Effects of divalent cations and pH on amiloride-sensitive Na+ fluxes into luminal membrane vesicles from pars recta of rabbit proximal tubule.
The effect of Ca2+, Cd2+, Ba2+, Mg2+ and pH on the renal epithelial Na(+)-channel was investigated by measuring the amiloride-sensitive 22Na+ fluxes into luminal membrane vesicles from pars recta of rabbit proximal tubule. It was found that intravesicular Ca2+ as well as extravesicular Ca2+ substantially lowered the channel-mediated flux. Amiloride sensitive Na+ uptake was nearly completely blocked by 10 microM Ca2+ at pH 7.4. The inhibitory effect of Ca2+ was dependent on pH. Thus, 10 microM Ca2+ produced 90% inhibition of 22Na+ uptake at pH 7.4, and only 40% inhibition at pH 7.0. The tracer fluxes measured in the absence of Ca2+ were pH independent over the range from 7.0 to 7.4. All the cations Ca2+, Cd2+, Ba2+ except Mg2+ inhibited the 22Na+ influx drastically when added extravesicularly in millimolar concentrations. The cations Cd2+, Ba2+ and Mg2+ in the same concentrations intravesicularly inhibited the 22Na+ influx only slightly. A millimolar concentration of Ca2+ intravesicularly blocked the amiloride-sensitive 22Na+ flux completely. The data indicate that Ca2+ inhibits Na+ influx specifically by binding to sites composed of one or several deprotonated groups on the channel proteins.
Blokkebak-Poulsen
J
J
Institute of Medical Biochemistry, University of Aarhus, Denmark.
Sheikh
M I
MI
Jacobsen
C
C
D000584:Amiloride / Q000494:pharmacology*; D000818:Animals; D001485:Basement Membrane / Q000378:metabolism / Q000648:ultrastructure; D002118:Calcium / Q000494:pharmacology*; D002413:Cations, Divalent; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D004305:Dose-Response Relationship, Drug; D004848:Epithelium / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D011817:Rabbits; D012964:Sodium / Q000378:metabolism*; D015222:Sodium Channels / Q000187:drug effects* / Q000378:metabolism
D002413:Cations, Divalent; D015222:Sodium Channels; D000584:Amiloride; D012964:Sodium; D002118:Calcium