蒙医温针改善睡眠作用机制的实验研究
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摘要
失眠是指睡眠的始发和睡眠的维持发生障碍致使睡眠缺失,不能满足机体的生理需求引发的一系列精神症状,属蒙医“赫依”病范畴。西医治疗本病仍存在难以根治、不同程度地产生不良反应、耐药性和依赖性。蒙医温针对该病治疗通过多层次、多途径、多手段结合调整机体而达到改善睡眠的目的,其最大优势在于可以避免口服药物所致的副作用。因此,寻找适当有效的针刺干预措施是临床与实验研究的重要课题之一
蒙医温针治疗具有镇赫依、疏通气血、调体素的作用。近年来研究表明,温针疗法影响神经内分泌免疫网络中神经递质和细胞因子的作用和抗疲劳的作用。
研究目的本课题从实验方面,观察蒙医温针对戊巴比妥钠致大鼠睡眠时间的影响,探讨蒙医温针的改善睡眠作用。以对氯苯丙氨酸致失眠模型大鼠为研究对象,观察模型大鼠的行为学变化,以脑内单胺类类神经递质5-HT、5-HIAA及细胞因子IL-1β、IL-2、IL-6、TNF-a为主要指标,初步研究蒙医温针干预对睡眠的可能机制,以期为后续研究做基础。
研究方法
1.选用戊巴比妥钠诱导的睡眠的方法,将健康雄性SD大鼠30只分为空白组和蒙医温针组,每组15只。2.温针组采用温针疗法,选择顶穴、赫依穴、心穴,采用戊巴比妥钠致睡眠时间为观察指标。3.采用对氯苯丙氨酸致失眠大鼠模型,将健康雄性SD大鼠60只分为空白对照组、失眠模型组、蒙医温针治疗组和地西泮注射液组,每组15只。4.采用行为学指标观察大鼠的自主活动和一般情况。5.采用高效液相色谱法检测各组大鼠脑内5-HT、5-HIAA含量变化。6.采用ELISA法检查各组大鼠脑内细胞因子IL-1β、IL-2、IL-6、TNF-a含量变化。
研究结果
1.睡眠时间：蒙医温针组与空白组比较睡眠时间延长,有明显差异(P<0.01)；蒙医温针对大鼠入睡率无作用,与空白对照组比较无显著性差异(P>0.05),对睡眠潜伏期无明显作用,与空白对照组比较无显著性差异(P>0.05)
2.体重变化：失眠第三天和第七天,与模型组比较,蒙医温针组大鼠体重减轻量有统计学差异(P<0.05)；与其余三组体重量呈现减少的表现相反,空白对照组大鼠体重量呈现增加的表现。
3.行为学：在水平活动上,各组大鼠失眠第七天,模型组在活动时间上明显高于空白对照组(P<0.01)；模型组在中央格停留时间高于空白对照组,但无明显差异(P>0.05)；蒙医温针组大鼠在中央停留低于模型组,但无明显差异(P>0.05)。在垂直活动上,模型组站立次数明显高于对照组,有明显差异(P<0.05)；蒙医温针组明显低于模型组(P<0.05)。
4.5-HT、5-HIAA含量：蒙医温针组对大鼠下丘脑、海马、脑干、大脑前额皮层5-HT.5-HIAA含量的影响研究结果表明,蒙医温针能显著增加失眠大鼠下丘脑、海马5-HT、5-HIAA含量,与模型组比较有显著性差异(P<0.05)；对大脑前额皮层和脑干5-HT、5-HIAA含量略有增加,但与模型组比较无显著性差异(P<0.05)
5.IL-1β含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层IL-1β含量的影响研究结果表明,蒙医温针能显著降低下丘脑、大脑前额皮层异常增加的IL-1β含量,与模型组比较有显著差异(P<0.05,P<0.01)；对海马IL-1β含量略有降低作用,但与模型组无明显差异(P>0.05)；对脑干IL-1β含量无影响,与模型组比较无明显差异(P>0.05)。
IL-2含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层IL-2含量的影响研究结果表明,蒙医温针能显著降低下丘脑、海马、大脑前额皮层异常增加的IL-2含量,与模型组比较有显著差异(P<0.05,P<0.05,P<0.01)；对脑干IL-2含量无影响,与模型组比较无明显差异(P>0.05)。
IL-6含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层IL-6含量的影响研究结果表明,蒙医温针能显著降低下丘脑、海马、大脑前额皮层异常增加的IL-6含量,与模型组比较有显著差异(P<0.05,P<0.01,P<0.01)；对脑干IL-6含量无影响,与模型组比较无明显差异(P>0.05)。
TNF-a含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层TNF-a含量的影响研究结果表明,蒙医温针能显著降低下丘脑异常增加的TNF-a含量,与模型组比较有显著差异(P<0.05)；对海马、大脑皮层、脑干TNF-a含量无影响,与模型组比较无明显差异(P>0.05,P>0.05,P>0.05)。
结论
1.蒙医温针具有改善睡眠的作用,明显延长戊巴比妥钠诱导的睡眠时间。
2.蒙医温针显著改善失眠大鼠的精神状态,改善其行为能力。
3.蒙医温针具有改善睡眠的作用机制可能有5-HT能神经系统、细胞因子(IL-1β、IL-2、IL-6、TNF-a)介导。
4.蒙医温针对失眠大鼠下丘脑、海马、大脑皮层的影响为明显,特别是对下丘脑和海马5-HT的含量和下丘脑、大脑皮层的IL-1、IL-2、IL-6的含量影响明显。
5.蒙医温针操作简便、安全、有效、副作用小。
Insomnia is a series of psychiatric symptoms that a sleep onset and sleep maintenance disorder result in lack of sleep and can not meet the physiological needs of the organism. In Mongolian Medicine,it belongs to the Mongolian "Heyi" disease category.Using the western therapy still remain problems which are hardly to cure and different degrees of adverse reactions,drug resistance and dependence.The Mongolian Medicine Warm Acupuncture treatment can imprve the qulity of sleep through multi-level and various ways and methods. The most advantage is avoiding the side effects by the oral medication.Therefore,searching a proper,effctive acupuncture method is one of the key projects.
