Amiloride sensitivity of proton-conductive pathways in gastric and intestinal apical membrane vesicles.
Passive proton permeability of gastrointestinal apical membrane vesicles was determined. The nature of the pathways for proton permeation was investigated using amiloride. The rate of proton permeation (kH+) was determined by addition of vesicles (pHi = 6.5) to a pH 8.0 solution containing acridine orange. The rate of recovery of acridine orange fluorescence after quenching by the acidic vesicles ranged from 4 x 10(-3) (gastric parietal cell stimulation-associated vesicles; SAV) and 5 x 10(-3) (duodenal brush-border membrane vesicles; dBBMV) to 11 x 10(-3) sec-1 (ileal BBMV; iBBMV). Amiloride, 0.03 and 0.1 mM, significantly reduced the rate of proton permeation in dBBMV and iBBMV, but not gastric SAV. The decreases in kH+ were proportionately greater in iBBMV as compared with dBBMV. The presence of Na+/H+ exchange was demonstrated in both dBBMV and iBBMV by proton-driven (pHi less than pHo) 22Na+ uptake. Evidence was also sought for the conductive nature of pathways for proton permeation. Intravesicular acidification, again determined by quenching of acridine orange fluorescence, was observed during imposition of K(+)-diffusion potential ([K+]i much much greater than [K+]o). In dBBMV and iBBMV, intravesicular acidification was enhanced in the presence of the K(+)-ionophore valinomycin, indicating that the native K+ permeability is rate limiting. In the presence of valinomycin, the K(+)-diffusion potential drove BBMV intravesicular acidification to levels close to the electrochemical potential. In gastric SAV, acidification was not limited by the K+ permeability. Valinomycin was without effect, but the K+/H+ ionophore nigericin enhanced acidification in gastric SAV, illustrating the low proton permeability of these membranes.(ABSTRACT TRUNCATED AT 250 WORDS)
Wilkes
J M
JM
Department of Physiological Sciences, University of Newcastle upon Tyne, Medical School, England.
Hirst
B H
BH
D000584:Amiloride / Q000494:pharmacology*; D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects; D004058:Diffusion; D004553:Electric Conductivity; D005753:Gastric Mucosa / Q000187:drug effects / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008564:Membrane Potentials; D011522:Protons; D011817:Rabbits; D012964:Sodium / Q000378:metabolism
D011522:Protons; D000584:Amiloride; D012964:Sodium