Glutamate In "Brain Stem-gut Axis" Involves In Rat Restraint Water-immersion Stress

The incidence of gastrointestinal diseases is very high in China,which is prone to relapse in a long time and is difficult to cure completely.Therefore,it has always been regarded as a difficult problem by the medical profession.The gastrointestinal diseases are closely related to poor eating habits,in addition,Stress such as major surgical operations,psychological trauma and craniocerebral injury plays an important role in the progression of gastrointestinal diseases.Restraint water-immersion stress(RWIS)is considered to be acomplex and psychological stressor in rats,resulting in hypothermia and disordered gastrointestinal function in a short time,which can mimic clinical gastric ulcer caused by surgery,trauma.Therefore,it is widely used to study the pathogenesis and treatment of stress-induced gastric damage.Sensory information from the viscera,including the gastrointestinal(GI)tract,is transmitted through the afferent vagus and activate second-order the nucleus tractus solitarius(NTS)neurons via glutamate action.The NTS integrate this sensory information to regulate autonomic functions and homeostasis.The integrated response is conveyed to the preganglionic neurons of the dorsal motor nucleus of the vagus(DMV)using mainly GABA,glutamate and catecholamines as neurotransmitters.Our previous studies have shown that A large number of experimental studies our laboratory shown that the RWIS significantly increase the release of neurotransmitters such as nitric oxide,acetylcholine and catecholamines in NTS,DMV and intestinal myenteric plexus.Glutamate(Glu),the most important excitatory neurotransmitter,is involved in a variety of stresses process(such as chronic stress depression,oxidative stress injury,psychological stress,brain oxidative stress).However,whether the Glu involved in the RWIS has not been reported,if so:1.Does Glu in "brain stem-gut axis" participate in the RWIS process,and do its synthetase,transporter and receptor change significantly after RWIS?2.The effect of NTS injection of Glu receptor inhibitor on neurons and glial cells in the enteric nervous system.3.The effect of NTS injection of Glu receptor inhibitor on gastrointestinal function.To solve the three problems,we designed the following experiments:Experimental Research ?: The rats were randomly divided into three groups(RWIS 0h,1h,3h)and the expression of GLS1,VGLUT2,GLUR3,NMDAR1 and EAAT2 in brain stem-gut axis were detected by Western blot,immunohistochemical and RT-PCR techniques.Experimental Research ?: The rats were randomly divided into four groups(control group,RWIS 1h group,normal saline + RWIS 1h group,CNQX + RWIS 1h group).Immunohistochemical staining and Western Blot technology were used to detect the expression of p-Erk1/2,NOS,CHAT and GFAP in the intestinal myenteric plexus,which explore the effect of Glu on neurons and glial cells in the gastrointestinal tract.Experimental Research ?: The rats were randomly divided into four groups(control group,RWIS 1h group,normal saline + RWIS 1h group,CNQX + RWIS 1h group).Western Blot technology was used to detect the expression of occludin and claudin1 in gut,and ELISA technology was used to detect the expression of MUC5 AC in stomach,which explore the effect of Glu on gastrointestinal function in RWIS.Experimental results showed that:1.Compared with RWIS 0h group,the expression of VGLUT2,GLS1,GLUR3 and NMDAR1 in the brain stem DVC in RWIS 1h and 3h group of rats significantly increased at the mRNA and protein level.The expression of eaat2 was significantly increased at the mRNA level but significantly decreased at the protein level.This may be a compensatory response to stress-induced increase in Glu,while the expression of functional proteins is reduced,thereby increasing extracellular Glu levels.This study showed that Glu in "brain stem-gut axis" isinvolved in the RWIS process.2.Compared with the normal saline + RWIS 1h group,the expressions of GFAP,p-Erk1/2,NOS,and CHAT in the enteric nervous system of the CNQX + RWIS 1h group significantly reduced.These results suggested that Glu in "brain stem-gut axis" may directly bind to the AMPA receptors and affect the activity of neurons and glial cells in the enteric nervous system.3.Compared with the normal saline + RWIS 1h group,the expressions of occludin and claudin1 in the intestinal wall of the CNQX + RWIS 1h group significantly increased,meanwhile,the expression of MUC5 AC in the gastric mucosa also significantly increased.These results suggested that Glu in "brain stem-gut axis" may affect gastrointestinal function through AMPA receptors.Based on the above experimental results,Glu in the "brain stem-gut axis" regulates the nervous system through AMPA receptors and participated in the RWIS process,which may play a critical role in RWIS-induced gastric mucosa damage.