电针“神门”“三阴交”对失眠大鼠下丘脑室旁核能量代谢的影响
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摘要
目的:观察电针"神门""三阴交"对失眠大鼠下丘脑室旁核(PVN)能量代谢的影响,探讨电针治疗失眠症的中枢调节机制。方法:SD大鼠随机分为正常组、模型组和电针组,每组22只。模型组和电针组采用慢性束缚法束缚15d复制失眠模型。电针组电针"神门""三阴交",每日1次,每次15min,治疗5d。用24h小动物自发活动数据采集系统检测大鼠24h自发活动量,以负重力竭游泳实验检测大鼠负重力竭游泳时间,采用蛋白免疫印迹法检测PVN内的腺苷酸活化蛋白激酶(AMPK)表达,酶联免疫法检测PVN内的乙酰辅酶A(Ac-CoA)、钠-钾三磷酸腺苷酶(Na+-K+-ATPase)含量以及血浆皮质酮(CORT)含量,透射电镜观察PVN的细胞结构。结果:与正常组比较,模型组大鼠白天活动量增多、夜晚活动量减少(P<0.05),负重力竭游泳时间缩短(P<0.05);PVN内AMPK表达增强(P<0.05)、Ac-CoA和Na~+-K~+-ATPase含量降低(P<0.05),血浆CORT含量降低(P<0.05);电镜下可见模型大鼠PVN内线粒体肿胀,核糖体部分消失。电针后,与模型组比较,电针组大鼠白天活动量减少、夜晚活动量增多(P<0.05),负重力竭游泳时间延长(P<0.05);PVN内AMPK表达减弱(P<0.05)、Ac-CoA和NA~+-K~+-ATPase含量升高(P<0.05),血浆CORT含量升高(P<0.05);PVN内线粒体仅轻度肿胀。结论:电针"神门""三阴交"对失眠大鼠下丘脑室旁核能量代谢有良好调整作用,通过调节下丘脑室旁核AMPK表达、Ac-CoA和Na~+-K~+-ATPase含量,电针可以有效改善失眠及失眠后疲劳症状。
Objective To observe the effect of electroacupuncture(EA)at"Shenmen"(HT7)and"Sanyinjiao"(SP6)on energy metabolism in paraventricular nucleus(PVN)of hypothalamus in insomnia rats,so as to explore its mechanism underlying improvement of insomnia.Methods A total of 66 SD rats(half male and half female)were randomized into 3 groups:normal control,model and EA groups(n=22 per group).The insomnia model was established by binding the rat for at least 4 h(step increase of 30 min per day),once daily for 15 days.EA(5 Hz/25 Hz,0.5-1.0 mA)was applied to unilateral HT7 and SP6 for 15 min,once daily for 5 days.The rats' spontaneous activities during day and night were recorded by using the ClockLab Data Collection and Analysis System,and the duration of exhausted swimming was detected by using load-bearing endurance swimming test.The expression of adenosine 5′-monophosphate(AMP)-activated protein kinase(AMPK)of PVN tissue was assayed by Western blot,and the contents of acetyl coenzyme A(Ac-CoA)and Na~+-K~+adenosine triphosphatase(Na+-K+-ATPase)in the PVN tissue,and corticosterone(CORT)in plasma were assayed by ELISA.Changes of the ultrastructure of PVN cells were observed by transmission electron microscope.Results After modeling,the rats' daytime and nocturnal locomotor activities were significantly increased and decreased,respectively(P<0.05),and the duration of exhausted swimming was considerably shortened in the model group compared with that of the normal control group(P<0.05).The expression level of AMPK protein in the PVN was obviously up-regulated(P<0.05),and the contents of Ac-CoA and Na~+-K~+-ATPase in PVN and CORT in plasma were markedly decreased in the model control group relevant to the normal group(P<0.05).After EA intervention,the increased daytime locomotion and the decreased nocturnal activities,the shortened duration of exhausted swimming,the up-regulated expression of AMPK,and the decreased Ac-CoA,Na+-K+-ATPase and CORT contents were all reversed in the EA treated rats relevant to those of the insomnia rats(all P<0.05).Moreover,ultrastructural observation showed mitochondrial swelling and disappearance of partial ribosomes in the plasma of PVN cells in the model group,while in the EA group,only mild swelling of some mitochondria was found,being with basically normal nuclear membrane,mitochondria,rough endoplasmic reticulum,Golgi complex and ribosomes.Conclusion EA at HT7 and SP6 has a positive effect in improving insomnia and insomnia-induced fatigue in insomnia rats,which may be associated with its effects in restraining the expression of AMPK protein,and up-regulating the contents of Ac-CoA and Na~+-K~+-ATPase in PVN and CORT in plasma.
