长时程睡眠剥夺对大鼠行为记忆和海马区递质的影响及四神聪电针的预防作用
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摘要
【目的】探讨连续长时程睡眠剥夺对大鼠行为、记忆和海马区递质的影响及四神聪电针的预防作用。【方法】采用多平台法进行连续7 d睡眠剥夺造模。实验分阶段进行,一阶段(实验一):将30只SD雌性大鼠随机分成环境对照组(TC组),睡眠剥夺组(SD组),睡眠剥夺结束后3 d组(3 d after SD)、6 d组(6 d after SD)和11 d组(11d after SD),即7 d睡眠剥夺结束后再在常规大鼠饲养笼内饲养不同时间,并让大鼠恢复正常睡眠;每组大鼠均为6只。采用旷场实验观察大鼠自发活动。二阶段(实验二):将18只大鼠随机分成环境对照组(TC组)、睡眠剥夺组(SD组)、电针组(SD+EA组),每组6只。SD+EA组于睡眠剥夺期间每天上午电针一次,每次20 min,共7 d。采用Morris水迷宫实验观察各组大鼠空间学习记忆能力。另外,两阶段的酶联免疫吸附测定均在旷场实验和水迷宫检测结束后取材进行。【结果】实验一,旷场实验:SD组与TC组比,大鼠水平运动和垂直运动的得分减少明显(P<0.05);但睡眠恢复3 d、6 d、11 d后,3 d组、6 d组、11 d组的大鼠水平运动和垂直运动得分恢复明显,与SD组比较差异有统计学意义(P<0.05),但与TC组比,3 d组、6 d组虽存在差异(P<0.05),但11 d组差异已无显著性(P>0.05)。大鼠海马区GABA检测:SD组与TC组比,睡眠剥夺后导致GABA表达下降(P<0.05);但睡眠恢复3 d、6 d后,GABA的表达明显上升,与SD组比较差异有统计学意义(P<0.05),而与TC组比差异已无显著性(P>0.05);但到第11天时,GABA表达相对于TC组和6 d组又有下降趋势,但与TC组和6 d组相比较无统计学差异(P>0.05)。实验二:Morris水迷宫检测结果显示,与TC组和电针组比,SD组水下平台的逃避潜伏期明显延长(P<0.05),而穿越原平台所在区次数明显减少(P<0.05);相反,电针组与TC组相比,无论是水下平台的逃避潜伏期还是穿越原平台所在区次数,均无统计学意义(P>0.05)。大鼠海马区的5-HT和GABA检测:5-HT的表达,与TC组相比,SD组变化不大(P>0.05);但电针组与TC组比有下降趋势,且差异有统计学意义(P<0.05)。GABA的表达与5-HT的表达刚好相反,与TC组比,SD组海马GABA的表达有下降趋势(P<0.05);但电针组与TC组比变化不大(P>0.05)。【结论】7 d连续睡眠剥夺可导致大鼠行为、记忆能力明显下降,且对大鼠海马区GABA和5-HT表达的影响有异于既往文献报道的少于4 d连续睡眠剥夺的影响。四神聪电针具有预防长时程睡眠剥夺对大鼠行为记忆损害的作用,其机制可能是通过电针改善因睡眠剥夺而导致的紊乱的中枢神经递质。
【Objective】To investigate the effects of continuous long-term sleep deprivation on rat's behavior,memory and neurotransmitters in hippocampus,as well as determine the effects of prevention of electro-acupuncture.【Method】We used multi-platform water environment to make the rat model of insomnia.The experiment was divided into two sections. Section one:thirty rats were divided into 5 groups(six rats each group):tank control group(TC),sleep-deprived for continuous 7 days group(SD group),three days of recovery after SD group(3 d after SD group),six days of recovery after SD group(6 d after SD group),eleven days of recovery after SD group(11 d after SD group). The term of recovery refers to normal feeding and sleeping when rats return to cages after sleep-deprivation. The open-field test was performed following the end time of each group experiment and then the rats were perfused and fixed. Section two:Eighteen rats were divided into three groups(six rats each group):tank control group(TC),sleep-deprived for continuous 7 days group(SD group),sleep-deprived rats treated with electro-acupuncture group(SD+EA group). Rats of SD group and SD+EA group were deprived sleep for continuous 7 days. During the period of sleep deprivation,the rats of SD+EA group were treated with electro-acupuncture 20 minutes every morning. At the end time of experiment,rats were tested by the Morris water maze and then perfused and get specimens.【Results】Section one:the statistical analysis of outcome of open-field test showed:as compared to the TC group,the scores of rat's horizontal movement and vertical movement were significantly fewer in the SD group(P<0.05). After 3,6,or 11 days recovery of sleep,the scores increased significantly as compared with SD group(P<0.05). Although,the outcome of open-field test in the 3 d after SD and 6 d after SD groups had the significant difference with TC group(P<0.05),there was no different between the 11 d after SD group and TC group(P>0.05).The quantitative analysis of gamma aminobutyric acid(GABA)expression in the hippocampus showed:the SD group shows lower level of GABA when compared to the TC group(P<0.05). After 