| Background and ObjectivePostoperative ileus is an iatrogenic disorder characterized by a transient inhibition of gastrointestinal motility,which develops after each abdominal surgical procedure.Postoperative ileus clearly has a significant impact on patient morbidity with symptoms such as pain,nausea and vomiting,abdominal distension,absence of defecation and intolerance to oral feeding.Postoperative ileus not only increases patients’ pain and reduces their quality of life,but also is the most common cause of prolonged hospital stay following abdominal surgery and there is still lack of effective targeted agents for treating postoperative ileus.As a unique external treatment of Traditional Chinese Medicine,acupuncture has the functions of dredge channels and collaterals,invigorate qi and activate blood circulation.The advantage of acupuncture and moxibustion lies in the active mobilization of the body’s own potential ability to restore the maladjusted function.In recent years,neuro-mediated immune response in gastrointestinal system has become a new hotspot in the study of acupuncture in regulating gastrointestinal motility,especially the cholinergic anti-inflammatory pathway mediated by vagus nerve.Postoperative ileus originates from intestinal inflammation triggered by resident muscularis macrophages.Acupuncture has been shown to increase gastrointestinal motility and alleviate intestinal inflammatory response.Our previous studies indicated that EA attenuated POI via the activation of the α7 nicotinic acetylcholine receptor(α7nAChR)-mediated cholinergic anti-inflammatory pathway in resident macrophages.Cholinergic antiinflammatory pathway is mediated by the vagus nerve.How does acupuncture stimulate the vagus nerve to promote gastrointestinal motility?Therefore,on the basis of previous studies,this study explored the mechanism of EA at ST36 in postoperative ileus mice based on the vagus pathway mediated by GABAA receptors on cholinergic neurons in DMV.Based on the vagus nerve promoting gastrointestinal motility,this study explored the potential neurological mechanism of EA in mice with postoperative ileus after ST36 treatment by neurotomy and intraventricular injection,so as to clarify the neurobiological basis of electroacupuncture and provide scientific basis for clarifying the biological mechanism of acupuncture in preventing and treating gastrointestinal dynamic diseases.Methods1.The peripheral vagus nerve mechanism of ST36 in postoperative ileus:left cervical vagotomy and sub-diaphragmatic vagotomy were performed separately.C57BL/6 mice was randomly divided into seven groups,namely:model group,model+left cervical vagotomy group,model+left cervical vagotomy+electroacupuncture group,model+sham left cervical vagotomy+electroacupuncture group;model+sub-diaphragmatic vagotomy group,model+sub-diaphragmatic vagotomy+electroacupuncture group,model+ sham sub-diaphragmatic vagotomy + electroacupuncture group;In each group of 12 mice,6 mice were tested for gastrointestinal motility and 6 mice were detected the expression of inflammatory factors TNFα and IL-6 via Real-time PCR and ELISA.2.The central mechanism of ST36 in postoperative ileus:Firstly,3 mice were used for DMV immunohistochemistry in sham group,model group,electroacupuncture group and nonacupoint group.Secondly,C57BL/6 mice was randomly divided into five groups with DMV microinjection,namely:model+Nacl+electroacupuncture group,model+Mus group,model+Mus+electroacupuncture group,model+BIM group,model+BIM+electroacupuncture group.6 mice were tested for gastrointestinal motility and 6 mice were detected the expression of inflammatory factors TNF-α and IL-6 via Real-time PCR and ELISA.3.The peripheral somatic nerve mechanism of ST36 in postoperative ileus:Firstly,C57BL/6 mice was randomly divided into four groups,namely:model group,model+ sciatic section group,model+sciatic section+electroacupuncture group,model+sham sciatic section+electroacupuncture group.6 mice were tested for gastrointestinal motility and 6 mice were detected the expression of inflammatory factors TNF-α and IL-6 via Real-time PCR and ELISA.Secondly,mice were randomly divided into four groups,namely:sham group,model group,electroacupuncture group and non-acupoint group.3 mice were used for spinal cord immunohistochemistry in each group.Results1.The peripheral vagus nerve mechanism of ST36 in postoperative ileus:Compared with model group,the gastrointestinal motility and the levels of TNF-α and IL-6 were no significant difference in model+sub-diaphragmatic vagotomy group(P>0.05).Compared with model group or model+sub-diaphragmatic vagotomy group,the gastrointestinal motility of model+sub-diaphragmatic sham vagotomy+electroacupuncture group was better,and the level