Objective To observe the effect of electroacupuncture(EA)at"Shenmen"(HT7)and"Sanyinjiao"(SP6)on energy metabolism in paraventricular nucleus(PVN)of hypothalamus in insomnia rats,so as to explore its mechanism underlying improvement of insomnia.Methods A total of 66 SD rats(half male and half female)were randomized into 3 groups:normal control,model and EA groups(n=22 per group).The insomnia model was established by binding the rat for at least 4 h(step increase of 30 min per day),once daily for 15 days.EA(5 Hz/25 Hz,0.5-1.0 mA)was applied to unilateral HT7 and SP6 for 15 min,once daily for 5 days.The rats' spontaneous activities during day and night were recorded by using the ClockLab Data Collection and Analysis System,and the duration of exhausted swimming was detected by using load-bearing endurance swimming test.The expression of adenosine 5′-monophosphate(AMP)-activated protein kinase(AMPK)of PVN tissue was assayed by Western blot,and the contents of acetyl coenzyme A(Ac-CoA)and Na~+-K~+adenosine triphosphatase(Na+-K+-ATPase)in the PVN tissue,and corticosterone(CORT)in plasma were assayed by ELISA.Changes of the ultrastructure of PVN cells were observed by transmission electron microscope.Results After modeling,the rats' daytime and nocturnal locomotor activities were significantly increased and decreased,respectively(P<0.05),and the duration of exhausted swimming was considerably shortened in the model group compared with that of the normal control group(P<0.05).The expression level of AMPK protein in the PVN was obviously up-regulated(P<0.05),and the contents of Ac-CoA and Na~+-K~+-ATPase in PVN and CORT in plasma were markedly decreased in the model control group relevant to the normal group(P<0.05).After EA intervention,the increased daytime locomotion and the decreased nocturnal activities,the shortened duration of exhausted swimming,the up-regulated expression of AMPK,and the decreased Ac-CoA,Na+-K+-ATPase and CORT contents were all reversed in the EA treated rats relevant to those of the insomnia rats(all P<0.05).Moreover,ultrastructural observation showed mitochondrial swelling and disappearance of partial ribosomes in the plasma of PVN cells in the model group,while in the EA group,only mild swelling of some mitochondria was found,being with basically normal nuclear membrane,mitochondria,rough endoplasmic reticulum,Golgi complex and ribosomes.Conclusion EA at HT7 and SP6 has a positive effect in improving insomnia and insomnia-induced fatigue in insomnia rats,which may be associated with its effects in restraining the expression of AMPK protein,and up-regulating the contents of Ac-CoA and Na~+-K~+-ATPase in PVN and CORT in plasma.
引文
[1]刘艳琴,任玉兰,赵凌,等.基于数据挖掘技术探析针刺治疗失眠症的经穴使用特点[J].中华中医药杂志,2015,30(8):2734-2737.
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[12]刘红,杨劲,唐向东.原发性失眠主客观睡眠质量与日间功能损害的关系[J].华西医学,2013,28(2):209-213.
[13]徐淼,潘霄.老年失眠症心理治疗疗效的系统评价[J].中国老年学杂志,2012,32(2):278-281.
[14]闫明启,朱立红.失眠症患者心理健康状况调查[J].中国健康心理学杂志,2013,21(10):1505-1507.
[15]毛洪祥.认知疗法治疗慢性失眠症的对照研究[J].中国健康心理学杂志,2010,18(6):655-656.
[16]王琳,向静,付华斌,等.慢性失眠患者不同负性生活事件后的睡眠质量分析[J].重庆医学,2009,38(10):1155-1157.
[17]马捷,李峰,宋月晗,等.基于“失眠恶性循环”假说探讨四逆散对失眠症的干预效应[J].中国中医药信息杂志,2013,20(1):93-94.
[18]成词松,刘萍,吴绮雯,等.电针对失眠模型大鼠焦虑状态及交感-肾上腺髓质系统的影响[J].中国针灸,2015,35(9):923-926.
