Abstract:

Abstract:Objective  To study the neuroprotective effects of Humanin(HN) on exciatory neurotoxicity  based on inhibiting  the expressions of NMDA receptor NR1 subunits and  phosphorylation of NR2B subunits, and to explore the possible mechanisms.Methods Primary cultured  cerebral cortical neurons of Wistar rats were used to set up excitatory neuotoxicity model and  were divided into control group, NMDA group,NMDA+MK-801 (MK-801) group and NMDA+HN group.The expressions  of NR1 subunit proteins of NMDA receptors  were detected by flow cytometry  (FCM).The levels of NR2B subunits of NMDA receptors phosphorylation at Tyr1472 in various  groups were detected by Western blotting.Results  The results of inverted phase contrast microscope showed that in NMDA group the cerebral cortical neurons were obvious neuronal loss,few cells became round and there was halofuzzy around the cells and the neurices were mostly gone, and the cells showed obvirous  excitatory neurotoxicity.The mean fluorescence intensity value of NR1 subuint protein of NMDA receptor  dsignificantly was increased  compared with control group（P<0.05）,the mean fluorescence intesity values of NR1 subunit protein were  reduced in MK-801  group and HN group compared with NMDA group（P<0.05）.The Western blotting results showed that there were no significant changes of total NR2B subunit proteins in various groups.The level of NR2B phosphorylation at Tyr1472 induced by NMDA  was increased significantly compared with control group（P<0.05）, while the pretreatment of HN(1  μmol/L) attenuated the level of phosphorylated of NR2B subunit  in primary cultured rat cerebral cortical neurons compared with control group（P<0.05）.Conclusion The up-regulation of the expression level of NR1 subuint protein of NMDA receptor and the  increaing  of the level of the phosphorylated of NR2B subunit  might be responsible for the excitatory       neurotoxicity induced by NNDA.HN might contribute to the neuroprotection against excitatory neurotoxicity  by inhibiting the expression of NR1 subunits and reduction of phosphorylated of NR2B.

Key words: Humanin, N-methyl-D-aspartate receptor, NR1 subunit, NR2B subunit, excitatory neurotoxicity