运动抗抑郁作用及其分子机制研究
摘要
研究目的
本研究采用慢性不可预知性应激(chronic unpredictable stress,CUS)制备大鼠抑郁模型,并伴随进行规律性运动和氟西汀(fluoxetine,Flu)干预,从整体行为学表现、海马神经元损伤、神经营养通路重要分子表达水平多个方面,研究运动及其与氟西汀联合使用的抗抑郁效果,并探讨其可能的机制,为运动提高身心健康,丰富抑郁症的治疗手段提供理论依据。
研究方法
65只雄性SD大鼠,安静饲养适应环境3天后进行1周适应性跑台训练,挑选出56只大鼠(淘汰不愿意进行跑步的动物),根据体重,均衡分为Control组、CUS组、Flu组、小强度跑台运动组(low intensity treadmill running,LIR)、中等强度跑台运动组(moderate intensity treadmill running,MIR)、Flu+LIR组和Flu+MIR组。
除Control组外,其余各组大鼠均接受为期4周的慢性不可预知性应激刺激。Flu组、Flu+LIR组、Flu+MIR组每次应激前1h灌胃给予氟西汀(10mg/kg/day,给药体积2ml/kg,1次/日),其余组大鼠灌胃给予相应体积的生理盐水。LIR组和Flu+LIR组、MIR组和Flu+MIR组大鼠每天上午分别进行小强度、中等强度跑台训练。
采用巧克力牛奶快感缺乏实验、开场实验、新奇抑制摄食实验,并测定每周体重,观察运动、氟西汀对CUS大鼠行为学表现和体重增长的影响;应用免疫组织化学法和western-blotting法,检测运动和氟西汀对CUS大鼠海马神经元形态、结构、数目,海马神经细胞微管相关蛋白-2C(MAP-2C)及神经营养通路重要信号分子细胞外信号调节激酶(ERK)、磷酸化ERK(pERK)、脑源性神经营养因子(BDNF)、VGF(一种神经肽,非缩写形式)蛋白表达的影响。
研究结果
(1)行为学研究
CUS明显使大鼠巧克力牛奶偏嗜度下降、开场活动减少、新奇环境中的摄食潜伏期延长;氟西汀可以明显对抗上述改变。慢性应激伴随进行运动具有与氟西汀一致的抗抑郁样作用。
(2)体重变化
CUS显著抑制大鼠体重的增长,Flu,Flu+MIR、Flu+LIR均未能改善CUS大鼠体重增长的缓慢。MIR、LIR能够改善CUS大鼠体重增长缓慢的情况,每周体重与CUS组相比没有显著性差异。
(3)神经生物学研究
①免疫组织化学研究
CUS破坏了大鼠海马神经元正常的形态结构,减少大鼠海马CA1区锥体细胞数量。Flu、MIR、Flu+MIR和Flu+LIR均能够对抗海马神经元的应激性损伤和CA1区锥体细胞数量的减少。Flu对抗CUS大鼠海马CA1区锥体细胞数量减少的作用分别显著优于MIR和LIR。
②western-blotting研究
CUS显著下调海马MAP-2C、pERK、BDNF和VGF的表达,Flu、MIR、Flu+MIR和Flu+LIR,均能够显著改善这种情况。Flu+LIR显著上调CUS大鼠海马pERK和BDNF的表达,未能明显上调MAP-2C和VGF的表达。
对CUS大鼠海马pERK表达的上调,MIR、Flu+LIR显著优于LIR。对CUS大鼠BDNF表达的上调,Flu+MIR显著优于Flu或者MIR的单独干预;Flu+LIR显著优于LIR的单独干预。
研究结论
(1)成功制备大鼠慢性不可预见应激抑郁模型,其较好地模拟了人类抑郁症的行为特点:CUS大鼠出现快感缺乏、自发活动下降、摄食潜伏期延长的抑郁样行为表现和体重增长缓慢的躯体症状。
(2)运动对CUS大鼠具有与抗抑郁药氟西汀一致的抗抑郁行为学效应。
(3)运动抗抑郁作用可能与以下因素有关:①保护抑郁模型大鼠海马神经元形态结构;②阻止锥体细胞数量的减少;③上调海马MAP-2C表达;④上调海马BDNF及其相关的神经营养通路重要信号分子pERK和VGF的表达。这有利于保护应激状态下海马结构和功能的进一步被破坏,促进神经营养和神经可塑性。
(4)运动能够增强氟西汀的神经营养作用。
Purpose
To investigate effects of regular exercise,the selective serotonin reputake inhibitor, fluoxetine(Flu),and the combination of them on behavior,neuronal impairment,and the neurotrophic factor signal pathway in the chronic unpredictable stress(CUS)animal model of depression,and to elucidate possible molecular mechanisms of antidepressive-like effect of exercise and it's combination with fluoxetine.
