睡眠剥夺对慢性应激抑郁模型大鼠单胺递质及腺苷的影响
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摘要
背景:腺苷作为一种神经调质,可以通过与受体结合调节各种神经递质的释放。此外,越来越多的人认为腺苷是一种睡眠因子,主要通过与其受体结合在睡眠调节中起作用。有研究认为腺苷及其受体与抑郁及抗抑郁过程有一定关系。睡眠剥夺可以发挥快速抗抑郁效应,但其中的神经生物学机制至今未明,目前的研究多集中于单胺类神经递质系统,我们认为睡眠剥夺快速抗抑郁的过程可能是多物质相互作用、共同参与的过程。即然如此,腺苷是否参与了睡眠剥夺的快速抗抑郁过程呢?在此过程中,腺苷在不同脑区的含量是否有所变化呢?它与单胺递质的变化情况有何关系?目前尚无这方面的报道。本研究拟通过对慢性应激抑郁模型大鼠进行连续72小时快眼动睡眠剥夺(rapid eye movement sleep deprivation,REMSD)观察大鼠不同脑区单胺类神经递质以及腺苷、肌苷含量的改变,旨在探讨腺苷在睡眠剥夺快速抗抑郁起效机制中可能的作用。
目的:观察睡眠剥夺对慢性应激抑郁模型大鼠不同脑区组织匀浆液中单胺递质及腺苷、肌苷含量的影响。
材料和方法:实验动物为Sprague Dawley 2-3月龄雄性大鼠,随机分为正常对照组,抑郁模型组,抑郁模型+72小时快眼动睡眠剥夺组(以下称REMSD组),抑郁模型组+72小时大平台对照组(以下称大平台对照组),每组8只。应激造模采用慢性轻度不可预见性应激(chronic mild unpredicted stress, CMUS)和分养两种经典模型结合的方式来建立抑郁模型,REMSD组采用小平台水环境法,用大平台水环境做对照,排除水环境的影响。采用强迫游泳的方法记录大鼠在24℃-25℃水温中连续5min强迫游泳的不动时间,观察大鼠抑郁行为及情绪的变化;采用高效液相色谱仪紫外检测器(high-performance liquid chromatography with Photodiode Array Detector,HPLC-PAD)检测大鼠海马、下丘脑、纹状体脑区匀浆液中腺苷(adenosine)、肌苷(inosine)含量的变化,采用高效液相色谱仪-荧光检测器(high-performance liquid chromatography with fluorescence detection,HPLC-FLD)检测大鼠海马、下丘脑、纹状体脑区单胺类神经递质的含量。实验结果用EXCEL 2003和SPSS 13.0统计软件进行分析处理。结果以均数±标准误(X±SE)表示,采用单因素方差分析(ANOVA)和t检验(T-test)进行组间均数比较,P<0.05为差异显著。
结果:1.经过21天慢性应激后,抑郁模型组与REMSD组大鼠5min强迫游泳的不动时间与应激前相比显著延长;抑郁模型大鼠经过72小时REMSD后5min强迫游泳的不动时间显著减少;而大平台对照组大鼠经过21天慢性应激后5min强迫游泳的不动时间与应激前相比差别却无显著性(造模失败),经过72小时大平台水环境处理后大鼠5min强迫游泳的不动时间与慢性应激后相比差别依旧无显著性。
2.正常对照组海马去甲肾上腺素(norepinephrine, NE)含量为(0.7854±0.2230)ng/mg,与抑郁模型组海马NE含量(0.7039±0.2256)ng/mg相比差异不显著(P﹥0.05),REMSD组海马NE含量(0.9589±0.2056)ng/mg显著高于抑郁模型组(P﹤0.05);正常对照组海马5-羟色胺(Serotonin, 5-HT)含量为(0.2502±0.1253)ng/mg,抑郁模型组海马5-HT含量(0.1562±0.0696)ng/mg与之相比显著减少(P﹤0.05), REMSD组海马5-HT含量(0.3903±0.0996)ng/mg与抑郁模型组相比显著增高,差别有极显著性(P﹤0.001);利用HPLC-FLD未能检测出海马多巴胺(dopamine, DA)含量。
3.正常对照组下丘脑NE含量为(4.7066±1.2337)ng/mg,与抑郁模型组下丘脑NE含量(4.3204±0.9312)ng/mg相比差异不显著(P﹥0.05),REMSD组下丘脑NE含量(4.9031±0.9962)ng/mg与抑郁模型组相比有一定程度的升高,但无显著性差异;正常对照组下丘脑5-HT含量为(0.4850±0.0878)ng/mg,抑郁模型组含量(0.3350±0.1923)ng/mg与之相比有一定程度的减少,但无显著性差异,REMSD组下丘脑5-HT含量为(0.7088±0.2903)ng/mg与抑郁模型组相比显著增高(P﹤0.01);利用HPLC-FLD仅能检测到正常对照组下丘脑的DA含量(3.8347±2.3783)ng/mg,其它几组未检测出。
4.利用HPLC-FLD未能检测出各组纹状体脑区NE的含量;正常对照组纹状体脑区5-HT含量为(0.4612±0.3313)ng/mg, REMSD组为(0.3474±0.1677)ng/mg,两者相比差别无显著性(P﹥0.05),利用HPLC-FLD未能检测出抑郁模型组纹状体脑区5-HT的含量;正常对照组纹状体DA含量为(10.6081±2.8886)ng/mg,抑郁模型组为(10.3086±0.3029)ng/mg,与正常对照组相比差别无显著性(P﹥0.05),REMSD组纹状体DA含量为(15.3682±2.6326)ng/mg,较抑郁模型组显著增高,差别有极显著性(P=0.000)。
