Identification Of The Nociceptors Of Gastrointestinal Tract On Vagal Afferent Nerves And The Effect And Mechanism Of 6-shogaol On Gastrointestinal Nociceptor

Objectives: In somatosensory,the pain mediated by nociceptor is an important protective mechanism of the body.With the discover of TRP channels,we now could make progress in the elucidation of the molecular mechanism of pain.However,due to some objective reasons,limited progress was made in visceral sensations,especially the role of vagal nerve.Whether the vagal nerve is involved in the transduction of nociception is still controversial.In this study,we aimed to identify the nociceptors of gastrointestinal tract on vagal afferent nerves,and studied the effect and mechanism of 6-shogaol on gastrointestinal nociceptor.And we also investigated the underlying peripheral mechanism of 6-shogaol on CINV.Methods:(1)The method used to develop the mouse gastroesophageal-vagal preparations with intact nerve ending in either the esophagus or the stomach was modified from a extensively used to record from guinea-pig esophageal-vagal;(2)Immunofluorescence histochemical staining of the vagus ganglion was performed to analyze the expression of TRPV1;(3)we developed a novel approach by using two-photon neuron imaging technique to address the neuronal population response in ex vivo gastroesophageal-vagal preparation from our newly-established Pirt-GCa MP6 s transgenic mouse line;(4)we performed extra-cellular single-unit recording of action potential discharges in ex vivo gastroesophageal-vagal preparation from wild type mouse;(5)Combined two-photon neuron imaging and extra-cellular single-unit recording,we studied the effect and mechanism of 6-shogaol on gastrointestinal nociceptor;(6)we established a model of chemotherapy-induced nausea and vomiting to explore whether 6-shogaol can exert anti-emetic effects and related mechanisms through the vagus nerve.Results:(1)We successfully develop the mouse gastroesophageal-vagal preparations with intact nerve ending in either the esophagus or the stomach,which has been experimentally proven,can maintain the activity of the tissues and nerve for more than 8 hours.(2)The conduction velocity of the gastroesophageal nociceptors on the vagus nerve was <1 m/s;the immunofluorescence results showed that the TRPV1 was highly expressed in the neurons in the vagus ganglion;all of them belonged to the characteristics of unmyelinated C fibers.(3)The vagus nerve terminals can sense the noxious stimuli from the gastrointestinal tract,such as noxious mechanical distension,acid,?,?-methylene-ATP,AITC,and Capsaicin.(4)6-shogaol,an important bioactive phytochemical from ginger,can directly activate nociceptors in the esophagus and stomach of the vagus nerve,and this effect is mainly mediated by the TRPA1 channel.(5)After 6-shogaol pretreatment,desensitization of vagal nerve endings reduced the re-stimulation of chemical stimuli(included itself,AITC and Cap).(6)VCR treatment can cause a decreased food intake in mice and delay of gastric emptying.(7)VCR can activate vagus nerve endings in the gastrointestinal tract.(8)The 5-HT3 receptor antagonist palonosetron,TRPA1 channel inhibitor HC-030031 and 6-shogaol can inhibit the effect of VCR on vagus nerve endings,respectively.Conclusions:(1)There are many gastroesophageal nociceptors on the vagus ganglion,and their nerve ending are distributed in tissues such as esophagus and gastrointestinal tract.Like most somatosensory nociceptors,they are unmyelinated C fibers and the conduction velocity are slow,and they can also sense and transmit various noxious stimuli from target organs,such as mechanical distension,acid and various chemical stimuli.(2)6-shogaol,an important bioactive phytochemical from ginger,activated esophageal and gastric nociceptors on the vagus nerve mainly via TRPA1.The desensitization caused by 6-shogaol was closely related to the effects of ginger,such as analgesia and relief of nausea and vomiting.The TRPA1 channel may be a target for developing new antiemetics or visceral pain in the future.(3)VCR can stimulate the vagus nerve endings in the gastrointestinal tract,and desensitization of vagal nerve endings after 6-shogaol pretreatment can inhibit the effect of VCR on the vagus nerve endings.It may be one underlying mechanism of ginger to alleviate CINV.