Methods
65 Male SD rats were habitually housed for three days and then adaptively exercised on treadmill for one week.According to running performance and body weight 56 rats were balancedly grouped into seven groups:Control,CUS,Flu,low intensity treadmill running(LIR),moderate intensity treadmill running(MIR),Flu+LIR and Flu+MIR.
CUS was exercted on all groups exception of the Control group for four weeks.The Flu,Flu+LIR,and Flu+MIR group,fluoxetine(10mg/kg/day,2ml/kg)was applied to rats before one hour of daily stress.Other groups were administrationed same volume of saline.The LIR,MIR,Flu+LIR,and Flu+MIR group were exercised different intensity of treadmill running.After four weeks of CUS,chocolate milk anhedonia test,open-field test,novelty-suppressed feeding and together with weekly body weight were determined. Morphological,cytoarchitectural and quantitative changes of hippocampal CA1 pyramidal neurons were detected using immunohistochemistry,MAP-2C expression was determined using western-blotting.The expression of hippocampal brain-derived neurotrophic factor(BDNF),extracellular signal regulated kinase(ERK),phosphor-ERK (pERK),VGF(a neuropeptide,nonacronymic)were determined using western-blotting.
Results
(1)Behavioral studies
CUS significantly decreased the chocolate milk preference and spontaneous activity, prolonged latency to feed,and concomitant treatment with Flu could reverse these changes.CUS concomitant with exercise hasthe same antidepressive-like effect as Flu.
(2)Body weight
CUS could significantly suppress the increase of body weight.Flu,Flu+MIR and Flu+LIR did not improve CUS-induced the suppression of body weight.LIR and MIR reversed the low increase of body weight and they did not have significant difference compared with that of CUS.
(3)Neurobiological studies
①immmunohistochemistry staining
CUS destructed the normally hippocampal morphosis,reduced the number of hippocampal CA1 pyramidal neurons.Flu,MIR,Flu+LIR and Flu+MIR could antagonize the CUS-induced hippocampal impairement and the reduction of the number of hippocampal CA1 pyramidal neurons.The antagonistic effect of Flu on the reduced number of hippcampal CA1 pyramidal neurons was better than that of LIR and MIR,respectively.
②western-blotting
CUS down-regulated significantly the expression of MAP-2C,pERK,BDNF and VGF in hippocampus.Flu,MIR,Flu+LIR and Flu+MIR could improve these changes. Flu+LIR up-reguated the expression of hippocampal pERK and BDNF but the expression of hippocampal MAP-2C and VGF.
The up-regulaed effect of MIR and Flu+LIR on the expression of hippocampal pERK was better than that of LIR.The up-regulated effect of Flu+MIR on the expression of hippocampal BDNF was better than that of Flu or MIR,respectively;Flu+LIR was better than LIR.
Conclusion
(1)We successfully duplicated the CUS animal model of depression,which simulated the depressive-like behaviors as following:significantly decreased in chocolate milk preference in and spontaneous activity,prolonged latency to feed,and slowed growth of body weight.
(2)Exercise could significantly improve the reduction of chocolate milk preference and spontaneous activity,and the augmentation of latency to feed in CUS model.These behavioral changes were consistent with the effect of fluoxetine in CUS model.
(3)The antidepressant mechanisms of exercise may be related to following: exercise can antagonize the chronic stress-induced hipocampal impairment via:①protecting the morphology and cyto-architecture of hippocampal CA1 pyramidal neurons;②preventing reduction of the number of hippocampal CA1 pyramidal neurons;③up-regalting the expression of hippocampal MAP-2C in CUS model of depression;④up-regalting the expression of hippocampal BDNF,pERK and VGF which were related to BDNF's signal pathway may be underlying the neurotrophic and neuroplastic effects of exercise.
(4)The neuroprotective and neurotrophic effect of fluoxetine can be enhanced further by exercise.