Mongolian Medicine Warm Acupuncture can adjust unbalanced Heyi,can improve the flow of Heyi and Blood and can restore the harmonious blance of Mahbud. According to the recent study,the Mongolian Medicine Warm Acupucture can affect certain neurotransmitter levels and cytokin levels.
Objective
From experiment research,to observe the influence of the sleeping time of rats induced by sodium pentobarbital and explicit that Mongolian Medicine Warm Acupuncture improving effect the sleep. Use the chronic intraperitoneal injection of amphetamine acid(PCPA) animal model of insomnia as the objects, put warm acupuncture on "Ori"."Heyi ","Zurih" acupuncture point and choose capability of behavioral changes of each group, changes of content of5-HT,5-HIAA,IL-1β,IL-2,IL-6, TNF-a as indicators. The object of this study is to explore the mechanism of using Mongolian Medicine Warm Acupuncture to cure insomnia.
Methods
①We chose30healthy male SD rats and random divided them into nonnal guoup and Mongolian Medicine Warm Acupuncture treatment group and15rats for each group, then treated the cure group with Mongolian Medicine Warm Acupuncture for7days. After that, observed the sleep time.②We chose60healthy male SD rats and random divided them into normal guoup, insomnia group, Mongolian Medicine Warm Acupuncture treatment group and diazepam injection group and15rats for each group. Used the chronic intraperitoneal injection of amphetamine acid(PCPA) animal model of insomnia, and then treated the cure groups with Mongolian Medicine Warm Acupuncture for7days.④Observe the rat's jeneral status and free activities through open field test.⑤According to the HPLC method to measure the content of5-HT.5-HIAA in the cerebrum.⑥Using the ELISA method to measue the content of L-1β,IL-2,IL-6,TNF-a.
Result
1. Sleep time:Mongolian Medicine Warm Acupuncture treatment group can prolonge the pentobarbital sodium induced sleep time,compared with the normal group there were significant differences(P<0.01).
2. Changes of body weight:In the third days and seventh days of insomnia, compared with the model group, Changes of body weight of Mongolian Medicine Warm Acupuncture treatment group were significant differences(P<0.05);compared with other groups, body weight of the rat of the nomal group.
3. Behavioral changes:In open field test, the time of movement of rat of model group higher than normal group with highly significant differences(P<0.01);compared with model group, Mongolian Medicine Warm Acupuncture treatment group were significant differences (P<0.05).
The time of inner zone of rat of model group more than control group> but there were not significant differences (P>0.05);compared with model group, Mongolian Medicine Warm Acupuncture treatment group were not significant differences (P>0.05).
rats of model group made significantly more than normal group in stand-up times; Mongolian Medicine Warm Acupuncture treatment group less than model group in stand-up times(P<0.05).
