Activation of Na:2Cl:K cotransport by luminal chloride in macula densa cells.
Changes in macula densa intracellular pH (pHi) were used to monitor the direction of flux mediated by the apical Na:2Cl:K cotransporter. At the macula densa, a decrease in luminal [Cl] ([Cl]1) from 60 to 1 mM produced cellular alkalinization secondary to a cascade of events involving a decrease in apical Na:2Cl:K cotransport, a fall in intracellular [Na] ([Na]i) and a stimulation of Na:H exchange. This is supported by the fact that 97% of the change in macula densa pHi with reduction in [Cl]1 was bumetanide-sensitive whereas 92% of this pH change was amiloride-sensitive. We found that, in the presence of 20 mM Na and 5 mM K, a [Cl]1 of 14.3 +/- 2.4 mM (N = 7) produced equilibrium of the apical cotransporter since the pHi obtained under this condition was identical to the pHi found after reducing the net ionic flux to zero with bumetanide. Using this value together with the expected stoichiometry for the bumetanide-sensitive cotransporter, it was estimated that the intracellular [Cl] ([Cl]i) at equilibrium (or in the presence of bumetanide) could be as low as 5 mM. Also, using a Hill number of 2 which is consistent with the present data, the affinity for [Cl]1 was found to be 32.5 mM. Under physiological luminal conditions prevailing at the end of the thick ascending limb (approximately 3.5 mM K, and approximately 25 to 30 mM NaCl), macula densa cells are probably operating close to equilibrium while maintaining a small net reabsorption of Na/K and Cl. Since macula densa cells appear capable of reducing [Cl]i to very low levels, a reabsorptive flux should continue to occur until [NaCl]1 is reduced to 18 mM.
Lapointe
J Y
JY
Groupe de Recherche en Transport Membranaire, Université de Montréal, Québec, Canada.
Laamarti
A
A
Hurst
A M
AM
Fowler
B C
BC
Bell
P D
PD
D000818:Animals; D002034:Bumetanide / Q000494:pharmacology; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D003494:Cyclamates / Q000494:pharmacology; D005260:Female; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007424:Intracellular Fluid / Q000378:metabolism*; D017136:Ion Transport / Q000187:drug effects; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism*; D011817:Rabbits; D012964:Sodium / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000187:drug effects / Q000378:metabolism; D028021:Sodium-Potassium-Chloride Symporters
D002352:Carrier Proteins; D002712:Chlorides; D003494:Cyclamates; D017923:Sodium-Hydrogen Exchangers; D028021:Sodium-Potassium-Chloride Symporters; D002034:Bumetanide; D012964:Sodium; D011188:Potassium