Effects Of Diazoxide Preconditioning On The Expressions Of Apoptosis And C-Jun N-terminal Kinase In Cultured Rat Hippocampal Neurons After Anoxia-reoxygenation In Vitro

Objective To investigate the effects and possible relationship of diazoxide (a selective opener of mitochondrial ATP-sensitive potassium channel,mitoKATP) precon-ditioning on the expressions of apoptosis and c-Jun N-terminal kinase (JNK) signali-ng transduction pathway in primary cultured rat hippocampal neurons in vitro cause-d by anoxia-reoxygenation injury.Methods 100 Sprage-Dawley rats (<24 h after birth) weighing 5～6 g were decapitated,hippocampal tissue was isolated.After being cultured for 9～10 d,the hippocampal neurons were randomly assigned to one of 5 groups:Control group (Gr-oup A),diazoxide 0,30,100 umol/ L (Group B,C,D) and 5-hydroxydecanoate (a specific inhibitor of mitoKATP) 100 umol/ L + diazoxide l00 umol/ L (Group E). The neurons were treated with diazoxide l h per day for 3 d before being subjected to 4 h oxygen deprivation followed by 24 h reoxygenation.The neuronal viability was measured by MTT assay.Expressions and variability of Bcl-2,Bax and JNK proteins were detected by Western blot analysis.Results The results obtained are as follows:①The survival rates of the rat hippocampal neurons subjected to anoxia-reoxygenation were diminished than that inGroup A significantly (P<0.01) .Bc1-2,Bax and JNK proteins were up-regulated than those in Group A (P<0.01,P<0.05);②Compared with that in Group B,the neuronal survival rates rise in both Group C and Group D (P<0.01,P<0.05) .The survival rate in Group C was increased by about 5% and by nearly 12% in Group D.The expression of Bcl-2 protein in both Group C and Group D was upgraded (P<0.05,P<0.01)by approximately 25% and 100% each.Bax and JNK proteins were decreased (P<0.05) .The expression of Bax protein was descended by almost 22% and 42% each,while JNK protein was lower by 19% and 35% respectively. No difference was detected between Group B and Group E (P>0.05);③There was significant difference between Group C and Group D (P<0.05,P<0.01) .By contra-st to Group C,the neuronal survival rate in Group D was advanced by almost 7%,Bcl-2 expression was increaseed by 25%,whereas Bax and JNK expressions were decreased by 27% and 19% respectively.Conclusion Diazoxide (a selective opener of mitoKATP) preconditioning may at-tenuate neuronal apoptosis and enhance the tolerance against anoxia-reoxygenation injury in primary cultured hippocampal neurons.JNK signaling transduction pathway plays a vivtal role in the neuronal apoptosis.Diazoxide may lower the expression of JNK protein,and increase cell survival ratio through up-regulation of Bcl-2 and down-regulation of Bax expression by JNK signaling transduction pathway.