HIGH OXYGEN ATMOSPHERE FOR NEURONAL CELL-CULTURE WITH NERVE GROWTH-FACTOR .2. SURVIVAL AND GROWTH OF CLONAL RAT PHEOCHROMOCYTOMA PC12H CELLS

When clonal rat pheochromocytoma PC12h cells were cultured in a 50% O2 atmosphere, cells gradually died during the cultivation. On the other hand, the addition of NGF at the final concentration of 50 ng/ml could rescue the cells from death. The culture in a 40% O2 atmosphere had little effect on the growth of PC12h cells, as compared with the culture in a normal 20% O2 condition. A very high O2 concentration, as 60%, caused severe damage to PC12h cell growth, and the restoration of cell growth by NGF seemed to be insufficient. PC12h cells were fully differentiated and extended dense long neurities by NGF even in a 50% O2 atmosphere. However, the neurite extension in the culture in a 60% O2 atmosphere was suppressed. The cell-saving effect of NGF on cell death in culture under a 50% O2 atmosphere was dose-dependent, and the ED50 value of NGF was 5 ng/ml. Basic fibroblast growth factor and epidermal growth factor also had a potent effect to rescue the cell death in the high O2 culture, but insulin had no effect. Since the differentiation effects of NGF on PC12h cells are thought to offer a model system to investigate the effect of NGF on neurons, the present observations suggest that a protection machinery for high O2 toxicity to neurons may exist in the neuronal differentiated PC12h cells by NGF, but not in the undifferentiated cells.