4.5-HT,5-HIAA content:From the study on the influence on5-HT,5-HIAA content of hypothalamus, hippocampus, pallium, brianstem showed that it can significantly increase the5-HT,5-HIAA content of hypothalamus, hippocampus of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05); it can not significantly increase the5-HT,5-HIAA of pallium, brianstem (P>0.05).
5. IL-1β content:From the study on the influence on IL-1β content of hypothalamus, hippocampus, pallium,brianstem showed that it can significantly reduce the IL-1β content of hypothalamus, pallium of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05); there is a little reduce the IL-1β of hippocampus, but there is not significantly different compared with that of model group (P>0.05).it not influence the IL-1β of brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
IL-2content:From the study on the influence on IL-2content of hypothalamus. hippocampus, pallium, brianstem showed that it can significantly reduce the IL-2content of hypothalamus、hippocampus、 pallium of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05, P<0.05, P<0.01);there is no influence the IL-2of brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
IL-6content:From the study on the influence on IL-6Content of hypothalamus, hippocampus, pallium, brianstem showed that it can significantly reduce the IL-6content of hypothalamus, hippocampus, pallium of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group (P<0.05, P<0.01, P<0.01); there is no influence the IL-6of brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
TNF-a content:From the study on the influence on IL-6Content of hypothalamus、 hippocampus、 pallium、 brianstem showed that it can significantly reduce the IL-6content of hypothalamus of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05);there is no influence the IL-6of hippocampus、 pallium、 brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
Conclusion
1. Mongolian Medicine Warm Acupuncture can improve sleep, can prolong the sleeping time of rats indued by sodium pentobarbital.
2. Mongolian Medicine Warm Acupuncture can reduce the spontaneous activity, improve behavioral effects as reaction.
3.5-HT,L-1β,IL-2,IL-6,TNF-a participate in improving sleep of Mongolian Medicine Warm Acupuncture.
4. Effect of Mongolian Medicine warm acupuncture on the hypothalamus, hippocampus, cerebral cortex of insomnia rats was significant. Mongolian Medicine Warm Acupuncture could improve rat's sleep,which might be possibly achieved through changing the5-HT content of hypothalamus, hippocampus and IL-1, IL-2, IL-6content of hypothalamus、 hippocampus and pallium.
5.Mongolian Medicine Warm Acupunture with the advangates of simple oeration, safety,effective,few side effects.
蒙医温针治疗具有镇赫依、疏通气血、调体素的作用。近年来研究表明,温针疗法影响神经内分泌免疫网络中神经递质和细胞因子的作用和抗疲劳的作用。
研究目的本课题从实验方面,观察蒙医温针对戊巴比妥钠致大鼠睡眠时间的影响,探讨蒙医温针的改善睡眠作用。以对氯苯丙氨酸致失眠模型大鼠为研究对象,观察模型大鼠的行为学变化,以脑内单胺类类神经递质5-HT、5-HIAA及细胞因子IL-1β、IL-2、IL-6、TNF-a为主要指标,初步研究蒙医温针干预对睡眠的可能机制,以期为后续研究做基础。
研究方法
1.选用戊巴比妥钠诱导的睡眠的方法,将健康雄性SD大鼠30只分为空白组和蒙医温针组,每组15只。2.温针组采用温针疗法,选择顶穴、赫依穴、心穴,采用戊巴比妥钠致睡眠时间为观察指标。3.采用对氯苯丙氨酸致失眠大鼠模型,将健康雄性SD大鼠60只分为空白对照组、失眠模型组、蒙医温针治疗组和地西泮注射液组,每组15只。4.采用行为学指标观察大鼠的自主活动和一般情况。5.采用高效液相色谱法检测各组大鼠脑内5-HT、5-HIAA含量变化。6.采用ELISA法检查各组大鼠脑内细胞因子IL-1β、IL-2、IL-6、TNF-a含量变化。
