TEMPORAL ANALYSIS OF EVENTS ASSOCIATED WITH PROGRAMMED CELL-DEATH (APOPTOSIS) OF SYMPATHETIC NEURONS DEPRIVED OF NERVE GROWTH-FACTOR

The time course of molecular events that accompany degeneration and de ath after nerve growth factor (NGF) deprivation and neuroprotection by NGF and other agents was examined in cultures of NGF-dependent neonat al rat sympathetic neurons and compared to death by apoptosis. Within 12 h after onset of NGF deprivation, glucose uptake, protein synthesis , and RNA synthesis fell precipitously followed by a moderate decrease of mitochondrial function. The molecular mechanisms underlying the NG F deprivation-induced decrease of protein synthesis and neuronal death were compared and found to be different, demonstrating that this decr ease of protein synthesis is insufficient to cause death subsequently. After these early changes and during the onset of neuronal atrophy, i nhibition of protein synthesis ceased to halt neuronal degeneration wh ile readdition of NGF or a cAMP analogue remained neuroprotective for 6 h. This suggests a model in which a putative killer protein reaches lethal levels several hours before the neurons cease to respond to rea ddition of NGF with survival and become committed to die. Preceding lo ss of viability by 5 h and concurrent with commitment to die, the neur onal DNA fragmented into oligonucleosomes. The temporal and pharmacolo gical characteristics of DNA fragmentation is consistent with DNA frag mentation being part of the mechanism that commits the neuron to die. The antimitotic and neurotoxin cytosine arabinoside induced DNA fragme ntation in the presence of NGF, supporting previous evidence that it m imicked NGF deprivation-induced death closely. Thus trophic factor dep rivation-induced death occurs by apoptosis and is an example of progra mmed cell death.