Objective To determine whether or not apoptosis occurs in the delayed cell death following hypoxic ischemic injury, and assess the sensibility and specificity of various techniques. Methods Using HE staining and electromicroscopic technique, in situ end labeling (ISEL) and DNA electrophoresis, we observed the histological features of apoptotic cells in the cerebral cortex and hippocampus, and compared the ipsilateral hemisphere at various time points following hypoxic ischemia (HI) with that of the sham group. Results In the cerebral cortex and hippocampus at the ipsilateral hemisphere in neonatal rat models of HI, light and electronic microscopy exhibited cell shrinkage, chromatin condensation, and apoptotic bodies. ISEL and DNA electrophoresis also provided evidence of DNA fragmentation. It was found that apoptotic cell death generally began 6～12 h after the initiating insult, and reached the peak at 24 h in the cortex and hippocampus. Conclusions Cerebral HI can induce neuronal apoptosis in neonatal rats. Apoptosis is mainly involved in the delayed cell death following hypoxic ischemic injury except for the existence of necrosis. Apoptosis appears to be a major form of delayed cell death during the selective neuronal loss following hypoxic ischemic injury. Owing to the limitation of different techniques, it is necessary to adopt several ways to determine the existence of apoptosis.