Na+ and H+ transport in human jejunal brush-border membrane vesicles.
We have examined pH gradient-driven Na+ uptake and Na+-driven H+ transport in brush-border membrane vesicles prepared from jejunal tissue obtained from organ donors by measuring the influx of 22Na and the fluorescence quenching of acridine orange (AO). Vesicle preparation by either Ca2+ or Mg2+ precipitation showed no difference in 22Na uptake or AO fluorescence quenching and dissipation. An outward H+ gradient [intravesicular pH (pHi) 5.5; extravesicular pH (pHo) 7.5] induced a Na+ uptake "overshoot" of threefold over equilibrium, whereas the absence of an H+ gradient (at either pH 5.5 or 7.5) did not produce an overshoot. Voltage clamping by Ki+ = Ko+ plus valinomycin reduced the overshoot by 50%. The initial rate of pH-driven Na+ uptake in voltage-clamped vesicles was related to [Nao+] (Km = 29 mM and Vmax = 9.5 nmol.mg protein-1.3 s-1). Amiloride inhibited this uptake in voltage-clamped vesicles (Ki = 99 microM). Dissipation of AO fluorescence quench in vesicles with a preformed internal acid gradient was hastened by Nao+ as well as voltage clamping in the absence of Na+. In vesicles without a pH gradient, internal Na+, as well as a diffusion potential (Ki+ 100; Ko+ 0 plus valinomycin) in the absence of Na+, induced AO quenching. External Na+ and Li+, but not choline, acted to dissipate AO quenching induced by a diffusion potential, and the rate of dissipation was unaffected by the presence of Cl-.Li+ and NH4+, but not Cs+, K+, Rb+, or choline+, inhibited pH gradient-driven 22Na uptake. We conclude that human jejunal brush-border membrane vesicles contain conductive pathways for both Na+ and H+ and an Na+-H+ exchanger.
Kleinman
J G
JG
Nephrology Section, Zablocki Veterans Administration Medical Center, Milwaukee, Wisconsin.
Harig
J M
JM
Barry
J A
JA
Ramaswamy
K
K
D000165:Acridine Orange; D000584:Amiloride / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007583:Jejunum / Q000378:metabolism*; D007700:Kinetics; D008871:Microvilli / Q000187:drug effects / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers; D013050:Spectrometry, Fluorescence; D014634:Valinomycin / Q000494:pharmacology
D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D014634:Valinomycin; D000584:Amiloride; D012964:Sodium; D000165:Acridine Orange; D011188:Potassium