【作者】 王艳；

【导师】 李晓白；

【作者基本信息】 中国医科大学 ， 精神病与精神卫生学， 2018， 博士

【摘要】 目的:生命早期应激与个体的认知、行为、情感障碍的发生密切相关。本实验通过建立生命早期应激(Early life stress,ELS)模型,观察生命早期应激及不同年龄阶段(婴幼儿期、青春期、成年期)艾司西酞普兰和丙戊酸钠干预对小鼠行为学及海马区Gsk-3β/β-Catenin/p-Glu R1信号系统的影响,从而探讨生命早期应激对子代小鼠的影响及其病理生理学机制,以期为与生命早期应激相关疾病的发病机制的研究提供理论依据。方法:采用母婴分离模型,将C57BL/6J小鼠分为2组,对照组(control,CON):正常饲养组,和生命早期应激组(ELS):母一代(F0)及小鼠自身(F1)从哺乳期至断奶期(第2-20天)给予母婴分离应激。对照组和ELS组分别在小鼠婴幼儿期、青春期、成年期给予生理盐水、艾司西酞普兰和丙戊酸钠干预,待小鼠成年后进行旷场实验、社交实验、高架十字迷宫、悬尾实验和强迫游泳实验,并检测小鼠海马区Gsk-3β,p-Gsk-3β,β-Catenin,Non-phospho(Active)β-Catenin,Glu R1,p-Glu R1和Glu R2的表达。结果:1.生命早期应激增加了小鼠的类焦虑样行为,早期干预有利于减少小鼠类焦虑样行为。在旷场实验中,各组小鼠在10min内的运动距离没有明显差异,即生命早期应激(ELS)和各个阶段药物干预对小鼠的运动活性没有显著影响。在婴幼儿时期给药阶段,生命早期应激减少小鼠进入中央区的时间,婴幼儿时期给予艾司西酞普兰干预可逆转早期应激的影响。在青春期给药阶段,ELS减少小鼠进入中央区的时间,艾司西酞普兰或丙戊酸钠干预可缓解早期应激的影响。在成年期给药阶段,生命早期应激减少小鼠进入中央区的时间,应激组给予艾司西酞普兰或丙戊酸钠干预可逆转早期应激的影响,然而对照组给予艾司西酞普兰或丙戊酸钠干预减少了小鼠进入中央区的时间。不同阶段疗效比较发现,在青春期给予丙戊酸钠干预更能增加小鼠进入中央区的时间。在社交实验中,在婴幼儿时期给药阶段,丙戊酸钠干预可以增加小鼠在社交箱中的活动距离;ELS减少小鼠进入社交区的时间,艾司西酞普兰或丙戊酸钠干预可缓解早期应激的影响。青春期给药阶段,生命早期应激减少小鼠进入社交区的时间,艾司西酞普兰或丙戊酸钠干预可逆转早期应激的影响,但并不改变对照组小鼠进入社交区的时间。在成年期给药阶段,ELS减少小鼠进入社交区的时间,对照组成年期艾司西酞普兰干预增加小鼠进入社交区的时间,但并不能缓解早期应激的影响。不同阶段疗效比较发现,在青春期给予艾司西酞普兰干预更能增加小鼠进入社交区的时间。在高架十字迷宫实验中,在婴幼儿时期给药阶段,ELS减少小鼠在高架十字迷宫中的活动距离及进入开放臂的相对时间,丙戊酸钠干预增加小鼠在高架十字迷宫中的活动距离。在青春期给药阶段,ELS减少小鼠进入开放臂的相对时间,丙戊酸钠干预可增加小鼠在高架十字迷宫中的活动距离。成年期给药阶段,戊酸钠干预可增加小鼠在高架十字迷宫中的活动距离,并逆转ELS组小鼠进入开放臂的相对时间,但减少对照组小鼠进入开放臂的相对时间。2.生命早期应激增加了小鼠的类抑郁样行为,早期干预有利于减少小鼠类抑郁样行为。在悬尾实验中,在婴幼儿时期给药阶段,ELS增加小鼠的immobility时间,丙戊酸钠干预可减少小鼠的immobility时间。在成年期给药阶段,ELS增加小鼠的immobility时间,但艾司西酞普兰或丙戊酸钠干预无显著作用。强迫游泳实验结果显示,在婴幼儿时期给药阶段,艾司西酞普兰干预或丙戊酸钠干预可减少小鼠的immobility时间。在成年期给药阶段,应激增加了小鼠的immobility时间,丙戊酸钠干预增加小鼠的immobility时间。阶段疗效结果显示,婴儿期丙戊酸钠干预可缓解ELS造成的影响。3.生命早期应激改变了Gsk-3β/β-Catenin/p-Glu R1信号系统功能改变,早期干预有利于逆转Gsk-3β/β-Catenin/p-GluR1信号系统功能。生化结果显示,各处理组小鼠海马区Glu R1和Glu R2的m RNA和蛋白表达量及Gsk-3β、β-Catenin蛋白表达量无明显差异。在婴幼儿期给药阶段,ELS减少了小鼠海马区p-Gsk-3β的表达量,艾司西酞普兰或丙戊酸钠干预可逆转ELS的影响。在青春期给药阶段,艾司西酞普兰或丙戊酸钠干预可增加p-Gsk-3β的表达量。在成年期给药阶段,ELS减少了小鼠海马区p-Gsk-3β的表达量,艾司西酞普兰可逆转ELS的影响,丙戊酸钠干预增加对照组小鼠海马区p-Gsk-3β的表达量,但未能逆转ELS的影响。在婴幼儿期给药阶段,艾司西酞普兰或丙戊酸干预可增加Non-phospho(Active)β-Catenin的表达量。在青春期给药阶段,ELS减少了小鼠海马区Non-phospho(Active)β-Catenin的表达量,艾司西酞普兰或丙戊酸钠干预可逆转ELS的影响。在成年期给药阶段,应激显著减少了小鼠海马区Non-phospho(Active)β-Catenin的表达量,但艾司西酞普兰或丙戊酸钠干预未能逆转ELS的影响。在婴幼儿期给药阶段,ELS减少了小鼠海马区p-Glu R1的表达量,艾司西酞普兰或丙戊酸钠干预可增加p-Glu R1的表达量。在青春期给药阶段,ELS干预减少了p-Glu R1的表达量,但艾司西酞普兰或丙戊酸钠干预未能逆转ELS的影响。在成年期给药阶段,ELS导致p-Glu R1的表达量下降,艾司西酞普兰或丙戊酸钠干预未能逆转ELS的影响。结论:生命早期应激导致Gsk-3β/β-Catenin/p-Glu R1信号系统功能改变,增加了子代对应激反应的脆弱性。更多还原