5.正常对照组海马腺苷含量为(138.7945±27.1076)ng/mg,抑郁模型组含量(101.3403±28.0018)ng/mg与之相比显著减少(P﹤0.05), REMSD组海马腺苷含量为(69.3919±9.7863)ng/mg,与抑郁模型组相比亦显著减少(P﹤0.05);正常对照组海马肌苷含量为(25.5273±6.6599)ng/mg,抑郁模型组含量(17.9310±5.5740)ng/mg与之相比显著减少(P﹤0.05),REMSD组海马肌苷含量(10.0350±1.5874)ng/mg与抑郁模型组相比亦显著减少(P﹤0.05)。
6.正常对照组下丘脑腺苷含量为(96.6095±30.0754)ng/mg,抑郁模型组含量(64.3065±19.4130)ng/mg与之相比显著减少(P﹤0.05),REMSD组下丘脑腺苷含量(34.6534±6.0269)ng/mg与抑郁模型组相比亦显著减少(P﹤0.01);正常对照组下丘脑肌苷含量为(52.6599±13.1163)ng/mg,抑郁模型组含量(37.9938±3.7118)ng/mg与之比较显著减少(P﹤0.05),REMSD组下丘脑肌苷含量(27.6484±6.7490)ng/mg与抑郁模型组相比亦显著减少(P﹤0.01)。
7.正常对照组纹状体腺苷含量为(180.7181±36.0231)ng/mg,抑郁模型组含量(137.0903±25.4030)ng/mg与之相比显著减少(P﹤0.05),REMSD组下丘脑腺苷含量(143.6780±43.0931)ng/mg与抑郁模型组相比差别不显著(P﹥0.05);正常对照组大鼠纹状体肌苷含量为(27.2072±6.2801)ng/mg,抑郁模型组含量(21.7120±5.1758)ng/mg与之相比有一定程度减少,但无显著性差异(P=0.077),REMSD组纹状体肌苷含量(13.8737±2.6891)ng/mg与抑郁模型组相比显著减少(P﹤0.01)。
结论:1.72小时REMSD可以明显改善慢性轻度不可预见性应激引起的大鼠抑郁样行为;
2.海马部位5-HT和NE含量的增高可能在REMSD快速抗抑郁过程中起着重要的作用;
3.下丘脑部位5-HT含量的增高可能在REMSD快速抗抑郁过程中起一定的作用,但下丘脑NE神经传递可能在REMSD的抗抑郁过程中不起主要作用;
4.海马部位和下丘脑部位的腺苷含量变化可能参与了REMSD的抗抑郁过程。上述结果提示:REMSD可以逆转CMUS抑郁模型大鼠的抑郁样行为,其快速抗抑郁作用可能与增强海马及下丘脑部位的5-HT神经传递有关,腺苷可能参与了REMSD的抗抑郁过程。
Backgrounds and Objective: Adenosine as a neuromodulator can modulate some neurotransmitters by integrating with adenosine receptors. On the other hand, adenosine is also thought to be a sleep factor.At present, a few studies indicate that adenosine and its receptors have some relations with the process of depression and antidepressant. It is known that sleep deprivation has an antidepressant-like effect. But up to now, the mechanisms of the acute antidepressant effect of sleep deprivation have not been clarified.Most of previous studies focused mainly on monoamine neurotransmitter systems. Some interactional factors may take part in this progress,we think. Is adenosine involved in sleep deprivation? Whether the concentration of adenosine in different encephalic regions change or not during the process? What are the relations between the adenosine and the monoamine neurotransmitters? There are questions remained at present.The current study is designed to investigate the effects of 72-hour REM (rapid eye movement) sleep deprivation on monoamines and adenosine in rats treated with chronic mild unpredicted stresses.
