A NEW CHROMATOGRAPHIC METHOD FOR MEASUREMENT OF RUBIDIUM TRANSPORT ACTIVITIES IN CULTURED BOVINE RETINAL-PIGMENT EPITHELIAL-CELLS

A new method for measuring cellular rubidium (Rb+) uptake activities b ased on cation chromatography was developed and compared with the stan dard technique, uptake of the radioisotope Rb-86(+), using cultured bo vine retinal pigment epithelial (RPE) cells, The Rb+ response was stri ctly linear from 0.25 nmol(detection limit) to 25 nmol. The Na+/K+-ATP ase inhibitor ouabain inhibited Rb+ uptake with IC50 values of 128.7 /- 23.5 nM (n = 8; radioactive method) and 56.6 +/- 9.3 nM (n = 9; non -radioactive method, p < 0.01), The latter value is identical to the I C50 value of 54.4 +/- 16.2 nM (n = 3) for ouabain binding to the intac t RPE cells. Ouabain and bumetanide, an inhibitor of the Na+/K+/Cl--co transporter, each inhibited Rb+ uptake maximally by 66 and 30%, respec tively, This new technique allows sensitive measurement of intracellul ar Rb+, as well as K+ and Na+, and, thus, should prove useful for stud ying the effects of pharmacologic agents and simulated disease conditi ons on cation transport and cation balance in RPE and other cell types .