3 days or 6 days of recovery,the expression of GABA is higher than the SD group(P<0.05),and similar to the TC group(P>0.05). In the 11 days after SD group,the expression of GABA declined as compared to the TC group and the 6 d after SD group(P<0.05). Section two:In the Morris water maze experiment,when compared to the TC group and the SD+EA group,the escape latency period was significantly prolonged and the times of across the platform were significantly reduced in the SD group(P<0.05). On the contrary,there were no statistical differences in the escape latency period or the across platform times between the SD+EA group and the TC group(P>0.05).The results of serotonin(5-HT)and GABA in the hippocampus showed:the 5-HT level in the SD group was no significant difference as compared to the TC group and the SD group on 5-HT level,while the 5-HT level in the SD+EA group was reduced as compared to the TC group(P<0.05). In the opposite,the GABA level in the SD group was decreased as compared to the TC group(P<0.05),while the SD+EA group and the TC group appear similarly level(P>0.05)【.Conclusions】The effect of 7 days' continuous long-term sleep-deprivation may lead to the reduction in the ability of learning and memory behavior of rats. Our results is different with 4 days' continuously sleep-deprivation which was reported on the rats' expression of GABA and 5-HT.Electro-acupuncture stimulating the Sishengcong Acupoints seems preventing the harming of long-term sleep-deprivation on the ability of learning and memory,which probably is through adjusting the disorder of central neurotransmitter expression caused by sleep deprivation.
【Objective】To investigate the effects of continuous long-term sleep deprivation on rat's behavior,memory and neurotransmitters in hippocampus,as well as determine the effects of prevention of electro-acupuncture.【Method】We used multi-platform water environment to make the rat model of insomnia.The experiment was divided into two sections. Section one:thirty rats were divided into 5 groups(six rats each group):tank control group(TC),sleep-deprived for continuous 7 days group(SD group),three days of recovery after SD group(3 d after SD group),six days of recovery after SD group(6 d after SD group),eleven days of recovery after SD group(11 d after SD group). The term of recovery refers to normal feeding and sleeping when rats return to cages after sleep-deprivation. The open-field test was performed following the end time of each group experiment and then the rats were perfused and fixed. Section two:Eighteen rats were divided into three groups(six rats each group):tank control group(TC),sleep-deprived for continuous 7 days group(SD group),sleep-deprived rats treated with electro-acupuncture group(SD+EA group). Rats of SD group and SD+EA group were deprived sleep for continuous 7 days. During the period of sleep deprivation,the rats of SD+EA group were treated with electro-acupuncture 20 minutes every morning. At the end time of experiment,rats were tested by the Morris water maze and then perfused and get specimens.