DNA fragmentation is a biochemical hallmark of programmed cell death under developing and pathological conditions. The aim of this study is to examine hypoxia-induced programmed cell death in hypoxic fetal guinea pig brain after a period of one-day and three-day following hypoxia. The time course and localization of DNA fragmentation in fetal guinea pig brain were assessed by means of the terminal transferase-mediated biotin dUTP nick end labeling(TUNEL). Pregnant guinea pigs of 58 days gestation were divided into three groups: normoxic, one day after hypoxia and three days after hypoxia, and exposed to either 21% or 7% oxygen for 60 minutes. The fetal brains were obtained and preserved in 4% paraformaldehyde. Cerebral cortex, hippocampus, thalamus, cerebellum, and brain stem areas were evaluated for DNA fragmentation and morphologic characteristics of apoptotic cell death using TUNEL stain. Apoptotic cells were defined as cells with shrunken dark nucleus, large nucleus/cytoplasm ratio, and/or DNA fragmentation. Non-apoptotic cells were defined as cells with shrunken nucleus, however, the size of the nucleus was not so small and dark, the nucleus/cytoplasm ratio was not as large as in apoptotic cells. TUNEL-positive cells were detected in all above areas. There was no difference in cell number between control and hypoxic groups in all areas except in cerebellum there was a significant increase of TUNEL positive non-apoptotic cells (22.86 cells/μm ± 6.74 cells/μm vs. 1.09 cells/μm ± 0.23 cells/μm, p<0.05) in the group of one day after hypoxia. The number of TUNEL positive non-apoptotic cells returned to a level close to that of normoxic group after three days. The results show that one-day after hypoxia, there was DNA fragmentation in all the areas described above however, the DNA fragmentation reached to maximal degree in cerebellum. We conclude that in fetal guinea pig brain, one day after a hypoxic insult, a significant number of cerebral cells are TUNEL positive and are undergoing programmed cell death.