On the water and proton permeabilities across membranes from erythrocyte ghosts.
The diffusional permeability of water across membranes from bovine and human erythrocyte ghosts was measured by a recently developed method which is based on the different indices of refraction of H2O and 2H2O. Resealed erythrocyte ghosts were prepared by a gel-filtration technique. Pd (2H2O/H2O) values of 1.2 X 10(-3) cm/s (human) and 1.7 X 10(-3) cm/s (bovine) were calculated at 20 degrees C. The activation energies of the water exchange were 23.5 kJ/mol (human) and 25.4 kJ/mol (bovine). Treatment of the ghosts with p-chloromercuribenzenesulfonic acid (PCMBS) led to a 60-70% inhibition of the diffusional water exchange. The pH equilibration across membranes of erythrocyte ghosts was measured by intracellular carboxyfluorescein. The rates of proton flux after pH-jumps (pH 7.3 to pH 6.1) were about 100-fold lower than those of the water exchange and dependent on the kind of anions present (Cl-, NO-3, SO2-4). The activation energies of proton flux were 60-70 kJ/mol. 4,4'-Diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) inhibited the exchange by 97-98% and lowered the activation energy. The inhibitor of water exchange, PCMBS, increased the proton-permeation rate by a factor of 4-5. It is assumed that the rate-limiting step for the proton permeation is determined by the anion exchange. Under this condition our results are not in accord with one channel as a common pathway for both the passive water and anion transport.
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D002731:4-Chloromercuribenzenesulfonate / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism; D002417:Cattle; D002463:Cell Membrane Permeability / Q000187:drug effects; D002712:Chlorides / Q000378:metabolism; D004058:Diffusion; D004910:Erythrocyte Membrane / Q000502:physiology*; D005374:Filtration; D005452:Fluoresceins; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008433:Mathematics; D011188:Potassium / Q000378:metabolism; D011522:Protons; D012964:Sodium / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014867:Water
D001639:Bicarbonates; D002712:Chlorides; D005452:Fluoresceins; D011522:Protons; D014867:Water; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; C024098:6-carboxyfluorescein; D002731:4-Chloromercuribenzenesulfonate; D012964:Sodium; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D011188:Potassium