本研究采用慢性不可预知性应激(chronic unpredictable stress,CUS)制备大鼠抑郁模型,并伴随进行规律性运动和氟西汀(fluoxetine,Flu)干预,从整体行为学表现、海马神经元损伤、神经营养通路重要分子表达水平多个方面,研究运动及其与氟西汀联合使用的抗抑郁效果,并探讨其可能的机制,为运动提高身心健康,丰富抑郁症的治疗手段提供理论依据。
研究方法
65只雄性SD大鼠,安静饲养适应环境3天后进行1周适应性跑台训练,挑选出56只大鼠(淘汰不愿意进行跑步的动物),根据体重,均衡分为Control组、CUS组、Flu组、小强度跑台运动组(low intensity treadmill running,LIR)、中等强度跑台运动组(moderate intensity treadmill running,MIR)、Flu+LIR组和Flu+MIR组。
除Control组外,其余各组大鼠均接受为期4周的慢性不可预知性应激刺激。Flu组、Flu+LIR组、Flu+MIR组每次应激前1h灌胃给予氟西汀(10mg/kg/day,给药体积2ml/kg,1次/日),其余组大鼠灌胃给予相应体积的生理盐水。LIR组和Flu+LIR组、MIR组和Flu+MIR组大鼠每天上午分别进行小强度、中等强度跑台训练。
采用巧克力牛奶快感缺乏实验、开场实验、新奇抑制摄食实验,并测定每周体重,观察运动、氟西汀对CUS大鼠行为学表现和体重增长的影响;应用免疫组织化学法和western-blotting法,检测运动和氟西汀对CUS大鼠海马神经元形态、结构、数目,海马神经细胞微管相关蛋白-2C(MAP-2C)及神经营养通路重要信号分子细胞外信号调节激酶(ERK)、磷酸化ERK(pERK)、脑源性神经营养因子(BDNF)、VGF(一种神经肽,非缩写形式)蛋白表达的影响。
研究结果
(1)行为学研究
CUS明显使大鼠巧克力牛奶偏嗜度下降、开场活动减少、新奇环境中的摄食潜伏期延长;氟西汀可以明显对抗上述改变。慢性应激伴随进行运动具有与氟西汀一致的抗抑郁样作用。
(2)体重变化
CUS显著抑制大鼠体重的增长,Flu,Flu+MIR、Flu+LIR均未能改善CUS大鼠体重增长的缓慢。MIR、LIR能够改善CUS大鼠体重增长缓慢的情况,每周体重与CUS组相比没有显著性差异。
(3)神经生物学研究
①免疫组织化学研究
CUS破坏了大鼠海马神经元正常的形态结构,减少大鼠海马CA1区锥体细胞数量。Flu、MIR、Flu+MIR和Flu+LIR均能够对抗海马神经元的应激性损伤和CA1区锥体细胞数量的减少。Flu对抗CUS大鼠海马CA1区锥体细胞数量减少的作用分别显著优于MIR和LIR。
②western-blotting研究
CUS显著下调海马MAP-2C、pERK、BDNF和VGF的表达,Flu、MIR、Flu+MIR和Flu+LIR,均能够显著改善这种情况。Flu+LIR显著上调CUS大鼠海马pERK和BDNF的表达,未能明显上调MAP-2C和VGF的表达。
对CUS大鼠海马pERK表达的上调,MIR、Flu+LIR显著优于LIR。对CUS大鼠BDNF表达的上调,Flu+MIR显著优于Flu或者MIR的单独干预;Flu+LIR显著优于LIR的单独干预。
研究结论
(1)成功制备大鼠慢性不可预见应激抑郁模型,其较好地模拟了人类抑郁症的行为特点:CUS大鼠出现快感缺乏、自发活动下降、摄食潜伏期延长的抑郁样行为表现和体重增长缓慢的躯体症状。
(2)运动对CUS大鼠具有与抗抑郁药氟西汀一致的抗抑郁行为学效应。
(3)运动抗抑郁作用可能与以下因素有关:①保护抑郁模型大鼠海马神经元形态结构;②阻止锥体细胞数量的减少;③上调海马MAP-2C表达;④上调海马BDNF及其相关的神经营养通路重要信号分子pERK和VGF的表达。这有利于保护应激状态下海马结构和功能的进一步被破坏,促进神经营养和神经可塑性。
(4)运动能够增强氟西汀的神经营养作用。
Purpose
To investigate effects of regular exercise,the selective serotonin reputake inhibitor, fluoxetine(Flu),and the combination of them on behavior,neuronal impairment,and the neurotrophic factor signal pathway in the chronic unpredictable stress(CUS)animal model of depression,and to elucidate possible molecular mechanisms of antidepressive-like effect of exercise and it's combination with fluoxetine.
