Fig. 5. Examples of atypical interconnected mitochondrial networks. (A) HUVEC cell loaded with TMRE was imaged with moderate laser intensity to trigger depolarisation of mitochondria by spontaneous PTP activation. This unusual HUVEC cell showed synchronous depolarisation across a large mitochondrial network. At 48 s (Ai) the mitochondria in the cell are all polarised, at 52 s (Aii) a large electrically continuous area was seen to depolarise, and 8 s later (Aiii) it had repolarised. After 120 s (Aiv) of imaging, this large electrically continuous mitochondrion depolarised again and did not recover during the experiment (5 min). (B) A cortical astrocyte that was found to have lumenally continuous mitochondria using FRAP. (Bii–iv) The region of the cell bounded by the dashed box in Bi on an expanded scale. The cell was imaged before (Bi,ii), approximately 10 s (Biii) and 120 s after (Biv) after bleaching a small area of the mitochondrion (the bleached region is denoted by the solid white box). During the photobleach process, the laser intensity was increased and sufficient irradiation was applied to reduce the fluorescence in the bleached area to zero. Despite this complete bleach, the fluorescence can already be seen to have partially recovered 10 s later and fully equilibrated after 120 s (Biv). Scale bar, 5 µm.