Cell Density-Dependent Death Mode Switch of Cultured Cortical Neurons Under Serum-Free Starvation Stress

1. Cell death mode switch of cortical neurons from E17 rats was studied. Cells rapidly died under the serum-free condition. The time-course of cell death was markedly delayed by increasing cell density for primary culture in the trypan blue exclusion, LDH release, and MTT assays.2. By analyzing cell death by the use of double staining using PI/TUNEL and PI/Annexin V combinations, the mode in the low density culture was found to be necrosis, while that in the high density culture was apoptosis.3. The intracellular ATP level after the start of serum-free culture rapidly decline to 25% of 0-time level in the low density culture, but it was 60% in the high density culture. Both oligomycin and zVAD-fmk markedly decreased ATP levels and the population of TUNEL-positive neurons, while 3-aminobenzamide slightly increased these indices.4. Thus, it is strongly suggested that the cell death mode switch from necrosis to apoptosis is closely related to intracellular ATP levels, and some conditioned medium factors observed in the high density culture may affect both ATP level and cell death mode switch.