【Results】Section one:the statistical analysis of outcome of open-field test showed:as compared to the TC group,the scores of rat's horizontal movement and vertical movement were significantly fewer in the SD group(P<0.05). After 3,6,or 11 days recovery of sleep,the scores increased significantly as compared with SD group(P<0.05). Although,the outcome of open-field test in the 3 d after SD and 6 d after SD groups had the significant difference with TC group(P<0.05),there was no different between the 11 d after SD group and TC group(P>0.05).The quantitative analysis of gamma aminobutyric acid(GABA)expression in the hippocampus showed:the SD group shows lower level of GABA when compared to the TC group(P<0.05). After 3 days or 6 days of recovery,the expression of GABA is higher than the SD group(P<0.05),and similar to the TC group(P>0.05). In the 11 days after SD group,the expression of GABA declined as compared to the TC group and the 6 d after SD group(P<0.05). Section two:In the Morris water maze experiment,when compared to the TC group and the SD+EA group,the escape latency period was significantly prolonged and the times of across the platform were significantly reduced in the SD group(P<0.05). On the contrary,there were no statistical differences in the escape latency period or the across platform times between the SD+EA group and the TC group(P>0.05).The results of serotonin(5-HT)and GABA in the hippocampus showed:the 5-HT level in the SD group was no significant difference as compared to the TC group and the SD group on 5-HT level,while the 5-HT level in the SD+EA group was reduced as compared to the TC group(P<0.05). In the opposite,the GABA level in the SD group was decreased as compared to the TC group(P<0.05),while the SD+EA group and the TC group appear similarly level(P>0.05)【.Conclusions】The effect of 7 days' continuous long-term sleep-deprivation may lead to the reduction in the ability of learning and memory behavior of rats. Our results is different with 4 days' continuously sleep-deprivation which was reported on the rats' expression of GABA and 5-HT.Electro-acupuncture stimulating the Sishengcong Acupoints seems preventing the harming of long-term sleep-deprivation on the ability of learning and memory,which probably is through adjusting the disorder of central neurotransmitter expression caused by sleep deprivation.
引文
[1]王博文,徐兆慧,杜勇,等.不同方式的睡眠剥夺对大鼠学习记忆的影响[J].西安交通大学学报(医学版),2014,35(02):213-217.Wang BW,Xu ZH,Du Y,et al.Effects of different ways of sleep deprivation on learning and memory in rats[J].J Xi'an Jiaotong Univ(Med Sci),2014,35(02):213-217.
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[4]Alhaider IA,Alkadhi KA.Caffeine treatment prevents rapid eye movement sleep deprivation-induced impairment of late-phase long-term potentiation in the dentate gyrus[J].Eur J Neurosci,2015,42(10):2843-2850.
[5]Mandiroglu S,Ozdilekcan C.Impact of acupuncture on chronic insomnia:A report of two cases with polysomnographic evaluation[J].J Acupunct Meridian Stud,2017,10(2):135-138.
[6]李继元,宋云龙,齐建林.睡眠剥夺对健康成人学习记忆网络影响的功能磁共振研究[J].安徽医科大学学报,2017,52(09):1335-1340.Li JY,Song YL,Qi JL,et al.Study on the effects of sleep deprivation on the learning and memory network of healthy adults[J].J Anhui Med Univ,2017,52(09):1335-1340.
[7]马双双,齐阳,王莹,等.连续睡眠剥夺对大鼠认知能力?神经递质及海马结构的影响[J].海军医学杂志,2013,34(02):15-18.Ma SS,Qi Y,Wang Y,et al.Effects of continuous sleep deprivation on cognitive ability,neurotransmitter and hippocampus in rats[J].J Naval Med Sci,2013,34(02):15-18.
[8]王新兴,梅竹松,刘伟丽,等.睡眠剥夺应激对大鼠海马的损伤作用及其对探究行为的影响[J].军事医学,2011,35(09):663-665.Wang XX,Mei ZS,Liu WL,et al.Effects of sleep deprivation stress on hippocampus injury in rats and its effect on exploratory behavior[J].J Mil Med,2011,35(09):663-665.
