to study the expression of VEGF and BDNF in the hippocampus (DG region, CA1 region, CA3 area) and prefrontal cortex in the brain of the depressive model mice and their relationship with the occurrence of depression, and to explore its role in the occurrence of depression and its related molecular mechanism. The research method: the 2 month old Kunming system was randomly selected. The mice in the control group were divided into the control group and the stress group. The mice in the control group were reared normally. The mice in the stress group adopted 7 different stress factors (water prohibition, fasting, high platform, oblique cage, foot shock, ice water swimming, all night lighting) to establish a chronic unpredictable stress and depression model. A test of open field experiment, tail suspension experiment, and Morris water maze experiment was used. The changes in the behavior of mice and the ability of spatial learning and memory were detected. The changes in the expression of BDNF, VEGF and PI3K in the hippocampus (DG area, CA1 area, CA3 area) and prefrontal cortex were detected by immunohistochemistry, and the morphological changes of neurons in the hippocampus and prefrontal cortex were observed by HE staining. Experiment two: the use of brain Stereotactic technique, the mice bilateral hippocampus was injected with saline and LY294002.C-ns group respectively: the control group was injected with saline (NS) in the hippocampus, group C-ly: the control group was injected with LY294002 in the hippocampus, S-ns group: the stress group was injected with saline in the hippocampus, group S-ly: the behavior and spatial learning and memory of the mice were detected after the hippocampus injection LY294002.24 hours in the stress group. Changes in various indexes, the changes in the expression of BDNF, VEGF and PI3K in the hippocampus and prefrontal cortex of the mice were detected by immunohistochemistry, and the morphological changes in the hippocampal and prefrontal cortex neurons were observed by HE staining. The results were as follows: Experiment 1: 1. open field experiment: the creeping number of mice in the stress group, compared with the control group, was compared with the control group. The number of modification and erect times were significantly reduced, and the residence time of the central lattice was significantly increased by the.2. suspension experiment. Compared with the control group, the first static time (incubation period) of the mice in the stress group decreased significantly, and the cumulative time of the latter 4 minutes increased significantly in the.3.Morris water maze test: the control group and the stress group increased with the increasing number of training times. The escape latency and the total swimming course were shortened, the escape latency and the total swimming course of the control group were significantly lower than those in the stress group; the time of the stress group in the target quadrant of the stress group was significantly less than the.4. immunization test in the control group. Compared with the control group, the mice in the stress group, the CA1 area, the CA3 region and the BDN of the prefrontal cortex were BDN. The expression of F and VEGF decreased significantly, and the expression of PI3K decreased significantly in the DG and prefrontal cortex. It showed that chronic stress could cause the expression of BDNF, VEGF and PI3K in the hippocampus and prefrontal cortex of mice. Experiment two: 1. open field experiment: the number of creeping, the number of modification and the erect times of the C-ly group were significantly decreased compared to the C-ns group, and the S-ns mice were significantly reduced. The number of crawling lattices and the number of modification decreased significantly. Compared with the S-ns group, the residence time of the central lattice, the number of creeping and the number of modifications in the S-ly group reduced the.2. suspension test significantly: compared with the C-ns group, the first time of static immobility in the C-ly group was significantly reduced, the cumulative time for the latter 4 minutes increased significantly, and the cumulative effect of the S-ns group was not moved after 4 minutes. Compared with the S-ns group, the cumulative time of 4 minutes after the S-ly group was significantly increased by the.3.Morris water maze experiment. As the number of training times increased, the escape latency and the total swimming distance of the C-ns and S-ns mice were significantly changed, and there was no significant change in the C-ly and S-ly groups. The escape latency and total route of mice in group C-ly and group S-ns were significantly higher than that in group S-ly. The escape latency and total swimming course of mice in group S-ly were significantly higher than that in group S-ns. Compared with the C-ns group, the stay time of the C-ly and S-ns mice in the target quadrant was significantly lower than that in the C-ns group; the time for the S-ly mice to stay in the target quadrant was significant. Compared with group.4., the expression of VEGF in DG area, CA1 area and CA3 area in group C-ly was significantly decreased, and PI3K expression in the DG area and prefrontal cortex decreased significantly compared with the group C-ns, and the expression in the hippocampus of the hippocampus, the region of the hippocampus, the frontal cortex and the cortex of the prefrontal cortex decreased significantly compared with those in the S-ns group. The expression of VEGF in the hippocampal DG region, CA1 area and CA3 region decreased significantly. The results showed that the expression of BDNF, VEGF and PI3K decreased significantly after the injection of PI3K-Akt signaling blocking agent LY294002 after chronic stress. The conclusion: 1. chronic stress can cause obvious anxiety, depressive behavior, spatial learning and memory impairment in mice; hippocampus After intramuscular injection of LY294002, the behavior and learning and memory function of mice were more severe. The expression of BDNF, VEGF and PI3K in the hippocampus and prefrontal cortex of mice decreased after.2. chronic stress, and the expression of BDNF, VEGF and PI3K decreased more significantly in the hippocampus and the lower.3. hippocampus and the downregulation of BDNF and VEGF expressions in the hippocampus and the prefrontal cortex and the decrease of chronic stress and chronic stress. Depression is closely related to mice, and may participate in the occurrence of depression through the PI3K/Akt signaling pathway.
【学位授予单位】：曲阜师范大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R749.4;R-332
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