Effects of Puerarin on the Levels of Calcium and Nitric Oxide in Cultured Hippocampal Neurons during Oxygen-Glucose Deprivation

OBJECTIVE To investigate the effects of puerarin(Pur) on the Levels of calcium and nitric oxide in cultured hippocampal neurons during oxygen-glucose deprivation.METHODS Primary hippocampal cultures were prepared from 2 day-old Sprague-Dawley rats.After 8 d in vitro,the cultures subjected to 3 h of oxygen/glucose deprivation(OGD) or 30 min of glutamate treatment.Pur at 40 or 100 μmol·L-1 were added into the cultures during OGD or treatment of glutamate and for the following 24 h respectively.Apoptosis and necrosis were detected using flow cytometric analysis of Annexin-V and PI labeling cells 24 h after OGD or treatment of glutamate.Futhermore,the changes in the levels of intracellular Ca2+ or nitric oxide(NO) were measured by confocal laser scanning microscopy with Fluo-3,Ca2+ probe,or DAF-2,NO probe respectively,during 30 min OGD.RESULTS OGD induced a rapid Ca2+ influx and NO synthesis,and the peaks of the intracellular Ca2+ or NO reached the levels of 2.9 or 1.9 times of the control.However,Pur(40,100 μmol·L-1) markedly slowed down OGD-induced Ca2+ influx and lowered the intracellular Ca2+ peak by 29.2% and 37.1%(P0.05 and P0.01),and significantly inhibited OGD-induced NO synthesis by 23% and 33%(P0.05 and P0.01).Meanwhile,apoptosis and necrosis resulted from OGD or glutamate excitotoxicity were significantly reduced in the cultured hippocampal neurons by addition of Pur.CONCLUSION Pur can ameliorate hippocampal neuronal death induced by OGD in vitro.The protective effects of Pur are associated with inhibiting the action of glutaminergic transmitter,intracellular Ca2+ elevation and neuronal NO synthesis.