Fig. 6

Mitochondrial damage and cytochrome c translocation caused by colchicine is not prevented by zVAD-fmk. (A) Neurons were treated with colchicine (1 µM) in the absence or presence of N-benzyloxycarbonyl-Val-Ala-aspartyl-fluoromethyl ketone (zVAD-fmk; 100 µM). After 16 hr, cells were fixed with 2.5% glutaraldehyde for transmission electron microscopy analysis. Arrows indicate blebbing of mitochondria, and arrowheads indicate rupture of the outer mitochondrial membrane. Neurons were exposed to colchicine (1 µM), either directly, or in the presence of zVAD-fmk (100 µM), or in the presence of cycloheximide (CHX; 1 µM). (B) Cells were fixed with 4% paraformaldehyde and used for cytochrome c immunocytochemistry. Nuclei were stained with H-33342 and correspond to the same field used for imaging cytochrome c staining. Neurons with released cytochrome c are indicated by arrows and neurons that entirely lost cytochrome c staining are indicated by arrowheads. The width of every scale bar corresponds to 30 µm. (C) Individual neuronal somata marked by the boxes in (B) are shown at higher magnification (width = 10 µm). cyt c, cytochrome c.