Concentration dependence of the chloride selfexchange and homoexchange fluxes in human red cell ghosts

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Abstract
The concentration dependence of the unidirectional chloride flux in human red cell ghosts was studied under selfexchange and under homoexchange conditions. Under selfexchange conditions the intracellular concentration of chloride [Cl]in is equal to the extracellular concentration [Cl]ex and [Cl]in and [Cl]ex are raised concomitantly. Under homoexchange conditions [Cl]in or [Cl]ex were varied separately at a fixed trans-concentration of chloride. The chloride fluxes were calculated from the rate of the tracer efflux and the intracellular chloride. All experiments were executed in isotonic (330 mosM) KCl/K-citrate/sorbitol solutions containing 0–100 mM KCl, 40 mM K-citrate and different concentrations of sorbitol for isoosmotic substitution. The chloride selfexchange and the chloride homoexchange fluxes exhibit a pure saturation kinetics. The halfsaturation constant for the chloride selfexchange was ∼ 20 mM, the maximal flux was approx. 3.5·10−4 mol/(min g cells). The apparent chloride halfsaturation constants from the homoexchange experiments were in the range of 0.9–4.5 mM for the outer and of 5.5–14.5 mM (0° C, pH 7.3) for the inner membrane surface, both halfsaturation constants increase with increasing trans-concentrations. At infinite trans-concentrations of chloride, the halfsaturation constant for the outer and the inner membrane surface amounts to ∼ 5 mM and ∼ 15 mM, respectively. The slope of the double reciprocal plots of flux versus cis-chloride concentration decreases with increasing trans-concentration of chloride. The kinetics of the chloride transport provides evidence for a carrier mediated transport mechanism with a single reciprocating transport site. The translocation of the loaded carrier appears to be much faster than the translocation of the unloaded carrier. At low concentrations of chloride, however, the slippage of the empty carrier seems to contribute to the back-diffusion of carrier to the cis-side of the membrane.

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