【Abstract】 Objectives: Early-life stress can lead to long-term consequence on cognitive,behavioral,and mood disorders in later life.Here,we assess the implications of an adverse early environment and the intervention of escitalopram or sodium valproate in different stages(infancy,adolescence and adulthood)on behavioral and hippocampal Gsk-3β/β-Catenin/ p-Glu R1 changes during adulthood in mice.And so as to study the pathophysiological mechanisms of early life stress,and provide a theoretical basis for the study of the pathogenesis of the early life stress-related diseases.Methods: In this study,a mouse model of maternal separation was adopted,the pregnant mice were divided into control group(CON)which was reared in normal condition,and early life stress group(ELS)whose mothers and themselves were experienced maternal separation from postnatal day 2 to day 21.Then,the saline,escitalopram and sodium valproate were administered separately to the mice during infancy,adolescence and adulthood in both the control group and the ELS group.We investigated behavioral and molecular changes resulting from ELS by measuring anxiety-like behavior in the open field test(OFT),social interaction test(SIT)and elevated plus maze(EPM)test,and depression-like behavior in tail suspension test(TST)and forced swimming test(FST),and the changes hippocampal Gsk-3β,p-Gsk-3β,β-Catenin,Non-phospho(Active)β-Catenin,Glu R1,p-Glu R1 and Glu R2 during adulthood in mice.Results: 1.In open field test,the distance moved during 10 minutes had no difference between each group,which demonstrated that neither early life stress,nor drug intervention in each stage had effect on motor activity in mice.In infancy intervention stage,ELS decreased time spent in center zone in mice,escitalopram intervention could increase the time spent in center zone.In adolescence intervention stage,ELS decreased time spent in center zone in mice,escitalopram or sodium valproate intervention could increase the time spent in center zone.In adult intervention stage,ELS decreased timespent in center zone in mice,escitalopram intervention could increased the time spent in center zone in ELS mice,however,escitalopram intervention decreased the time spent in center zone in CON mice.Stage curative effect revealed that sodium valproate intervention during adolescence stage could increase more time spent in center zone.2.In social interaction test,sodium valproate intervention could increase the distance moved during 10 minutes in social interaction box in infancy intervention stage.ELS decreased time spent in social interaction zone in mice,escitalopram or sodium valproate intervention could increase the time spent in social interaction zone in infancy intervention stage.In adolescence intervention stage,ELS decreased time spent in social interaction zone,escitalopram or sodium valproate intervention could increase the time spent in social interaction zone in ELS mice,but not in CON mice.In adult intervention