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 based on cation chromatography was developed and compared with the standard technique, uptake of the radioisotope 86Rb+, using cultured bovine retinal pigment epithelial (RPE) cells. The Rb+ response was strictly linear from 0.25 n...

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Bibliographic Details
Published inOphthalmic research Vol. 29; no. 3; p. 117
Main Authors Crider, J Y, Williams, G W, Yorio, T, Sharif, N A, Griffin, B W
Format Journal Article
LanguageEnglish
Published Switzerland 01.01.1997
Subjects
Animals
Bumetanide - pharmacology
Cattle
Cell Membrane - drug effects
Cell Membrane - metabolism
Cells, Cultured
Chromatography, Ion Exchange - methods
Diuretics - pharmacology
Enzyme Inhibitors - pharmacology
Intracellular Fluid - metabolism
Ion Transport
Ouabain - pharmacology
Pigment Epithelium of Eye - cytology
Pigment Epithelium of Eye - drug effects
Pigment Epithelium of Eye - metabolism
Potassium - metabolism
Rubidium Radioisotopes - pharmacokinetics
Sodium - metabolism
Sodium-Potassium-Exchanging ATPase - antagonists & inhibitors
Sodium-Potassium-Exchanging ATPase - metabolism
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Summary:A new method for measuring cellular rubidium (Rb+) uptake activities based on cation chromatography was developed and compared with the standard technique, uptake of the radioisotope 86Rb+, using cultured bovine retinal pigment epithelial (RPE) cells. The Rb+ response was strictly linear from 0.25 nmol (detection limit) to 25 nmol. The Na+/K(+)-ATPase 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 IC50 value of 54.4 +/- 16.2 nM (n = 3) for ouabain binding to the intact RPE cells. Ouabain and bumetanide, an inhibitor of the Na+/K+/Cl(-)-cotransporter, each inhibited Rb+ uptake maximally by 66 and 30%, respectively. This new technique allows sensitive measurement of intracellular Rb+, as well as K+ and Na+, and, thus, should prove useful for studying the effects of pharmacologic agents and simulated disease conditions on cation transport and cation balance in RPE and other cell types.
ISSN:0030-3747
DOI:10.1159/000268005