研究结果
1.睡眠时间：蒙医温针组与空白组比较睡眠时间延长,有明显差异(P<0.01)；蒙医温针对大鼠入睡率无作用,与空白对照组比较无显著性差异(P>0.05),对睡眠潜伏期无明显作用,与空白对照组比较无显著性差异(P>0.05)
2.体重变化：失眠第三天和第七天,与模型组比较,蒙医温针组大鼠体重减轻量有统计学差异(P<0.05)；与其余三组体重量呈现减少的表现相反,空白对照组大鼠体重量呈现增加的表现。
3.行为学：在水平活动上,各组大鼠失眠第七天,模型组在活动时间上明显高于空白对照组(P<0.01)；模型组在中央格停留时间高于空白对照组,但无明显差异(P>0.05)；蒙医温针组大鼠在中央停留低于模型组,但无明显差异(P>0.05)。在垂直活动上,模型组站立次数明显高于对照组,有明显差异(P<0.05)；蒙医温针组明显低于模型组(P<0.05)。
4.5-HT、5-HIAA含量：蒙医温针组对大鼠下丘脑、海马、脑干、大脑前额皮层5-HT.5-HIAA含量的影响研究结果表明,蒙医温针能显著增加失眠大鼠下丘脑、海马5-HT、5-HIAA含量,与模型组比较有显著性差异(P<0.05)；对大脑前额皮层和脑干5-HT、5-HIAA含量略有增加,但与模型组比较无显著性差异(P<0.05)
5.IL-1β含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层IL-1β含量的影响研究结果表明,蒙医温针能显著降低下丘脑、大脑前额皮层异常增加的IL-1β含量,与模型组比较有显著差异(P<0.05,P<0.01)；对海马IL-1β含量略有降低作用,但与模型组无明显差异(P>0.05)；对脑干IL-1β含量无影响,与模型组比较无明显差异(P>0.05)。
IL-2含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层IL-2含量的影响研究结果表明,蒙医温针能显著降低下丘脑、海马、大脑前额皮层异常增加的IL-2含量,与模型组比较有显著差异(P<0.05,P<0.05,P<0.01)；对脑干IL-2含量无影响,与模型组比较无明显差异(P>0.05)。
IL-6含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层IL-6含量的影响研究结果表明,蒙医温针能显著降低下丘脑、海马、大脑前额皮层异常增加的IL-6含量,与模型组比较有显著差异(P<0.05,P<0.01,P<0.01)；对脑干IL-6含量无影响,与模型组比较无明显差异(P>0.05)。
TNF-a含量：蒙医温针对大鼠下丘脑、海马、脑干、大脑前额皮层TNF-a含量的影响研究结果表明,蒙医温针能显著降低下丘脑异常增加的TNF-a含量,与模型组比较有显著差异(P<0.05)；对海马、大脑皮层、脑干TNF-a含量无影响,与模型组比较无明显差异(P>0.05,P>0.05,P>0.05)。
结论
1.蒙医温针具有改善睡眠的作用,明显延长戊巴比妥钠诱导的睡眠时间。
2.蒙医温针显著改善失眠大鼠的精神状态,改善其行为能力。
3.蒙医温针具有改善睡眠的作用机制可能有5-HT能神经系统、细胞因子(IL-1β、IL-2、IL-6、TNF-a)介导。
4.蒙医温针对失眠大鼠下丘脑、海马、大脑皮层的影响为明显,特别是对下丘脑和海马5-HT的含量和下丘脑、大脑皮层的IL-1、IL-2、IL-6的含量影响明显。
5.蒙医温针操作简便、安全、有效、副作用小。
Insomnia is a series of psychiatric symptoms that a sleep onset and sleep maintenance disorder result in lack of sleep and can not meet the physiological needs of the organism. In Mongolian Medicine,it belongs to the Mongolian "Heyi" disease category.Using the western therapy still remain problems which are hardly to cure and different degrees of adverse reactions,drug resistance and dependence.The Mongolian Medicine Warm Acupuncture treatment can imprve the qulity of sleep through multi-level and various ways and methods. The most advantage is avoiding the side effects by the oral medication.Therefore,searching a proper,effctive acupuncture method is one of the key projects.
Mongolian Medicine Warm Acupuncture can adjust unbalanced Heyi,can improve the flow of Heyi and Blood and can restore the harmonious blance of Mahbud. According to the recent study,the Mongolian Medicine Warm Acupucture can affect certain neurotransmitter levels and cytokin levels.
Objective
From experiment research,to observe the influence of the sleeping time of rats induced by sodium pentobarbital and explicit that Mongolian Medicine Warm Acupuncture improving effect the sleep. Use the chronic intraperitoneal injection of amphetamine acid(PCPA) animal model of insomnia as the objects, put warm acupuncture on "Ori"."Heyi ","Zurih" acupuncture point and choose capability of behavioral changes of each group, changes of content of5-HT,5-HIAA,IL-1β,IL-2,IL-6, TNF-a as indicators. The object of this study is to explore the mechanism of using Mongolian Medicine Warm Acupuncture to cure insomnia.