[9]杨波,岳明.酸枣仁汤对睡眠剥夺大鼠认知功能及海马NA,GABA,5-HT的影响[J].陕西中医,2015,36(1):122-123Yang B,Yue M.Effects of ziziphus jujuba decoction on cognitive function and NA,GABA and 5-HT in rats with sleep deprivation[J].Shanxi Tradit Chin Med,2015,36(1):122-123
[10]木日根吉雅,阿古拉.蒙医温针对失眠大鼠海马中Glu GABA ACh含量的影响[J].中国民族医药杂志,2014,20(6):43-44Murigen JY,Aguera.Effect of mongolian warming on Glu,GABA and ACh contents in hippocampus of insomnia rats[J].J Med Pharm Chin Minorities,2014,20(6):43-44
[11]李振.GABA能药物干预对REM睡眠剥夺大鼠认知功能影响的实验研究[D].第二军医大学,2008:20-22Li Z.Experimental study on the effect of GABAergic drug intervention on cognitive function in REM sleep deprived rats[D].Second Mil Med Univ,2008:20-22.
[12]Brady K,Brown JW,Thurmond JB.Behavioral and neurochemical effects of dietary tyrosine in young and aged mice following cold-swim stress[J].Pharmacol Biochem Behav,1980,12(5):667-674.
[13]刘晶,李峰,宋月晗,等.GABA能系统与失眠[J].现代生物医学进展,2012,12(18):3589-3591.Liu J,Li F,Song YH,et al.GABAergic system and insomnia[J].Adv Modern Biol,2012,12(18):3589-3591.
[14]Rechtschaffen A,Gilliland MA,Bergmann BM,et al.Physiological correlates of prolonged sleep deprivation in rats[J].Science,1983,221(4606):182-184.
[15]曾荣,邵淑丽,徐兴军,等.长期异相睡眠剥夺对大鼠能量代谢及血清甲状腺素水平的影响[J].中国应用生理学杂志,2010,26(1):105-108.Zen R,Shao SL,Xu XJ,et al.Effects of longterm sleep deprivation on energy metabolism and serum thyroxine levels in rats[J].Chin J Appl Physiol,2010,26(1):105-108.
[16]王云,岳云.异丙酚对异相睡眠剥夺大鼠海马谷氨酸和γ-氨基丁酸释放的影响[J].中华麻醉学杂志,2008,28(1):7-9.Wang Y,Yue Y.Effects of propofol on the release of glutamate andγ-aminobutyric acid in hippocampus of rats with heterogeneous sleep deprivation[J].Chin J Anesth,2008,28(1):7-9.
[17]王敏建,况利.脑海马结构与抑郁障碍的关系[J].中国临床康复,2005,9(48):116-118.Wang MJ,Kuang L.The relationship between hippocampus and depressive disorder[J].Chin J Clin Rehabil,2005,9(48):116-118.
[18]Amat J,Baratta MV,Paul E,et al.Medial prefrontal cortex determines how stressorcontrollabilityaffects behavior and dorsal raphe nucleus[J].Nat Neurosci,2005,8(3):365-371.
[19]钟静瑜,黄俊山.5-羟色胺与睡眠的研究进展[J].医学综述,2010,16(10):1471-1473.Zhong JY,Huang JS.Advances in serotonin and sleep[J].Med Rev,2010,16(10):1471-1473.
[2]项恒,梁曼.中国民航管制学员注意力差异分析[J].科学技术与工程,2014,14(12):141-145.Xiang H,Liang M.An analysis of attracting differences of chinese civil aviation control[J].Sci Tech Eng,2014,14(12):141-145
[3]How JM,Foo SC,Low E,et al.Effects of sleep deprivation on performance of Naval seamen:I.Total sleep deprivation on performance[J].Ann Acad Med Singapore,1994,23(5):669-675.
[4]Alhaider IA,Alkadhi KA.Caffeine treatment prevents rapid eye movement sleep deprivation-induced impairment of late-phase long-term potentiation in the dentate gyrus[J].Eur J Neurosci,2015,42(10):2843-2850.
[5]Mandiroglu S,Ozdilekcan C.Impact of acupuncture on chronic insomnia:A report of two cases with polysomnographic evaluation[J].J Acupunct Meridian Stud,2017,10(2):135-138.
[6]李继元,宋云龙,齐建林.睡眠剥夺对健康成人学习记忆网络影响的功能磁共振研究[J].安徽医科大学学报,2017,52(09):1335-1340.Li JY,Song YL,Qi JL,et al.Study on the effects of sleep deprivation on the learning and memory network of healthy adults[J].J Anhui Med Univ,2017,52(09):1335-1340.
