Photoinactivation of sodium-potassium-chloride cotransport in LLC-PK1/Cl 4 cells by bumetanide.
Rb+ uptake into LLC-PK1/Cl 4 cells can be subdivided into three components: 1) ouabain-sensitive uptake, 2) bumetanide-sensitive uptake, and 3) ouabain- and bumetanide-insensitive uptake. Exposure of cells to near-UV light in the presence of low concentrations of bumetanide produces a specific, irreversible inhibition of the bumetanide-sensitive uptake component, while not affecting the other two uptake components. Irreversible inhibition of bumetanide-sensitive transport is observed when measuring either cellular uptake or efflux and also when measuring 86Rb+ uptake into membrane vesicles. The irreversible inhibition is both concentration and time dependent and is blocked under conditions where the interaction of bumetanide with the Na+-K+-Cl- cotransporter is disturbed. We conclude that bumetanide, at low concentrations, can specifically and irreversibly inhibit the Na+-K+-Cl- cotransporter of LLC-PK1/Cl 4 cells. We suggest that this irreversible inhibition is the result of the photoactivation of an ether linkage in the bumetanide molecule, leading to a covalent binding of bumetanide to the Na+-K+-Cl- cotransporter.
D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D002034:Bumetanide / Q000494:pharmacology* / Q000528:radiation effects; D002352:Carrier Proteins / Q000037:antagonists & inhibitors*; D002460:Cell Line; D004232:Diuretics / Q000494:pharmacology*; D004848:Epithelium / Q000378:metabolism; D007668:Kidney; D007700:Kinetics; D010042:Ouabain / Q000494:pharmacology; D010777:Photochemistry; D011868:Radioisotopes; D012413:Rubidium / Q000378:metabolism; D028021:Sodium-Potassium-Chloride Symporters; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013552:Swine; D014466:Ultraviolet Rays
D002352:Carrier Proteins; D004232:Diuretics; D011868:Radioisotopes; D028021:Sodium-Potassium-Chloride Symporters; D002034:Bumetanide; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; D012413:Rubidium