Methods
①We chose30healthy male SD rats and random divided them into nonnal guoup and Mongolian Medicine Warm Acupuncture treatment group and15rats for each group, then treated the cure group with Mongolian Medicine Warm Acupuncture for7days. After that, observed the sleep time.②We chose60healthy male SD rats and random divided them into normal guoup, insomnia group, Mongolian Medicine Warm Acupuncture treatment group and diazepam injection group and15rats for each group. Used the chronic intraperitoneal injection of amphetamine acid(PCPA) animal model of insomnia, and then treated the cure groups with Mongolian Medicine Warm Acupuncture for7days.④Observe the rat's jeneral status and free activities through open field test.⑤According to the HPLC method to measure the content of5-HT.5-HIAA in the cerebrum.⑥Using the ELISA method to measue the content of L-1β,IL-2,IL-6,TNF-a.
Result
1. Sleep time:Mongolian Medicine Warm Acupuncture treatment group can prolonge the pentobarbital sodium induced sleep time,compared with the normal group there were significant differences(P<0.01).
2. Changes of body weight:In the third days and seventh days of insomnia, compared with the model group, Changes of body weight of Mongolian Medicine Warm Acupuncture treatment group were significant differences(P<0.05);compared with other groups, body weight of the rat of the nomal group.
3. Behavioral changes:In open field test, the time of movement of rat of model group higher than normal group with highly significant differences(P<0.01);compared with model group, Mongolian Medicine Warm Acupuncture treatment group were significant differences (P<0.05).
The time of inner zone of rat of model group more than control group> but there were not significant differences (P>0.05);compared with model group, Mongolian Medicine Warm Acupuncture treatment group were not significant differences (P>0.05).
rats of model group made significantly more than normal group in stand-up times; Mongolian Medicine Warm Acupuncture treatment group less than model group in stand-up times(P<0.05).
4.5-HT,5-HIAA content:From the study on the influence on5-HT,5-HIAA content of hypothalamus, hippocampus, pallium, brianstem showed that it can significantly increase the5-HT,5-HIAA content of hypothalamus, hippocampus of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05); it can not significantly increase the5-HT,5-HIAA of pallium, brianstem (P>0.05).
5. IL-1β content:From the study on the influence on IL-1β content of hypothalamus, hippocampus, pallium,brianstem showed that it can significantly reduce the IL-1β content of hypothalamus, pallium of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05); there is a little reduce the IL-1β of hippocampus, but there is not significantly different compared with that of model group (P>0.05).it not influence the IL-1β of brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
IL-2content:From the study on the influence on IL-2content of hypothalamus. hippocampus, pallium, brianstem showed that it can significantly reduce the IL-2content of hypothalamus、hippocampus、 pallium of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05, P<0.05, P<0.01);there is no influence the IL-2of brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
IL-6content:From the study on the influence on IL-6Content of hypothalamus, hippocampus, pallium, brianstem showed that it can significantly reduce the IL-6content of hypothalamus, hippocampus, pallium of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group (P<0.05, P<0.01, P<0.01); there is no influence the IL-6of brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
TNF-a content:From the study on the influence on IL-6Content of hypothalamus、 hippocampus、 pallium、 brianstem showed that it can significantly reduce the IL-6content of hypothalamus of rats of Mongolian Medicine Warm Acupuncture treatment group. Which were significantly different than that of model group(P<0.05);there is no influence the IL-6of hippocampus、 pallium、 brianstem of rats, there is not significantly different compared with that of model group (P>0.05).
Conclusion
1. Mongolian Medicine Warm Acupuncture can improve sleep, can prolong the sleeping time of rats indued by sodium pentobarbital.
2. Mongolian Medicine Warm Acupuncture can reduce the spontaneous activity, improve behavioral effects as reaction.
3.5-HT,L-1β,IL-2,IL-6,TNF-a participate in improving sleep of Mongolian Medicine Warm Acupuncture.
4. Effect of Mongolian Medicine warm acupuncture on the hypothalamus, hippocampus, cerebral cortex of insomnia rats was significant. Mongolian Medicine Warm Acupuncture could improve rat's sleep,which might be possibly achieved through changing the5-HT content of hypothalamus, hippocampus and IL-1, IL-2, IL-6content of hypothalamus、 hippocampus and pallium.
5.Mongolian Medicine Warm Acupunture with the advangates of simple oeration, safety,effective,few side effects.
引文
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