Fig. 9. Transient openings of the mPTP were innocuous: prolonged openings caused necrotic cell death. TMRE loaded cells were imaged using either a 1% neutral density filter, sufficient to image reversible mPTP openings, or a 30% filter, to induce global mitochondrial depolarisation. After a period of imaging, cells were returned to the incubator for 4 hours. They were then exposed to propidium iodide (PI) and Hoechst 33342 (A) in order to identify necrotic or apoptotic nuclei. PI stains damaged or necrotic cells (red arrow). Hoechst stains all nuclei (blue arrow), but apoptotic nuclei display a characteristic bright, condensed staining (not shown). Bar, 10 µm. Transient openings of the mPTP caused no increase in cell death, either by apoptosis or necrosis (B); however, global and complete mitochondrial depolarisation caused a significant increase in necrotic staining (P=0.0002). No sign of apoptotic morphology or Hoechst staining was apparent.

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