stage,ELS decreased time spent in social interaction zone in mice,escitalopram intervention could increased the time spent in social interaction zone in CON mice,however,it couldn’t increase the time spent in social interaction zone in ELS mice.Stage curative effect revealed that escitalopram intervention during adolescence stage could increase more time spent in social interaction zone.3.In elevated plus maze test,ELS decreased the distance moved in the elevated plus maze and the time spent in open arms,sodium valproate intervention could increase the distance moved in the elevated plus maze in infancy intervention stage.In adolescence intervention stage,ELS decreased the time spent in open arms,and sodium valproate intervention could increase the distance moved in the elevated plus maze.In adult intervention stage,sodium valproate intervention could increase the distance moved in the elevated plus maze and increased the time spent in open arms in ELS mice,but not in CON mice.4.In tail suspension test,ELS increased the immobility time and sodium valproate intervention could decrease the immobility time in infancy intervention stage.In adult intervention stage,ELS increased the immobility time in mice,but neither escitalopram,nor sodium valproate could reverse the effect of ELS.5.In forced swim test,escitalopram and sodium valproate intervention could decrease the immobility time in infancy intervention stage.In adult intervention stage,ELS increased the immobility time in mice,and sodium valproate also increased the immobility time in mice.Stage curative effect revealed that sodium valproate could reverse the effect of ELS only in infancy intervention stage.6.there areno difference in the expression of Gsk-3β,β-Catenin,Glu R1 and Glu R2 between each group.In infancy intervention stage,ELS decreased the expression of p-Gsk-3β and Non-phospho(Active)β-Catenin,and escitalopram and sodium valproate intervention could increase the expression of p-Gsk-3β,Non-phospho(Active)β-Catenin and p-Glu R1.In adolescence intervention stage,ELS decreased the expression of Non-phospho(Active)β-Catenin and p-Glu R1,and escitalopram and sodium valproate intervention could increase the expression of p-Gsk-3β.In adult intervention stage,ELS decreased the expression of p-Gsk-3β,Non-phospho(Active)β-Catenin and p-Glu R1,and escitalopram and sodium valproate intervention couldn’t reverse the decreased the expression of p-Gsk-3β,Non-phospho(Active)β-Catenin and p-Glu R1 induced by ELS.Conclusion: Our study revealed that early life stress resulted in changes in the function of Gsk-3β/β-Catenin/p-Glu R1 signaling system,which increased the neurobiological vulnerability to stress in later life.更多还原