针刺跷脉对失眠大鼠自发活动昼夜节律及海马内5-HT1A、5-HT2A受体mRNA及蛋白表达的影响

发布时间：2018-04-11 13:11

本文选题：失眠 + 针刺　；参考：《成都中医药大学》2016年硕士论文

【摘要】：目的：借助PCPA失眠动物模型,观察正常和失眠SD大鼠自发活动昼夜节律特征,以及针刺跷脉后自发活动昼夜节律的调整和变化；观察失眠大鼠海马内5-HT1A、5-HT2A受体mRNA及蛋白表达的改变,评价针刺跷脉对失眠大鼠的治疗作用,以探讨其内在作用机制,为针刺跷脉治疗失眠提供科学依据。方法：1.将30只雄性SD大鼠驯化与L：D(12：12)光暗同步后,利用动物自发活动自动记录及节律分析系统和美国CLOCKLAB数据采集分析系统,自动检测和记录自发活动昼夜节律。然后将SD大鼠随机分为空白组、模型组、针刺组,每组10只。2.对模型组和针刺组SD大鼠,运用腹腔注射对氯苯丙氨酸(PCPA)混悬液的方法,复制失眠模型。观察造模前后自发活动昼夜节律的改变。3.造模后,每日早上8：00-10：00,对空白组及模型组给予抓放固定、不针刺干预,对针刺组给予抓放固定、针刺“申脉”、“照海”干预,1次／天,20min/次,连续1周,观察针刺跷脉对自发活动昼夜节律的影响。4.应用实时荧光定量PCR(RT-PCR)检测海马内5-HT1A、5-HT2A受体mRNA的表达水平。5.应用蛋白免疫印迹(Western Blot)检测海马内5-HT1A、5-HT2A受体蛋白的表达水平。结果：1.在L：D(12：12)驯化稳定后,SD大鼠自发活动均表现出近似昼夜节律。模型组与针刺组在PCPA造模后,表现出明显的自发活动昼夜节律紊乱,休息期活动量明显增加,与空白组比较有统计学意义(P0.05)。2.针刺跷脉对针刺组大鼠自发活动昼夜节律具有调整作用,休息期活动量减少,与模型组比较,有统计学意义(P0.05)。3.造模后,模型组大鼠海马中5-HT1A受体mRNA相对表达值减少,5-HT2A受体mRNA相对表达值增加,与空白组比较,有统计学意义(P0.05)。4.针刺治疗后,针刺组大鼠海马中5-HT1A受体mRNA相对表达值增加,5-HT2A受体mRNA相对表达值减少,与模型组比较,有统计学意义(P0.05)。5.造模后,模型组大鼠海马中5-HT1A受体蛋白的表达水平减少,5-HT2A受体蛋白的表达水平增加,与空白组比较,有统计学意义(P0.05)。6.针刺治疗后,针刺组大鼠海马中5-HT1 A受体蛋白的表达水平增加,5-HT2A受体蛋白的表达水平减少,与模型组比较,有统计学意义(P0.05)。结论：1.PCPA造模后失眠SD大鼠一般状态表现异常,自发活动昼夜节律紊乱,休息期活动量明显增加。2.针刺跷脉可以使失眠SD大鼠自发活动昼夜节律恢复其规律性,休息期活动量相对减少,睡眠逐渐恢复正常。3.针刺跷脉可以使失眠SD大鼠海马中5-HT1A受体mRNA及蛋白表达增加,5-HT2A受体mRNA及蛋白表达减少。这可能是针刺跷脉治疗失眠的内在作用机制之一。
[Abstract]:Objective: to observe the circadian rhythm of spontaneous activity in normal and insomnia SD rats, and to observe the regulation and change of the circadian rhythm of spontaneous activity after acupuncture.To observe the changes of mRNA and protein expression of 5-HT _ (1) A _ (5-HT _ 2A) receptor in hippocampus of insomnia rats, to evaluate the therapeutic effect of acupuncture on stilt veins in insomnia rats, to explore its intrinsic mechanism, and to provide scientific basis for acupuncture of stilt-vein acupuncture in the treatment of insomnia.Method 1: 1.30 male Sprague-Dawley rats were acclimated to L: D12: 12) light and dark synchronization. The spontaneous activity circadian rhythm was automatically detected and recorded by the automatic recording and rhythm analysis system of animal spontaneous activity and the CLOCKLAB data acquisition and analysis system of the United States.Then SD rats were randomly divided into blank group, model group, acupuncture group, 10 rats in each group.Models of insomnia were established in SD rats in model group and acupuncture group by intraperitoneal injection of PCPA suspension.Observe the changes of circadian rhythm of spontaneous activity before and after modeling.After making the model, 8: 00-10: 00 every morning, the blank group and the model group were given grip fixation without acupuncture intervention, and the acupuncture group was given grasping fixation, acupuncture "Shenmai" and "Zhaohai" intervention once a day for 20 minutes every time for one week in a row.To observe the effect of acupuncture on the circadian rhythm of spontaneous activity.The expression level of 5-HT _ 1A _ (5-HT _ 2A) receptor mRNA in hippocampus was detected by real-time quantitative PCR- RT-PCR- PCR.Western blot was used to detect the expression of 5-HT _ 1A _ (5-HT _ 2A) receptor protein in hippocampus.The result is 1: 1.The spontaneous activity of SD rats showed an approximate circadian rhythm after acclimation and stabilization.After PCPA, the model group and acupuncture group showed obvious disturbance of the circadian rhythm of spontaneous activity and increased the amount of activity during rest period, which had statistical significance compared with the blank group (P0.05. 2).Acupuncture of still-vein can adjust the circadian rhythm of spontaneous activity of rats in acupuncture group and decrease the amount of activity in rest period. Compared with the model group, it has statistical significance (P0.05 路3).After modeling, the relative expression of 5-HT1A receptor mRNA in the hippocampus of the model group decreased and the relative expression value of 5-HT2A receptor mRNA increased. Compared with the blank group, the expression of 5-HT1A receptor mRNA in the model group was significantly higher than that in the blank group.After acupuncture treatment, the relative expression of 5-HT1A receptor mRNA in hippocampus of the acupuncture group increased and the relative expression value of 5-HT2A receptor mRNA decreased. Compared with the model group, the relative expression of 5-HT1A receptor mRNA was significantly higher than that of the model group.After modeling, the expression of 5-HT1A receptor protein in the hippocampus of the model group decreased and the expression level of 5-HT2A receptor protein increased. Compared with the blank group, the expression level of 5-HT1A receptor protein in the model group was significantly higher than that in the blank group.After acupuncture treatment, the expression level of 5-HT1 A receptor protein in hippocampus of the acupuncture group increased and the expression level of 5-HT2A receptor protein decreased. Compared with the model group, the expression level of 5-HT1 A receptor protein was significantly higher than that of the model group (P 0.05).Conclusion 1. The normal state of insomnia SD rats was abnormal, the circadian rhythm of spontaneous activity was disordered, and the amount of activity during rest period was increased significantly.Acupuncture can restore the circadian rhythm of spontaneous activity of insomnia SD rats, decrease the amount of activity during rest period, and gradually return to normal. 3.Acupuncture could increase the expression of 5-HT1A receptor mRNA and protein in hippocampus of SD rats and decrease the expression of 5-HT 2A receptor mRNA and protein.This may be one of the internal mechanisms of acupuncture on the treatment of insomnia.
【学位授予单位】：成都中医药大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R245

【参考文献】