Methods
65 Male SD rats were habitually housed for three days and then adaptively exercised on treadmill for one week.According to running performance and body weight 56 rats were balancedly grouped into seven groups:Control,CUS,Flu,low intensity treadmill running(LIR),moderate intensity treadmill running(MIR),Flu+LIR and Flu+MIR.
CUS was exercted on all groups exception of the Control group for four weeks.The Flu,Flu+LIR,and Flu+MIR group,fluoxetine(10mg/kg/day,2ml/kg)was applied to rats before one hour of daily stress.Other groups were administrationed same volume of saline.The LIR,MIR,Flu+LIR,and Flu+MIR group were exercised different intensity of treadmill running.After four weeks of CUS,chocolate milk anhedonia test,open-field test,novelty-suppressed feeding and together with weekly body weight were determined. Morphological,cytoarchitectural and quantitative changes of hippocampal CA1 pyramidal neurons were detected using immunohistochemistry,MAP-2C expression was determined using western-blotting.The expression of hippocampal brain-derived neurotrophic factor(BDNF),extracellular signal regulated kinase(ERK),phosphor-ERK (pERK),VGF(a neuropeptide,nonacronymic)were determined using western-blotting.
Results
(1)Behavioral studies
CUS significantly decreased the chocolate milk preference and spontaneous activity, prolonged latency to feed,and concomitant treatment with Flu could reverse these changes.CUS concomitant with exercise hasthe same antidepressive-like effect as Flu.
(2)Body weight
CUS could significantly suppress the increase of body weight.Flu,Flu+MIR and Flu+LIR did not improve CUS-induced the suppression of body weight.LIR and MIR reversed the low increase of body weight and they did not have significant difference compared with that of CUS.
(3)Neurobiological studies
①immmunohistochemistry staining
CUS destructed the normally hippocampal morphosis,reduced the number of hippocampal CA1 pyramidal neurons.Flu,MIR,Flu+LIR and Flu+MIR could antagonize the CUS-induced hippocampal impairement and the reduction of the number of hippocampal CA1 pyramidal neurons.The antagonistic effect of Flu on the reduced number of hippcampal CA1 pyramidal neurons was better than that of LIR and MIR,respectively.
②western-blotting
CUS down-regulated significantly the expression of MAP-2C,pERK,BDNF and VGF in hippocampus.Flu,MIR,Flu+LIR and Flu+MIR could improve these changes. Flu+LIR up-reguated the expression of hippocampal pERK and BDNF but the expression of hippocampal MAP-2C and VGF.
The up-regulaed effect of MIR and Flu+LIR on the expression of hippocampal pERK was better than that of LIR.The up-regulated effect of Flu+MIR on the expression of hippocampal BDNF was better than that of Flu or MIR,respectively;Flu+LIR was better than LIR.
Conclusion
(1)We successfully duplicated the CUS animal model of depression,which simulated the depressive-like behaviors as following:significantly decreased in chocolate milk preference in and spontaneous activity,prolonged latency to feed,and slowed growth of body weight.
(2)Exercise could significantly improve the reduction of chocolate milk preference and spontaneous activity,and the augmentation of latency to feed in CUS model.These behavioral changes were consistent with the effect of fluoxetine in CUS model.
(3)The antidepressant mechanisms of exercise may be related to following: exercise can antagonize the chronic stress-induced hipocampal impairment via:①protecting the morphology and cyto-architecture of hippocampal CA1 pyramidal neurons;②preventing reduction of the number of hippocampal CA1 pyramidal neurons;③up-regalting the expression of hippocampal MAP-2C in CUS model of depression;④up-regalting the expression of hippocampal BDNF,pERK and VGF which were related to BDNF's signal pathway may be underlying the neurotrophic and neuroplastic effects of exercise.
(4)The neuroprotective and neurotrophic effect of fluoxetine can be enhanced further by exercise.
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