Calcium-activated chloride currents in primary cultures of rabbit distal convoluted tubule.
Chloride (Cl-) conductances were studied in primary cultures of rabbit distal convoluted tubule (very early distal "bright" convoluted tubule, DCTb) by the whole cell patch-clamp technique. We identified a Cl- current activated by 2 microM extracellular ionomycin. The kinetics of the macroscopic current were time dependent for depolarizing potentials with a slow developing component. The steady state current presented outward rectification, and the ion selectivity sequence was I- > Br- > > Cl > glutamate. The current was inhibited by 0.1 mM 5-nitro-2-(3-phenylpropyl-amino)benzoic acid, 1 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, and 1 mM diphenylamine-2-carboxylate. To identify the location of the Cl- conductance, 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescence experiments were carried out in confluent cultures developed on collagen-coated permeable filters. Cl- removal from the apical solution induced a Cl- efflux that was stimulated by 10 microM forskolin. Forskolin had no effect on the basolateral Cl- permeability Cl- substitution in the basolateral solution induced an efflux stimulated by 2 microM ionomycin or 50 microM extracellular ATP Ionomycin had no effect on the apical Cl- fluxes. Thus cultured DCTb cells exhibit Ca(2+)-activated Cl- channels located in the basolateral membrane. This Cl- permeability was active at a resting membrane potential and could participate in the Cl- reabsorption across the DCTb in control conditions.
Unité de Recherche Associée Centre National de la Recherche Scientifique 1938, Université de Nice Sophia Antipolis, France.
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D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D002712:Chlorides / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D005576:Colforsin / Q000494:pharmacology; D004553:Electric Conductivity; D005456:Fluorescent Dyes; D007425:Intracellular Membranes / Q000378:metabolism; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000378:metabolism*; D008297:Male; D011806:Quinolinium Compounds; D011817:Rabbits
D002712:Chlorides; D005456:Fluorescent Dyes; D007476:Ionophores; D011806:Quinolinium Compounds; D005576:Colforsin; D015759:Ionomycin; C052335:6-methoxy-N-(3-sulfopropyl)quinolinium; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D002118:Calcium