The Role Of P2X7 Receptor In The Changes Of Protein Kinase Activity In The Cerebral Cortex Of Insomnia Rats Induced By PCPA And The Intervention Of Suanzaoren Decoction

Objective:The changes of mRNA and protein expression in cerebral cortex signal molecule P38MAPK,PKC and CaMK? after insomnia induced by PCPA in PCPA rats were observed.The effects of P2X receptor antagonist,P2X7 receptor antagonist and agonist on the activity of cerebral cortex protein kinase induced by PCPA in rats were observed by intraventricular injection of P2X receptor antagonist and P2X7 receptor antagonist.The effects of SZRD on the expression of P38MAPK,PKC,CaMK II mRNA and protein in the cerebral cortex of the insomnia rats were observed with SZRD intervention.Method:1.Healthy and clean grade SD rats were randomly divided into normal group,surgical control group,PCPA group,PCPA+OXATP group,PCPA+A438079 group and PCPA+BzATP group.There were 6 groups in each group.In addition to the normal group,each other group was intubated by ventricular intubation,and the animals had to recover 7 days after the intubation to observe if there was' any abnormal condition.The model of insomnia in rats was established by intraperitoneal injection of PCPA for 3 days.After intraperitoneal injection of each rat,intraventricular injection was performed immediately.P2X receptor antagonist OxATP was intracerebroventricularly injected into the PCPA+OXATP group,P2X7 receptor antagonist A438079 was intracerebroventricularly injected into the PCPA+A438079 group,and P2X7 receptor agonist BzATP was intraventricularly injected into the PCPA+BzATP group.The mRNA and protein expression of P38MARK,PKCand CaMKII were detected by RT-PCR and Western blot.2.Healthy and clean SD rats were randomly divided into normal group,surgical control group,PCPA group,PCPA+SZRD group,PCPA+OXATP+SZRD group,PCPA+AA438079+SZRD group,PCPA+BzATP+SZRD group,surgical control+SZRD A total of 8 groups,6 in each group.In addition to the normal group,the remaining animals in each group required lateral ventricle catheterization,animals need to recover after 7 days.Then the insomnia model of rats was prepared by continuous intraperitoneal injection of PCPA.In the normal control group,surgical control group,and surgical+SZRD group,an equal volume of physiological saline was intraperitoneally injected in the same manner.After intraperitoneal injection,intraventricular injection was performed immediately.In addition to normal saline injected intracerebroventricularly in the PCPA+SZRD group and the surgical control+SZRD group,each group of intracerebroventricular injections and methods were the same as the experiment one.The difference lies in the SZRD(15g·kg-1)gavage treatment performed on the basis of the first experiment for 7 consecutive days.The normal group,surgical control group and PCPA group were intragastrically administered with the same volume of saline.After intragastric administration,the mRNA and protein expressions of P38MAPK,PKC and CaMKII mRNA in P2X7 receptors in cerebral cortex of rats were measured by Western blot and RT-PCR.Result:1.Compared with the control group,the expression of P38MARK mRNA and protein in the cerebral cortex of PCPA group was significantly up-regulated(P<0.01).Compared with the model group,the mRNA and protein expression of P38MARK in PCPA+OXATP group and PCPA+A438079 group were significantly down-regulated(P<0.05).The expression of P38MARK mRNA and protein in PCPA+BzATP group was higher than that in model group(P<0.05).There was no significant difference in the expression of P38MARK mRNA and protein between the normal control group and the surgical control group.Compared with the control group,the expression of PKC and CaMKII mRNA and protein in the cerebral cortex of PCPA group was significantly down-regulated(P<0.01).Compared with the model group,the mRNA and protein expressions of PKC and CaMKII in the P2X receptor antagonist group and the P2X7 receptor antagonist group were significantly upregulated(P<0.01).The expression of PKC and CaMK? mRNA and protein in P2X7 receptor agonist group was significantly down-regulated(P<0.01).2.Compared with PCPA group,the expression of P38MAPK mRNA and protein in PCPA+SZRD group was down-regulated(P<0.01),and the expression of PKC and CaMK? mRNA and protein was up-regulated(P<0.01).Compared with PCPA+OXATP group,the expression of P38MAPK mRNA and protein in PCPA+OXATP+SZRD group was up-regulated(P<0.01),and the expression of PKC and CaMKII mRNA had no significant difference(P>0.05).The expression of PKC and CaMKII protein was up-regulated(P<0.01).Compared with PCPA+A438079 group,the expression of P38MAPK mRNA and protein in PCPA+A438079+SZRD group was up-regulated(,P<0.01).Compared with PCPA+BzATP group,the expression of P38MAPK mRNA and protein in PCPA+BzATP+SZRD group was up-regulated(P<0.01),the expression of PKC and CaMKII mRNA was significantly down-regulated(P<0.01),and the protein expression of PKC and CaMKII was up-regulated(P<0.01).There was no significant difference in the expression of PKC and CaMKII mRNA(P>0.05).The expression of PKC and CaMKII protein was up-regulated(P<0.01).There was no significant difference in the expression of P38MAPK,PKC,CaMKII mRNA and protein between the normal group and the control group(P>0.05).There was no significant difference in the expression of P38MAPK mRNA and protein between the surgical control+SZRD group and the surgical control group(P>0.05).Conclusion:1.PCPA can induce the expression of P38MAPK mRNA and protein in the cerebral cortex of insomnia rats,and down-regulate the expression of PKC and CaMKII mRNA and protein.2.P2X7 receptor mediates changes in P38MAPK,PKC,CaMKII mRNA and protein expression induced by PCPA.3.The mechanism of SZRD in improving insomnia is likely to reduce P38MAPK mRNA and protein expression through P2X7 receptors,and upregulate the expression of PKC and CaMKII mRNA and protein,thus improving insomnia.