交泰丸治疗心肾不交型失眠的疗效特点解析和诱导睡眠的机制研究

Jiaotaiwan has definite efficacy on insomnia of non-interaction between heart and kidney.However the therapeutic effect is not clear,the advantage is not outstanding, and it is also lack of discussion of related neural mechanism.Combined PSG with MRS to test sleeping time,quality and variation of GABA in brain after englobement of Jiaotaiwan in patients with insomnia of non-interaction between heart and kidney.HPLC and ELISA detect contents of adenosine and melatonin.Use PCPA method to establish insomnia rat model of non-interaction between heart and kidney,and controled with zolpidem,sleeping biological system recorded and compared in 72h as follows:the amount and state change of wake,NREM and REM;to analyze characters of influence on sleep time and quality.Isolate hippocampus,thalamus,brainstem and cortex.UHPLC-MS/MS examine the quantities of adenosine and melatonin.Real-time PCR and Western bolt test contents of A1R,A2AR,MT1,MT2,mRNA and protein.MRS detect contents of GABA and Cr.Combined with clinic and animal study,we aim to clear feature of remission of kinds of symptoms of insomnia of non-interaction between heart and kidney,so we can definite advantage efficacy of improving difficulty falling asleep.Based on micro transmitter and macro activity,and from gene,protein to metabonomics,we explored the possible mechanism of sleeping inducement of Jiaotaiwan in homeostasis,master clock and arousal inhibition and correlation of brain areas.

交泰丸对心肾不交型失眠疗效确切，但疗效特点不明晰，优势不突出，也缺乏相关神经机制的探讨。联合PSG和MRS，检测心肾不交型失眠患者口服交泰丸后睡眠时间和质量及脑内GABA的变化，HPLC和ELISA检测血中腺苷和褪黑素的含量。PCPA法建立心肾不交型失眠大鼠模型，对照唑吡坦，睡眠生物系统记录72h并比较wake、NREM和REM量及状态改变，解析交泰丸影响睡眠时间和质量的特点。分离海马、丘脑、脑干、皮质，UHPLC-MS/MS检测腺苷和褪黑素含量，Real-time PCR和Western bolt检测A1R、A2AR、MT1R、MT2R mRNA和蛋白含量，MRS检测GABA和Cr含量。临床与动物研究结合，明确交泰丸缓解心肾不交型失眠各症状的特点，确定改善入睡困难的优势疗效。微观递质和宏观活动结合，从基因、蛋白和代谢组，阐明交泰丸在内稳态、主时钟和觉醒抑制方面的睡眠诱导机制和脑区相关性。

失眠严重摧残人体身心健康。作为治疗心肾不交型失眠的经典方剂之一，交泰丸缺乏疗效特点和相关神经机制的研究。本课题组采用广泛靶向代谢组检测技术获得交泰丸的总离子电流，并鉴定25种主要化合物的特征；采用气相色谱/质谱分析并鉴定了交泰丸挥发油的50个主要成分；采用自发活动仪，比较不同剂量的交泰丸对小鼠的镇静作用，确定交泰丸最佳镇静比例为黄连：肉桂=20:2，最佳剂量为0.220 g·mL-1。上述结果不仅为交泰丸药效研究提供依据，而且为临床运用的剂量和剂型提供借鉴。睡眠解析平台分析了交泰丸改变PCPA失眠大鼠wake、NREM和REM睡眠量，昼夜不同持续时间睡眠-觉醒片段数、互相之间转换次数及脑电能量强度，明确交泰丸降低觉醒时间，增加慢波睡眠和快速动眼睡眠时间和改善睡眠结构的作用。运用高效液相色谱法和ELLISA方法分别检测大鼠血清及不同脑区腺苷、GABA和褪黑素，Western Blot和实时荧光定量PCR技术分别检测不同脑区A1R、A2AR、MTNR1B、MTNR2蛋白和mRNA含量表达，结果表明交泰丸对于A1R的调节作用强于A2AR，海马区可能是疗效优势脑区，而且交泰丸对不同脑区的褪黑素及褪黑素受体的浓度存在双向调节的可能，或者通过调节褪黑素在不同脑区的分布情况，从而调节睡眠。课题组采用随机、对照、双盲试验设计，随机纳入交泰丸组与安慰剂组共128例患者，均服药1周。基线期，两组在人口学资料和各项检测指标均无差异。治疗后，交泰丸组PSQI总分和睡眠质量、入睡时间、睡眠时间、睡眠效率等因子评分降低，左侧小脑Ins和Cho及血液褪黑素均升高。两组各项安全性指标均无差异。综合临床研究结果，交泰丸可改善心肾不交型失眠患者的睡眠质量，缩短入睡时间，可能通过小脑代谢变化调节睡眠，可能通过调节褪黑素浓度进而改善睡眠然。