To determine if calcium could release Cytochrome c (Cyt c) from brain mitochondria without activating the permeability transition (mPT), brain mitochondria were prepared in two different ways. Digitonin was used to lyse synaptosomes and release synaptosomal mitochondria or a Percoll gradient was used to separate non-synaptosomal mitochondria from the synaptosomes. In gradient-purified mitochondria, low levels of added digitonin produced swelling and Cyt c release. Digitonin augmented Ca2+-induced Cyt c release that was insensitive to the mPT inhibitors, cyclosporin A CsA and ADP. Similarly, in mitochondria prepared with digitonin, these inhibitors also failed to prevent Ca2+-induced Cyt c release. Thus the mPT-independent, Ca2+-induced Cyt c release pathway was attributable to alteration of the permeability properties of the outer mitochondrial membrane by digitonin.
Bibliographical noteFunding Information:
We would like to thank Dr Calvin Roff of R&D Systems for his support. This work was funded by the HDSA, NINDS 39414 to J.M.D., AHA 9804691X and 0130497Z to N.B., and NIH 1R21-AI45090 to R.J.
- Brain mitochondria
- Cytochrome c
- Permeability transition