Materials and Methods: Sprague-Dawley rats were divided into four groups randomly: 1) normal control group, 2)the depression-model group,3)depression-model +sleep deprivation group,4)depression-model +72 hours tank control group. Two classical models were adopted to build depression-model,one is chronic mild unpredicted stresses,and the other is sub-raising. The REMSD group used a small platform water environment, in order to remove the influence of water environment, a big platform was used as a control condition. Using the forced swimming test observe the change of the duration of immobility of rats which was immersed in the water of 25℃temperature for 5 min; using the high-performance liquid chromatography with Photodiode Array Detector detect the concentration of adenosine and inosine in hippocampus, hypothalamus and striatum; using the high-performance liquid chromatography with fluorescence detection detect concentration of monoamines. Excel 2003 and SPSS 13.0 statistical software were used for the analysis.
Results:1)After chronic mild unpredicted stresses, the duration of immobility of rats increased in the groups which were adopted the treatment,corresponding,it decreased in the REMSD group after 72h rapid eye movement sleep deprivation.2)5-HT and NE levels were found to have significant differences in the hippocampus or hypothalamus among the different groups .The concentration of dopamine in hippocampus and hypothalamus couldn’t be detected by HPLC-FLD. The concentration of dopamine in striatum of REMSD group is higher than that of depression-model group.3)Adenosine and inosine levels were found to have significant differences in hippocampus or hypothalamus among the different groups.
Conclusions:
1.72 hours of REMSD could significantly improve the depressive behavior induced by chronic mild unpredicted stresses;
2.REMSD increases 5-HT and NE concentrations in Hippocampus, which play an important role in rapid antidepressant;
3.5-HT in hypothalamus seems to be related to the process of REMSD antidepressant, but not norepinephrine;
4.The change of adenosine in Hippocampus and Hypothalamus may take part in the REMSD antidepressant process,
These results suggest that the REMSD reverses chronic mild unpredictable stresses-induced depressive behavior in rats. The enhanced 5-HT transportation by REMSD is one of the reasons in rapid antidepressant effect,and adenosine may take parts in this process.
目的:观察睡眠剥夺对慢性应激抑郁模型大鼠不同脑区组织匀浆液中单胺递质及腺苷、肌苷含量的影响。
材料和方法:实验动物为Sprague Dawley 2-3月龄雄性大鼠,随机分为正常对照组,抑郁模型组,抑郁模型+72小时快眼动睡眠剥夺组(以下称REMSD组),抑郁模型组+72小时大平台对照组(以下称大平台对照组),每组8只。应激造模采用慢性轻度不可预见性应激(chronic mild unpredicted stress, CMUS)和分养两种经典模型结合的方式来建立抑郁模型,REMSD组采用小平台水环境法,用大平台水环境做对照,排除水环境的影响。采用强迫游泳的方法记录大鼠在24℃-25℃水温中连续5min强迫游泳的不动时间,观察大鼠抑郁行为及情绪的变化;采用高效液相色谱仪紫外检测器(high-performance liquid chromatography with Photodiode Array Detector,HPLC-PAD)检测大鼠海马、下丘脑、纹状体脑区匀浆液中腺苷(adenosine)、肌苷(inosine)含量的变化,采用高效液相色谱仪-荧光检测器(high-performance liquid chromatography with fluorescence detection,HPLC-FLD)检测大鼠海马、下丘脑、纹状体脑区单胺类神经递质的含量。实验结果用EXCEL 2003和SPSS 13.0统计软件进行分析处理。结果以均数±标准误(X±SE)表示,采用单因素方差分析(ANOVA)和t检验(T-test)进行组间均数比较,P<0.05为差异显著。