[19]范丹,诸毅晖,贺智倩,等.神门、三阴交治疗失眠作用探析[J].中国中医基础医学杂志,2012,18(2):205-206.
[20]吴绮雯,诸毅晖,刘萍,等.从睡眠机制探讨神门、三阴交治疗失眠的作用[J].时珍国医国药,2014,25(2):402-403.
[21]熊延路,王明星,韩勇,等.AMPK:细胞能量中枢[J].现代生物医学进展,2014,14(31):6190-6196.
[22]CHIKAHISA S,FUJIKI N,KITAOKA K,et al.Central AMPK contributes to sleep homeostasis in mice[J].Neuropharm,2009,57(4):369-374.
[2]于心同,何天峰,邴兴红,等.针灸治疗失眠临床用穴频次分析[J].针灸临床杂志,2012,28(6):72-74.
[3]陈莉,孙丽丽,王欣,等.针灸治疗失眠处方取穴现代文献研究[J].中医杂志,2012,53(12):1051-1054.
[4]王家陟,彭楚湘,罗红柳.现代针灸治疗失眠的选穴相关性研究[J].中医外治杂志,2011,20(1):60-61.
[5]潘萍.针灸临床治疗失眠选穴规律研究[J].辽宁中医杂志,2009,36(5):818-820.
[6]严兴科,张燕,于璐,等.“镇静安神”针法对心脾两虚型失眠患者匹兹堡睡眠指数的影响[J].针刺研究,2010,35(3):222-225.
[7]李佳,刘娇萍.针刺“神门”穴对睡眠剥夺大鼠脑电波及其认知能力的影响[J].针刺研究,2017,42(6):502-506.
[8]魏歆然,魏高文,郑雪娜,等.不同经穴组合针刺对失眠大鼠下丘脑生物钟基因Clock和Bmal 1表达的影响[J].针刺研究,2017,42(5):429-433.
[9]周艳丽,高希言,王培育,等.针刺不同腧穴对失眠大鼠下丘脑γ-氨基丁酸和γ-氨基丁酸A受体的影响[J].针刺研究,2012,37(4):302-307.
[10]FLIGHT M H.Rise and shine(we are all morning people!)[J].Nat Rev Neurosci,2008,9(3):166-167.
[11]DWORAK M,MCCARLEY R W,KIM T,et al.Sleep and brain energy levels:ATP changes during sleep[J].J Neurosci,2010,30(26):9007-9016.
[12]刘红,杨劲,唐向东.原发性失眠主客观睡眠质量与日间功能损害的关系[J].华西医学,2013,28(2):209-213.
[13]徐淼,潘霄.老年失眠症心理治疗疗效的系统评价[J].中国老年学杂志,2012,32(2):278-281.
[14]闫明启,朱立红.失眠症患者心理健康状况调查[J].中国健康心理学杂志,2013,21(10):1505-1507.
[15]毛洪祥.认知疗法治疗慢性失眠症的对照研究[J].中国健康心理学杂志,2010,18(6):655-656.
[16]王琳,向静,付华斌,等.慢性失眠患者不同负性生活事件后的睡眠质量分析[J].重庆医学,2009,38(10):1155-1157.
[17]马捷,李峰,宋月晗,等.基于“失眠恶性循环”假说探讨四逆散对失眠症的干预效应[J].中国中医药信息杂志,2013,20(1):93-94.
[18]成词松,刘萍,吴绮雯,等.电针对失眠模型大鼠焦虑状态及交感-肾上腺髓质系统的影响[J].中国针灸,2015,35(9):923-926.
[19]范丹,诸毅晖,贺智倩,等.神门、三阴交治疗失眠作用探析[J].中国中医基础医学杂志,2012,18(2):205-206.
[20]吴绮雯,诸毅晖,刘萍,等.从睡眠机制探讨神门、三阴交治疗失眠的作用[J].时珍国医国药,2014,25(2):402-403.
[21]熊延路,王明星,韩勇,等.AMPK:细胞能量中枢[J].现代生物医学进展,2014,14(31):6190-6196.
[22]CHIKAHISA S,FUJIKI N,KITAOKA K,et al.Central AMPK contributes to sleep homeostasis in mice[J].Neuropharm,2009,57(4):369-374.