Transport and regulatory properties of the apical Na-K-2Cl cotransporter of macula densa cells.
NH+4/NH3 fluxes were used to probe apical Na-K-2Cl transport activity of macula densa (MD) cells from rabbit kidney. In the presence of 25 mM NaCl and 5 mM Ba2+, addition of 20 mM NH+4 to the lumen produced a profound intracellular acidification, and approximately 80% of the initial acidification rate was bumetanide sensitive. The NH+4-induced acidification rate was dependent on luminal Cl- and Na+ with apparent affinities of 17 +/- 4 mM (Hill number 1.45) and 1.0 +/- 0.3 mM, respectively. In the presence of saturating luminal NaCl concentration ([NaCl]L), blockade of basolateral Cl- efflux with 10 microM 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) reduced the NH+4-induced acidification rate by 51 +/- 6% (P > 0.01, n = 5). Under similar conditions, dibutyryl-cAMP (DBcAMP) + forskolin increased the NH+4-induced acidification rate by 27%, whereas it produced no detectable effect at low luminal NaCl concentration. Most of the observed DBcAMP + forskolin effect was probably due to the stimulation of the basolateral Cl- conductance, since, in the presence of basolateral NPPB, this activation was changed to a 17.1% and 16.6% inhibition of the NH+4-induced acidification rate observed at high or low [NaCl]L, respectively. We conclude that the cotransporter found in MD cells displays, with respect to other Na-K-2Cl cotransporters, a relatively high affinity for luminal Na+ and luminal Cl- and can be specifically inhibited by increases in intracellular Cl- and cAMP concentrations.
Laamarti
M A
MA
Groupe de Recherche en Transport Membranaire, Université de Montréal, Montreal, Quebec, Canada H3C 3J7.
Bell
P D
PD
Lapointe
J Y
JY
D000818:Animals; D003994:Bucladesine / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000378:metabolism*; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D011817:Rabbits; D028021:Sodium-Potassium-Chloride Symporters
D002352:Carrier Proteins; D000644:Quaternary Ammonium Compounds; D028021:Sodium-Potassium-Chloride Symporters; D003994:Bucladesine