结果:1.经过21天慢性应激后,抑郁模型组与REMSD组大鼠5min强迫游泳的不动时间与应激前相比显著延长;抑郁模型大鼠经过72小时REMSD后5min强迫游泳的不动时间显著减少;而大平台对照组大鼠经过21天慢性应激后5min强迫游泳的不动时间与应激前相比差别却无显著性(造模失败),经过72小时大平台水环境处理后大鼠5min强迫游泳的不动时间与慢性应激后相比差别依旧无显著性。
2.正常对照组海马去甲肾上腺素(norepinephrine, NE)含量为(0.7854±0.2230)ng/mg,与抑郁模型组海马NE含量(0.7039±0.2256)ng/mg相比差异不显著(P﹥0.05),REMSD组海马NE含量(0.9589±0.2056)ng/mg显著高于抑郁模型组(P﹤0.05);正常对照组海马5-羟色胺(Serotonin, 5-HT)含量为(0.2502±0.1253)ng/mg,抑郁模型组海马5-HT含量(0.1562±0.0696)ng/mg与之相比显著减少(P﹤0.05), REMSD组海马5-HT含量(0.3903±0.0996)ng/mg与抑郁模型组相比显著增高,差别有极显著性(P﹤0.001);利用HPLC-FLD未能检测出海马多巴胺(dopamine, DA)含量。
3.正常对照组下丘脑NE含量为(4.7066±1.2337)ng/mg,与抑郁模型组下丘脑NE含量(4.3204±0.9312)ng/mg相比差异不显著(P﹥0.05),REMSD组下丘脑NE含量(4.9031±0.9962)ng/mg与抑郁模型组相比有一定程度的升高,但无显著性差异;正常对照组下丘脑5-HT含量为(0.4850±0.0878)ng/mg,抑郁模型组含量(0.3350±0.1923)ng/mg与之相比有一定程度的减少,但无显著性差异,REMSD组下丘脑5-HT含量为(0.7088±0.2903)ng/mg与抑郁模型组相比显著增高(P﹤0.01);利用HPLC-FLD仅能检测到正常对照组下丘脑的DA含量(3.8347±2.3783)ng/mg,其它几组未检测出。
4.利用HPLC-FLD未能检测出各组纹状体脑区NE的含量;正常对照组纹状体脑区5-HT含量为(0.4612±0.3313)ng/mg, REMSD组为(0.3474±0.1677)ng/mg,两者相比差别无显著性(P﹥0.05),利用HPLC-FLD未能检测出抑郁模型组纹状体脑区5-HT的含量;正常对照组纹状体DA含量为(10.6081±2.8886)ng/mg,抑郁模型组为(10.3086±0.3029)ng/mg,与正常对照组相比差别无显著性(P﹥0.05),REMSD组纹状体DA含量为(15.3682±2.6326)ng/mg,较抑郁模型组显著增高,差别有极显著性(P=0.000)。
5.正常对照组海马腺苷含量为(138.7945±27.1076)ng/mg,抑郁模型组含量(101.3403±28.0018)ng/mg与之相比显著减少(P﹤0.05), REMSD组海马腺苷含量为(69.3919±9.7863)ng/mg,与抑郁模型组相比亦显著减少(P﹤0.05);正常对照组海马肌苷含量为(25.5273±6.6599)ng/mg,抑郁模型组含量(17.9310±5.5740)ng/mg与之相比显著减少(P﹤0.05),REMSD组海马肌苷含量(10.0350±1.5874)ng/mg与抑郁模型组相比亦显著减少(P﹤0.05)。
6.正常对照组下丘脑腺苷含量为(96.6095±30.0754)ng/mg,抑郁模型组含量(64.3065±19.4130)ng/mg与之相比显著减少(P﹤0.05),REMSD组下丘脑腺苷含量(34.6534±6.0269)ng/mg与抑郁模型组相比亦显著减少(P﹤0.01);正常对照组下丘脑肌苷含量为(52.6599±13.1163)ng/mg,抑郁模型组含量(37.9938±3.7118)ng/mg与之比较显著减少(P﹤0.05),REMSD组下丘脑肌苷含量(27.6484±6.7490)ng/mg与抑郁模型组相比亦显著减少(P﹤0.01)。
7.正常对照组纹状体腺苷含量为(180.7181±36.0231)ng/mg,抑郁模型组含量(137.0903±25.4030)ng/mg与之相比显著减少(P﹤0.05),REMSD组下丘脑腺苷含量(143.6780±43.0931)ng/mg与抑郁模型组相比差别不显著(P﹥0.05);正常对照组大鼠纹状体肌苷含量为(27.2072±6.2801)ng/mg,抑郁模型组含量(21.7120±5.1758)ng/mg与之相比有一定程度减少,但无显著性差异(P=0.077),REMSD组纹状体肌苷含量(13.8737±2.6891)ng/mg与抑郁模型组相比显著减少(P﹤0.01)。
结论:1.72小时REMSD可以明显改善慢性轻度不可预见性应激引起的大鼠抑郁样行为;
2.海马部位5-HT和NE含量的增高可能在REMSD快速抗抑郁过程中起着重要的作用;
3.下丘脑部位5-HT含量的增高可能在REMSD快速抗抑郁过程中起一定的作用,但下丘脑NE神经传递可能在REMSD的抗抑郁过程中不起主要作用;
4.海马部位和下丘脑部位的腺苷含量变化可能参与了REMSD的抗抑郁过程。上述结果提示:REMSD可以逆转CMUS抑郁模型大鼠的抑郁样行为,其快速抗抑郁作用可能与增强海马及下丘脑部位的5-HT神经传递有关,腺苷可能参与了REMSD的抗抑郁过程。
Backgrounds and Objective: Adenosine as a neuromodulator can modulate some neurotransmitters by integrating with adenosine receptors. On the other hand, adenosine is also thought to be a sleep factor.At present, a few studies indicate that adenosine and its receptors have some relations with the process of depression and antidepressant. It is known that sleep deprivation has an antidepressant-like effect. But up to now, the mechanisms of the acute antidepressant effect of sleep deprivation have not been clarified.Most of previous studies focused mainly on monoamine neurotransmitter systems. Some