Protective Effects Of Cromakalin On Rats Following Focal Cerebral Ischemia/reperfusion And Its Mechamism

Objective To investigate the effects of cromakalin on rats following focal cerebral ischemia/reperfusion,and its mechanism about neuronal apoptosis by mitochondria signaling pathways and cerebral energy supply.Methods 60 male Wistar rats were randomly divided into four groups:A(sham -operated group);B(ischemia/reperfusion group);C(K_ATP opener treatmeat group) and D(K_ATP opener and blocker treatment group).The middle cerebral artery occlusion (MCAO) models were established by using the Zea Longa intraluminal suture occlusion method,the infarct volumes were studied by TTC staining,scores of neurological function deficits were evaluated,neuronal apoptosis were detected by TUNEL staining,the expression of cytochrome C were detected by immunohistochemical,SOD activity of the brain tissue were detected according to the xanthine oxidase method,MDA content of the brain tissue were detected according to the thio-barbituric acid method,the activities of Na⁺-K⁺-ATPase and Ca²⁺-ATPase of the brain tissue were detected according to the inorganic phosphonium method,to investigate the protective effects of K_ATP openers and its mechanism about mitochondrial opoptosis pathway,and to take initial investaction on its influence about energy metabolism.All data were expressed as mean±SD.Statistical analysis was carried out by one-way ANOVA.Results1.Infarct volumesThere were not infarcts in A group.The infarcts in B,C and D groups were mainly located in right frontal-parietal cortex,caudate nucleus and putamen nucleus.The infarct volumes of C group were significantly less than these in B and D groups(P＜0.01).There were no differerences between B and D groups(P＞0.05).2.Evaluation of neuorological function deficitsCompared with A group,the scores of B,C,D groups were significantly different (P＜0.01),the scores of C group were significantly less than these in B and D groups (P＜0.05),there were no differences between B and D groups(P＞0.05).3.Neuronal apoptosisApoptotic neurons were mainly located around ischemic tissues,that was,ischemic penumbra.Apoptotic cells were rare in the ischemic core region.There were a few opoptotic cells in A group.In B group,there were many apoptotic cells,the rate of apoptosis positive cells increased obviously.In C group,the rate of apoptosis positive cells were lower than that in B group(P＜0.01).There was no difference about the rate of apoptosis positive cells between B group and D group(P＞0.05).4.ImmunohistochemistryThere were a few positive cells in A group.The positive cells of B,C and D groups were mainly located in ischemic penumbra.In B group,the expression of cytochrome C increased obviously.The most obvious area was ischemia penumbra.Positive cells were labeled brown in cytoplasm,grainy.Some cells were labeled brown in cytoplasm and also in the nuclei.In C group,the expressions of cytochrome C were obviously lower than that in B group,there was significant difference(P＜0.01).There was no difference about expressions of cytochrome C between B group and D group(P＞0.05).5.SOD activity and MDA content in brain tissuesIn B group,the SOD activity reduced,the MDA content increased,compared with group A(P＜0.05).It showed that there was oxidative stress injury during cerebral ischemia reperfusion.Compared with group B,the SOD activity increased,the MDA content decreased in group C(P＜0.05).There was no difference about the SOD activity and MDA content between B group and D group(P＞0.05).6.activity of Na⁺,K⁺-ATPase and Ca²⁺-ATPase in brain tissuesIn B group,the activity of Na⁺,K⁺-ATPase and Ca²⁺-ATPase reduced,compared with group A(P＜0.05).Compared with group B,the activity of Na⁺,K⁺-ATPase and Ca²⁺-ATPase increased in group C(P＜0.05).There was no difference about the activity of Na⁺,K⁺-ATPase and Ca²⁺-ATPase between B group and D group(P＞0.05).Conclusion1.K_ATP openers can signigicantly decrease the infarct volumes,improve neurological function deficits,this result indicates that K_ATP openers have a protective effect on cerebral ischemia/reperfusion.2.K_ATP openers have a protective effect on cerebral ischemia/reperfusion by decreasing neuronal apoptosis in ischemia penumbra.3.K_ATP openers can significantly decrease the expressions of cytochrome C following cerebral ischemia/reperfusion,this result indecates that K_ATP openers can decrease neuronal apoptosis by mitochondria signaling pathways.4.K_ATP openers can significantly increase the activities of Na⁺-K⁺-ATP, Ca²⁺-ATPase and SOD,reduce the content of MDA,these results indecate that K_ATP openers can improve cerebral energy supply,decrease production of oxygen free radical, have a protective effect on cerebral ischemia/reperfusion.