Reduction of K+ efflux in cultured mouse fibroblasts, by mutation or by diuretics, permits growth in K+-deficient medium.
The mouse fibroblastic cell line LM(TK-) is unable to grow at external K+ concentrations below a threshold value of 0.4 mM. At subthreshold K+ concentrations, LM(TK-) cells rapidly lose intracellular K+ and eventually lyse. We have analyzed the pathway primarily responsible for K+ efflux under these experimental conditions and reports its specific inhibition by two diuretics, furosemide and bumetanide. Bumetanide, an analog of furosemide, was a more potent inhibitor (by several orders of magnitude) than was furosemide itself. The effects of ouabain and bumetanide were additive, suggesting independence of diuretic-sensitive K+ efflux from Na+/K+ pump-mediated fluxes. Characterization of K+ efflux in LTK-5, a mutant derived from LM(TK-) and selected for its ability to grow at 0.2 mM K+ indicated that the mutant had lost the diuretic-sensitive K+ efflux pathway. Net cation fluxes, steady-state intracellular cation concentrations, and growth at reduced K+ concentrations were comparable for LM(TK-) cells maximally inhibited by diuretics and for the LTK-5 mutant grown either in the presence or absence of diuretics. Thus, reduction in K+ efflux, either by diuretic addition diuretics. Thus, reduction in K+ efflux, either by diuretic addition or by genetic alteration, can permit the cell to maintain normal cation gradients and to grow at otherwise subthreshold external K+ concentrations.
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002034:Bumetanide / Q000494:pharmacology*; D004232:Diuretics / Q000494:pharmacology*; D005665:Furosemide / Q000494:pharmacology*; D007700:Kinetics; D007739:L Cells / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D009154:Mutation; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism
D004232:Diuretics; D002034:Bumetanide; D010042:Ouabain; D005665:Furosemide; D012964:Sodium; D011188:Potassium