Objective：The purpose of this study was to investigate the role of autophagy in oxygen-glucose-deprivation/reoxygenation（OGD/R）injury in rat neurons. Methods：Cortical neurons were isolated from Sprague-Dawley rats and identified by immunofluorescence. The cortical neurons were randomly assigned to four groups：control group（Ⅰ），experimental group（OGD/R group，Ⅱ），JNK inhibitor pretreatment group（Ⅲ）and JNK inhibitor pretreatment+OGD/R group（Ⅳ）. Results：Neuronal cell viability significantly decreased after 6 h and 12 h of reoxygenation in Group Ⅳ（P < 0.05）. Electron microscopy results showed the presence of many autophagic vacuoles and the formation of autolysosomes in the neurons；the number of autophagic vacuoles decreased transiently at 6 h，while a new autophagic flux and a large number of empty autophagic vacuoles were observed at 12 h. In Group Ⅳ，a large number of autophagic vacuoles were present at 0.5 h and 2 h of reoxygenation，which gradually decreased with increasing reoxygenation time. No significant differences in the expression of the LC3Ⅱprotein were detected between the GroupⅡ and Ⅳ prior to 6 h of reoxygenation，and LC3Ⅱ expression showed an overall rise-decline pattern. However，LC3Ⅱ protein expression increased in Group Ⅱ at 12 h of reoxygenation，whereas a continuous decline was observed in Group Ⅳ. The levels of phosphorylated JNK and Bcl-2 and the expression of Beclin-1 increased gradually as the reoxygenation time going in Group Ⅱ，whereas they increased at 12 h of reoxygenation in Group Ⅳ（P < 0.05）. In addition，progressive dissociation of the Bcl-2/Beclin-1 complex was observed in the Group Ⅱ，while JNK inhibitor suppressed this dissociation. Conclusion：The regulation of the JNK/Bcl-2/Beclin-1 signaling pathway may be one of the mechanisms underlying the OGD/R-induced autophagic cell death of neurons.