Zinc transport and metallothionein induction in primary cultures of rabbit kidney proximal cells

Abstract

Primary cultures of isolated rabbit renal proximal cells were grown on collagen-coated permeable supports. The confluent epithelia were polarized, making possible the measurement of uptakes and effluxes across the apical and the basolateral membranes. Uptakes of 65Zn were assessed under initial rate conditions, after 0.5 min incubation. The kinetic parameters of apical uptake were a J max of 25.1 ± 5.3 pmol min −1 (μ DNA) −1, a K m of 43.3 ± 7.3 μ M and an unsaturable constant of 0.105 ± 0.029 ( n = 7) at 37°C. Cadmium competitively inhibited the zinc uptake, with a K i value of 24.5 ± 7.3 μ M . Basolateral uptake was characterized by a high capacity ( J max = 227.9 ± 46.6 pmol min −1 (μg DNA) −1) and an affinity similar to that of the apical uptake ( K m = 35.4 ± 14.2 μ M ). Cadmium had no effect on the basolateral zinc uptake. Effluxes across the basolateral face of the epithelium always exceeded those across the apical face. Excess zinc in the culture medium induced the synthesis of metallothionein in the epithelia, as judged by the rate of [ 35S]cysteine incorporation into a fraction of cytosolic proteins. Metallothionein induction did not appear to modify the kinetic parameters of the apical zinc uptake. These data suggest that separate saturable transport systems are responsible for the apical and basolateral zinc uptakes in proximal renal cells. Induction of metallothioncin had no apparent effect on apical zinc uptake in this system.