INTRACELLULAR ACCUMULATION OF RHODAMINE-110 IN SINGLE LIVING CELLS

To gain a better understanding of the internalization of rhodamines, v ital staining of living cells in situ by two different rhodamines, R11 0 and R123, was studied by microfluorometry. These dyes differ strongl y in their lipophilic properties because of differences in charge dist ribution. Microspectrofluorometry was used to study the fluorescence e mission spectra of R110-loaded cells to determine reliable loading con ditions. Cell uptake and cell efflux studies of R110 were performed by numerical microfluorescence imaging. A slower uptake was observed for R110 (14 hr) vs R123 (2 hr), but the R110 efflux was much more rapid (30 min) than that of R123 (>24 hr). Although it appeared in the R110 and R123 co-localization study that R110 was able to accumulate in mit ochondria, labeling with R110 was lower than with R123. Our results in dicate that, rhodamine 110 in its acid cationic form is able to cross the plasma and mitochondrial membrane and to accumulate in cell compar tments as does the cationic rhodamine 123. However, because of its aci do-basic properties, R110 should be able to decrease the pH of cell co mpartments, depending on their ability to regulate pH. In such a model , mitochondrial pH should be more greatly decreased than cytosolic pH, leading to a lower mitochondrial accumulation of R110 than of R123. S urprisingly, these effects, which should affect the energetic state of mitochondria, do not influence cell growth, because no cytotoxic effe ct was observed.