interactional factors may take part in this progress,we think. Is adenosine involved in sleep deprivation? Whether the concentration of adenosine in different encephalic regions change or not during the process? What are the relations between the adenosine and the monoamine neurotransmitters? There are questions remained at present.The current study is designed to investigate the effects of 72-hour REM (rapid eye movement) sleep deprivation on monoamines and adenosine in rats treated with chronic mild unpredicted stresses.
Materials and Methods: Sprague-Dawley rats were divided into four groups randomly: 1) normal control group, 2)the depression-model group,3)depression-model +sleep deprivation group,4)depression-model +72 hours tank control group. Two classical models were adopted to build depression-model,one is chronic mild unpredicted stresses,and the other is sub-raising. The REMSD group used a small platform water environment, in order to remove the influence of water environment, a big platform was used as a control condition. Using the forced swimming test observe the change of the duration of immobility of rats which was immersed in the water of 25℃temperature for 5 min; using the high-performance liquid chromatography with Photodiode Array Detector detect the concentration of adenosine and inosine in hippocampus, hypothalamus and striatum; using the high-performance liquid chromatography with fluorescence detection detect concentration of monoamines. Excel 2003 and SPSS 13.0 statistical software were used for the analysis.
Results:1)After chronic mild unpredicted stresses, the duration of immobility of rats increased in the groups which were adopted the treatment,corresponding,it decreased in the REMSD group after 72h rapid eye movement sleep deprivation.2)5-HT and NE levels were found to have significant differences in the hippocampus or hypothalamus among the different groups .The concentration of dopamine in hippocampus and hypothalamus couldn’t be detected by HPLC-FLD. The concentration of dopamine in striatum of REMSD group is higher than that of depression-model group.3)Adenosine and inosine levels were found to have significant differences in hippocampus or hypothalamus among the different groups.
Conclusions:
1.72 hours of REMSD could significantly improve the depressive behavior induced by chronic mild unpredicted stresses;
2.REMSD increases 5-HT and NE concentrations in Hippocampus, which play an important role in rapid antidepressant;
3.5-HT in hypothalamus seems to be related to the process of REMSD antidepressant, but not norepinephrine;
4.The change of adenosine in Hippocampus and Hypothalamus may take part in the REMSD antidepressant process,
These results suggest that the REMSD reverses chronic mild unpredictable stresses-induced depressive behavior in rats. The enhanced 5-HT transportation by REMSD is one of the reasons in rapid antidepressant effect,and adenosine may take parts in this process.
引文
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