[7]马双双,齐阳,王莹,等.连续睡眠剥夺对大鼠认知能力?神经递质及海马结构的影响[J].海军医学杂志,2013,34(02):15-18.Ma SS,Qi Y,Wang Y,et al.Effects of continuous sleep deprivation on cognitive ability,neurotransmitter and hippocampus in rats[J].J Naval Med Sci,2013,34(02):15-18.
[8]王新兴,梅竹松,刘伟丽,等.睡眠剥夺应激对大鼠海马的损伤作用及其对探究行为的影响[J].军事医学,2011,35(09):663-665.Wang XX,Mei ZS,Liu WL,et al.Effects of sleep deprivation stress on hippocampus injury in rats and its effect on exploratory behavior[J].J Mil Med,2011,35(09):663-665.
[9]杨波,岳明.酸枣仁汤对睡眠剥夺大鼠认知功能及海马NA,GABA,5-HT的影响[J].陕西中医,2015,36(1):122-123Yang B,Yue M.Effects of ziziphus jujuba decoction on cognitive function and NA,GABA and 5-HT in rats with sleep deprivation[J].Shanxi Tradit Chin Med,2015,36(1):122-123
[10]木日根吉雅,阿古拉.蒙医温针对失眠大鼠海马中Glu GABA ACh含量的影响[J].中国民族医药杂志,2014,20(6):43-44Murigen JY,Aguera.Effect of mongolian warming on Glu,GABA and ACh contents in hippocampus of insomnia rats[J].J Med Pharm Chin Minorities,2014,20(6):43-44
[11]李振.GABA能药物干预对REM睡眠剥夺大鼠认知功能影响的实验研究[D].第二军医大学,2008:20-22Li Z.Experimental study on the effect of GABAergic drug intervention on cognitive function in REM sleep deprived rats[D].Second Mil Med Univ,2008:20-22.
[12]Brady K,Brown JW,Thurmond JB.Behavioral and neurochemical effects of dietary tyrosine in young and aged mice following cold-swim stress[J].Pharmacol Biochem Behav,1980,12(5):667-674.
[13]刘晶,李峰,宋月晗,等.GABA能系统与失眠[J].现代生物医学进展,2012,12(18):3589-3591.Liu J,Li F,Song YH,et al.GABAergic system and insomnia[J].Adv Modern Biol,2012,12(18):3589-3591.
[14]Rechtschaffen A,Gilliland MA,Bergmann BM,et al.Physiological correlates of prolonged sleep deprivation in rats[J].Science,1983,221(4606):182-184.
[15]曾荣,邵淑丽,徐兴军,等.长期异相睡眠剥夺对大鼠能量代谢及血清甲状腺素水平的影响[J].中国应用生理学杂志,2010,26(1):105-108.Zen R,Shao SL,Xu XJ,et al.Effects of longterm sleep deprivation on energy metabolism and serum thyroxine levels in rats[J].Chin J Appl Physiol,2010,26(1):105-108.
[16]王云,岳云.异丙酚对异相睡眠剥夺大鼠海马谷氨酸和γ-氨基丁酸释放的影响[J].中华麻醉学杂志,2008,28(1):7-9.Wang Y,Yue Y.Effects of propofol on the release of glutamate andγ-aminobutyric acid in hippocampus of rats with heterogeneous sleep deprivation[J].Chin J Anesth,2008,28(1):7-9.
[17]王敏建,况利.脑海马结构与抑郁障碍的关系[J].中国临床康复,2005,9(48):116-118.Wang MJ,Kuang L.The relationship between hippocampus and depressive disorder[J].Chin J Clin Rehabil,2005,9(48):116-118.
[18]Amat J,Baratta MV,Paul E,et al.Medial prefrontal cortex determines how stressorcontrollabilityaffects behavior and dorsal raphe nucleus[J].Nat Neurosci,2005,8(3):365-371.
[19]钟静瑜,黄俊山.5-羟色胺与睡眠的研究进展[J].医学综述,2010,16(10):1471-1473.Zhong JY,Huang JS.Advances in serotonin and sleep[J].Med Rev,2010,16(10):1471-1473.