In order to elucidate the mechanisms of photosensitized injury to mitochondria, two photosensitizers have been compared. Both doxycycline (DOTC) and rhodamine-123 (R123) localize preferentially within the mitochondria of MGH-U1 bladder carcinoma cells in vitro, and both sensitize phototoxic injury that is selective for mitochondria. Mitochondria of cells pretreated with DOTC and irradiated with UVA (1 J/cm2, 320-400 nm) undergo massive swelling that begins by 10 min after irradiation, is maximal by 1 h, and is partially repaired by 4 h; damage caused by exposure to a higher UVA dose (6 J/cm2), however, is not repaired. In contrast, cells pretreated with R123 and irradiated with an argon-ion laser (10 J/cm2, 514.5 nm) undergo a different type of mitochondrial injury, characterized by the delayed (4 h) onset of moderate mitochondrial swelling and striking mitochondrial distortion and fragmentation, which is not repaired by 48 h after irradiation. These differences indicated that the reactions underlying cellular photoxicity can be distinguished even on an ultrastructural level. Probably both the primary photochemistry and the submitochondrial targets of these reactions differ with the two photosensitizers.