The Study Of Correlation Between Intra-dorsal Raphe Nucleus Injection Of5-HT_1A Affects Sleep Substances With The Yin-yang Sleeping And Waking Theory

Background and Aim:Sleep is a fundamental human physiological activity, but the mechanism is still unclear.By neural chemistry, pharmacology, physiology, pathology and a series of research suggest that sleeping-waking cycle with sleep substances change in the brain, including excitatory and inhibitory substances. The ancient books of traditional Chinese medicine have recorded many of sleep.《NeiJing》 with yin-yang theory as the platform, laid the foundation of traditional Chinese medicine in sleep, then many ancient physicians have made a point on the Yin and Yang theory, from the various aspects to explain the sleep.Combination of traditional Chinese medicine and western medicine, we could know the occurrence of sleep is the increase of the "Yin", and the waking is the increase of the "Yang".There fore, the purpose of this study is to observe how the GLU,GABA and GABAb receptors change in hypothalamus after intra-dorsal raphe nucleus injection of the5-HT1A receptor agonists8-OH-DPAT and the blockers WAY100635,based on The yin-yang sleeping and waking theory by Yougui Wan and Zuogui Wan. We’d like to explore the possibility of changes in the brain, in order to establish the physiology foundation of The yin-yang sleeping and waking theory.Methods:Pathogen-free adult male Sprague-Dawley rat(260g-320g) were randomized into7groups,blank control group(n=5),8-OH-DPAT group(n=5),8-OH-DPAT+Yougui Wan lavage group(n=5),8-OH-DPAT+saline lavage group(n=5),WAY100635group(n=5), WAY100635+Zuogui Wan lavage group (n=5), WAY100635+saline lavage group(n=5).All of the rats were anesthetized with sodium pentobarbital (35mg/KG,ip)o Using brain stereotaxic instrument for positioning:(-7.9mm,0.0mm,-6.4mm),a guide cannula was inserted and maintained in the DRN, fixed in the skull。The animals have a rest for7days after the surgery.Drug was injected into the DRN with an injection cannula, including the5-HT1A receptor agonists8-OH-DPAT(100ng,0.4μ1) and the blockers WAY100635(100ng,0.4μ1),also the saline(0.4μ1).Injection for3days,then beheaded kill on the ice.Keep the hypothalamus for testing and the brain stem for positioning. According to the result of substance changes in the rats to choose traditional Chinese medicine lavage to set up8-OH-DPAT+Yougui Wan lavage group(n=5),8-OH-DPAT+saline lavage group(n=5), WAY100635+Zuogui Wan lavage group(n=5) and WAY100635+saline lavage group (n=5).Traditional Chinese medicine lavage for7days,beheaded kill on the ice.Also keep the hypothalamus for testing and the brain stem for positioning.Frozen section technique are used to observe brain stem stereotaxic injection way;High performance liquid chromatography detect of the content of GLU and GAB A in the hypothalamus;Western Blotting detect of the GABAB receptors expression in hypothalamus.Result:(1)8-OH-DPAT group compared with the blank control group, the content of GLU is decreased, and the GABA is increased(p<0.05), inhibitory substances predominate in the hypothalamus; Compared with the blank control group, the content of GLU in WAY100635group is increased,GABA content is decreased(p<0.05), the dominant excitatory substances in the hypothalamus;(2) WAY100635+Zuogui Wan lavage group compared with saline lavage group,the content of GLU is decreased,GABA is increased (p<0.05);8-OH-DPAT+Yougui Wan lavage group rats compared with saline lavage group, the content of GLU is increased, GABA is decreased(p<0.05);(3) compared with the blank control group, the amount of GABABR1a and GABAeR1b expression in WAY100635group were decreased (p<0.05);The GABAeR1a and GABAeR1b expression in8-OH-DPAT group were increased (p<0.05);(4) WAY100635+Zuogui Wan lavage group,8-OH-DPAT+Yougui Wan lavage group compared with normal saline lavage group the amount of GABABR1a and GABABR1b expression did not see obviously difference (p>0.05),Chinese medicine what we chosen in the experiment haven’t a significant effect on GABAB.Conclusion:①Intra-dorsal raphe nucleus injection of5-HT1A receptor agonists8-OH-DPAT induced the hypothalamus inhibitory neurotransmitter GABA increasing and GLU decreasing.At the same time,intra-dorsal raphe nucleus injection of5-HT1A receptor blockers WAY100635had the opposit result;②Yougui Wan reduces the content of inhibitory substance,Zuogui Wan reduces the contents of inhibitory substance, eventually keep the balance.Yin-Yang sleeping and waking theory to guide the Chinese medicine treatment of sleep disorders has certainly effect;③GABAB receptor may have certain regulation of sleep, but the target of Chinese medicine is unclear yet.