Role of mitochondria in serum withdrawal-induced apoptosis of immortalized neuronal precursors.
The intracellular mechanisms controlling apoptosis in immature neurons are still largely unknown. Taking immortalized hippocampal neuronal precursors (mouse cell line HN9.10e) as a model, we have analyzed the cellular events associated to apoptosis induced by serum deprivation. We observed translocation of Bax from cytosol to mitochondria after 1 h of serum withdrawal followed, 2 h later, by cytochrome c release from mitochondria. These events occurred without mitochondrial membrane potential loss nor mitochondrial calcium raise. As calcium is implicated in several cell death pathways, we analyzed intracellular calcium levels after longer periods of serum deprivation. After 6 h, an increase of cytosolic Ca2+ was detected in HN9.10e cells loaded with the Ca2+ indicator Fluo3-AM. This increase of calcium preceded morphological signs of apoptosis such as cell shrinkage and nuclear fragmentation, and was followed by a more pronounced raise that persisted until the terminal phases of the apoptotic process. Cells serum-deprived for 4 h and then grown in complete medium for 20 h fully recovered viability. Summarizing, in HN9.10e cells, calcium deregulation occurs in the late phases of apoptosis; earlier events involve translocation of Bax, release of cytochrome c, and maintenance of mitochondrial functionality. This allows an enlargement of the temporal window in which commitment to death is reversible.
Colombaioni
Laura
L
Institute of Neurophysiology of CNR, Italian National Research Council, Area della Ricerca CNR, Via G. Moruzzi, 1, Pisa, Italy. laura.colombaioni@in.pi.cnr.it
Colombini
Luisella
L
Garcia-Gil
Mercedes
M
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002461:Cell Line, Transformed; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D003574:Cytochrome c Group / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D008564:Membrane Potentials; D051379:Mice; D008928:Mitochondria / Q000502:physiology*; D009474:Neurons / Q000187:drug effects* / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D013234:Stem Cells / Q000187:drug effects* / Q000502:physiology*; D051028:bcl-2-Associated X Protein
C494827:Bax protein, mouse; D016895:Culture Media, Serum-Free; D003574:Cytochrome c Group; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D002118:Calcium