Fig. 5. CAI depolarizes the mitochondrial inner-membrane potential in a concentration-dependent manner. (A) Digital images of TMRE fluorescence in a single HEK-293 cell 60 seconds prior to (i) and 10 seconds following (ii) FCCP application (indicated by bar). (iii) Line traces of intensity changes from mitochondria indicated by white chevrons in (i). Note the spontaneous changes in _m and loss of fluorescence on depolarization when TMRE was loaded at low concentration. (B) Conversely, following high concentration loading of TMRE (i), FCCP application induced an increase in fluorescence in a small cluster of HEK-293 cells (ii); a response to depolarization and unquenching of the fluorophore as dye redistributed to the cytosol. (iii) Averaged intensity trace of rises in fluorescence measured in individual cells. (C) Average fluorescence intensity changes of high concentration TMRE-loaded cells treated with vehicle only (filled symbols) or treated with 10 µM CAI (open symbols). Cell suspensions were analyzed using a multi-well fluorescence plate reader. The break in the records indicates application of the drug. Bars, 10 µm.