The Role Of Primary Motor Cortex In Stress-induced Gastric Mucosal Injury In Rats

Restraint Water-Immersion Stress(RWIS)is a compound stress that causes great psychological and physiological stimulation in rats.The stressful stimulus will disturb the balance between the central nervous system and the gastrointestinal axis in rats within a few hours,leading to gastric dysfunction and thus inducing stressful gastric mucosal injury in rats.Our group has long been engaged in the study of neural mechanisms of gastrointestinal dysfunction caused by restraint water immersion stress in restraints in rats.Our previous research has revealed that the brainstem vagus complex,parabrachial nucleus,hypothalamic paraventricular nucleus,supraoptic nucleus and dorsomedial thalamic nucleus in the parasympathetic nervous system are involved in the regulation of stress-related gastric mucosal injury caused by restraint immersion stress in rats.The sympathetic nervous system is also excited during stress and it has been found that most of the higher centers of the sympathetic nervous system are concentrated in the primary motor cortex,so is the primary motor cortex involved in the process of water immersion stress? What is the role of primary motor cortex in the regulation of stressful gastric mucosal injury? Does primary motor cortex modulate stressful gastric mucosal injury by affecting the rostral ventrolateral medulla(RVLM),a sympathetic hypocenter of the medulla oblongata? These questions have not been reported.The present study therefore raises the following questions.1.What is the status of gastric mucosal damage and the time-space changes of c-Fos and GFAP expression in M1 after RWIS in rats?2.What is the effect on stress gastric mucosal injury after inhibition of M1 mature neurons?3.Are M1 glutamatergic neurons involved in the stress process of restraint immersion?What is the effect on stressful gastric mucosal injury after inhibition/activation of M1 glutamatergic neurons?4.Is the RVLM involved in bound immersion stress and does the change in excitability of M1 affect the excitability of the RVLM?To address the above four questions,we designed the following experiments:1.The experimental rats were randomly divided into three groups: control group,RWIS 1h,RWIS 3h.The macroscopic statistics of gastric ulcer index were calculated by Guth counting method,and the expression of gastric Claudin1,Occludin and PCNA protein were determined by Western Blot.The changes in the time-space expression of c-Fos and GFAP,a specific marker of astrocytes in M1,were probed by immunohistochemistry after 1h and 3h of RWIS in rats.2.Wistar Rats were randomly divided into three groups: Empty virus group(AAV9-h Synm Cherry + intraperitoneal injection CNO),h Syn-h M4D(Gi)virus inactivated group(AAV9-h Syn-h M4D(Gi)-m Cherry + intraperitoneal injection SA),h Syn-h M4D(Gi)virus suppressed group(AAV9-h Syn-h M4D(Gi)-m Cherry+intraperitoneal injection CNO),and injected virustransfected rats were fasted for 24 h with injected drugs for three weeks.RWIS 3h after drug injection 0.5h.The expression of c-Fos and GFAP within M1 was detected by immunohistochemistry.Western Blot technique was used to detect the expression of Claudin1,Occludin and PCNA proteins in the stomach to investigate the role of primary motor cortex in the process of stressful gastric mucosal injury in rats.3.Rats were randomly divided into three groups: Empty virus group(p AAV-Ca MKIIaMCS-3Flag + intraperitoneal injection CNO),Ca MKIIa-h M4D(Gi)virus inactivated group(p AAV-Ca MKIIa-h M4D(Gi)-m Cherry-3Flag + intraperitoneal injection SA),Ca MKIIah M4D(Gi)virus suppressed group(p AAV-Ca MKIIa-h M4D(Gi)-m Cherry-3Flag + intraperitoneal injection CNO).and the same intraperitoneal injection operation as 2.The expression of c-Fos and GFAP within M1 was detected by immunohistochemistry.Western Blot technique was used to detect the expression of Claudin1,Occludin and PCNA proteins in the stomach;activation of M1 glutamatergic neurons was performed only by replacing the virus with p AAV-Ca MKIIah M3D(Gq)-m Cherry,the same empty virus group,and the rest of the operations were the same.To investigate the role of inhibiting glutamatergic neurons in primary motor cortex during stressful gastric mucosal injury in rats.4.The expression of c-Fos and GFAP within RVLM was detected by immunohistochemical techniques in the corresponding groups.To investigate whether the RVLM is involved in the fasciculation immersion stress process;whether the inhibition/activation of M1 neurons affects the excitability of neurons and astrocytes within the RVLM.The experimental results showed that:1.Compared with the control group,the gastric mucosal damage of rats after RWIS 1h and 3h was more serious,and the gastric ulcer index was extremely significantly higher than that of the control group.Our experimental results revealed that the expression of Claudin1,Occludin,and PCNA proteins in the gastric mucosa of rats in the RWIS 1h and 3h groups were significantly lower compared with the control group,and the expression of c-Fos and GFAP in the anastomotic,middle and caudal segments of rat M1 was increased.These results suggest that bound immersion stress significantly disrupts the gastric barrier and M1 is significantly excited.2.RWIS 3h after three weeks of viral transfection injection,the expression of c-Fos and GFAP in M1 was significantly reduced in h Syn-h M4D(Gi)virus suppressed group compared with the empty virus group and the h Syn-h M4D(Gi)virus inactivated group,and the content of Occludin,Claudin1 and PCNA protein in the stomach was significantly reduced.The experimental results indicated that successful inhibition of mature neurons in primary motor cortex exacerbated the extent of gastric mucosal damage,and M1 may play a protective role for gastric mucosa during RWIS.3.RWIS 3h after three weeks of viral transfection injection,the expression of c-Fos and GFAP in M1 was significantly reduced in Ca MKIIa-h M4D(Gi)virus suppressed group compared with the empty virus group and Ca MKIIa-h M4D(Gi)virus inactivated group,and the content of Occludin,Claudin1 and PCNA protein in the stomach was significantly reduced.The expression of c-Fos and GFAP in M1 was significantly higher in Ca MKIIa-h M3D(Gq)virus activated group compared with the empty virus group and Ca MKIIa-h M3D(Gq)virus inactivated group,and the content of Occludin,Claudin1 and PCNA protein in the stomach was significantly increased.The results showed that inhibition of M1 glutamatergic neurons increased the extent of gastric mucosal injury while activation reduced it,indicating that glutamatergic neurons play an important role in M1 protection against stressful gastric mucosal injury.4.Expression of c-Fos and GFAP within the RVLM of bound immersion-stressed rats was significantly increased,indicating a process of RVLM stress gastric mucosal injury.Inhibition/activation of neurons within M1 significantly decreased/increased the expression of cFos and GFAP in the RVLM of bound immersion-stressed rats,suggesting that M1 may be involved in the stressful gastric mucosal injury process through the RVLM downstream visceral sympathetic pathway.In summary,it can be seen that.1.M1 is involved in the process of fasciculation immersion stress in rats.2.Inhibition of neurons in M1 significantly aggravated the stressful gastric mucosal injury in rats caused by bound immersion stress.3.Inhibiting glutamatergic neurons in M1 can significantly aggravate the stressful gastric mucosal injury in rats caused by bound immersion,while activating glutamatergic neurons in M1 can reduce the stressful gastric mucosal injury.4.M1 may be involved in the regulation of stressful gastric mucosal injury through the M1-RVLM pathway.This study provides new clinical ideas for the treatment of stress-related gastric mucosal injury.