of TNF-α and IL-6 in serum or intestinal muscle layer was also lower than that in the model group or model+sub-diaphragmatic vagotomy group(P<0.05).Compared with the model+subdiaphragmatic sham vagotomy + electroacupuncture group,the model+sub-diaphragmatic vagotomy+electroacupuncture group had a decrease in gastrointestinal motility(P<0.01),and the levels of TNF-α and IL-6 were significant elevated(P<0.05),so sub-diaphragmatic vagotomy could inhibit the improvement effect of electroacupuncture on postoperative ileus.Compared with model group,the gastrointestinal motility and the levels of TNF-α and IL-6 were no significant difference in model + left cervical vagotomy group(P>0.05).Compared with the model group or the model+left cervical vagotomy group,the motility of the model+left cervical sham vagotomy+electroacupuncture group was increased(P<0.01),and the levels of TNF-α and IL-6 were lower(P<0.05).Compared with the model+electroacupuncture+left cervical sham vagotomy group,the model+electroacupuncture+left cervical vagotomy group had reduced gastrointestinal motility(P<0.01)and increased levels of inflammatory factors(P<0.05).Therefore,left cervical sham vagotomy could also inhibit the improvement effect of electroacupuncture on gastrointestinal motility and inflammation in mice with postoperative ileus.2.The central mechanism of ST36 in postoperative ileus:Compared with the sham group.the expression of GABAA receptors on the cholinergic neurons in the DMV of the model group was increased.Compared with the model group,the expression of GABAA receptors in the electroacupuncture group or the non-acupoint group was reduced.Compared with the model+Nacl+electroacupuncture group,the gastrointestinal motility of the model + Mus+electroacupuncture group was significantly weakened(P<0.01),and the expression levels of serum and intestinal muscular TNF-α and IL-6 were increased(P<0.05).There is no statistical difference in gastrointestinal motility and inflammatory factor expression between the model+Mus+electroacupuncture group and model+Mus group(P>0.05),so the GABAA receptor agonist Mus can inhibit the effects of electroacupuncture on the postoperative ileus.Compared with the model+Nacl+electroacupuncture group,the model+BIM group had no statistically significant difference in gastrointestinal motility and inflammatory factors(P>0.05).Therefore,the GABAA receptor inhibitor BIM could simulate electroacupuncture.However,compared with the model + BIM group,the gastrointestinal motility of the model+BIM+left cervical vagotomy group was weakened(P<0.05),and the levels of serum and intestinal muscularis TNF-α and IL-6 were increased(P<0.05),indicating that the left cervical vagotomy could inhibit the effect of BIM.3.The peripheral somatic nerve mechanism of ST36 in postoperative ileus:Compared with model group,the gastrointestinal motility and the levels of TNF-α and IL-6 were no significant difference in model+sciatic section group(P>0.05).The gastrointestinal motility of the model+electroacupuncture+sham sciatic section group was higher than that of the model group or model+sciatic section group(P<0.01),and the levels of inflammatory factors TNF-α and IL6 in serum or intestinal muscle layer were also lower(P<0.05).Compared with the model+electroacupuncture+sciatic sham section group,the model+electroacupuncture+sciatic section group had reduced gastrointestinal motility(P<0.01)and increased levels of inflammatory factors in serum and intestinal muscularis(P<0.01).Therefore,sciatic section could inhibit the effect of electroacupuncture on gastrointestinal motility and inflammation.Compared with the model group,the expression of c-Fos in the spinal dorsal horn neurons of the lumbar 4 to 6(L4-L6)group of the electroacupuncture group or the non-acupoint group was increased,and the electroacupuncture group was more increased.Conclusions1.Peripheral cervical vagus nerve or sub-diaphragmatic nerve may mediate electroacupuncture to improve the outflow of gastrointestinal motility signal on postoperative ileus.2.Electroacupuncture at Zusanli may stimulate the vagus nerve and improve gastrointestinal motility and inflammation by inhibiting the expression of GABAA receptor on cholinergic neurons in DMV of postoperative ileus mice.3.Sciatic nerve and L4-L6 spinal dorsal horn neurons may participate in electroacupuncture at Zusanli to improve the transmission of gastrointestinal motility signals on postoperative ileus.In summary,electroacupuncture at Zusanli may improve the gastrointestinal motility of postoperative ileus mice through the DMV-vagus nerve pathway,and the sciatic nerve and L4L6 dorsal horn neurons may participate in acupuncture signals. |
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