TY - JOUR
T1 - Laser micro-irradiation of mitochondria
T2 - Is there an amplified mitochondrial death signal in neural cells?
AU - Khodjakov, Alexey
AU - Rieder, Conly
AU - Mannella, Carmen A.
AU - Kinnally, Kathleen W.
N1 - Funding Information:
This research was supported by NIH grants GM57249 (KWK), GM59363 (AK), and GM40198 (CLR), and NSF grants MCB 0235834 and INT 003797 (KWK). Electron microscopy and image processing utilized the facilities of the Wadsworth Center's Resource for the Visualization of Biological Complexity (NIH/NCRR grant RR012019). We thank D. Bredesen (Buck Institute) for the CSM14.1 cell line, A.-L. Nieminen (Case Western) for the GFP–cytochrome c plasmid, and X. Wang (U Texas SW Med. Center) for the APAF-1 and SMAC. We thank D. Pietkiewicz, C. Hughes, R. Cole, G. Cassels and K. Buttle for their excellent technical assistance.
PY - 2004/3
Y1 - 2004/3
N2 - Several mitochondrial proteins, such as cytochrome c, are directly involved in the pathway for caspase activation following induction of apoptosis. Release of mitochondrial cytochrome c early in apoptosis is rapid and almost complete. Microinjection of cytochrome c into resting cells induces apoptosis, but the amount needed approaches the total cellular content. These observations suggest that mitochondrial protein release is an all-or-nothing process inside the cell and not an amplifiable apoptotic signal. To test this hypothesis, laser micro-irradiation was used to rupture membranes of individual mitochondria within living rat neural cells. Laser micro-irradiation caused swelling, fragmentation, depolarization, and cytochrome c depletion in targeted mitochondria. These effects were explained by correlative electron microscopic analysis showing local rupture of outer and inner membranes at the site of irradiation. In all cases, there were no detectable changes in the structure, membrane potential, or cytochrome c content of neighboring, non-irradiated organelles. Furthermore, irradiation of up to 15% of the mitochondria in a cell did not induce apoptosis. The results from these laser micro-irradiation experiments prove that local release of mitochondrial proteins does not constitute an amplifiable apoptotic signal in resting neural cells.
AB - Several mitochondrial proteins, such as cytochrome c, are directly involved in the pathway for caspase activation following induction of apoptosis. Release of mitochondrial cytochrome c early in apoptosis is rapid and almost complete. Microinjection of cytochrome c into resting cells induces apoptosis, but the amount needed approaches the total cellular content. These observations suggest that mitochondrial protein release is an all-or-nothing process inside the cell and not an amplifiable apoptotic signal. To test this hypothesis, laser micro-irradiation was used to rupture membranes of individual mitochondria within living rat neural cells. Laser micro-irradiation caused swelling, fragmentation, depolarization, and cytochrome c depletion in targeted mitochondria. These effects were explained by correlative electron microscopic analysis showing local rupture of outer and inner membranes at the site of irradiation. In all cases, there were no detectable changes in the structure, membrane potential, or cytochrome c content of neighboring, non-irradiated organelles. Furthermore, irradiation of up to 15% of the mitochondria in a cell did not induce apoptosis. The results from these laser micro-irradiation experiments prove that local release of mitochondrial proteins does not constitute an amplifiable apoptotic signal in resting neural cells.
KW - Ablation
KW - Apoptosis
KW - Cytochrome c
KW - Lasers
KW - Mitochondria
KW - Neural cells
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U2 - 10.1016/j.mito.2003.10.002
DO - 10.1016/j.mito.2003.10.002
M3 - Article
C2 - 16120356
AN - SCOPUS:1342303541
SN - 1567-7249
VL - 3
SP - 217
EP - 227
JO - Mitochondrion
JF - Mitochondrion
IS - 4
ER -