Non-electrolyte transport across renal proximal tubule cell membranes measured by tracer efflux and light scattering

Abstract

Non-electrolyte transport in brush border membrane vesicles (BBMV), basolateral membrane vesicles (BLMV) and in viable cells isolated from the proximal convoluted tubule (PCT) of the rabbit kidney were measured by rapid filtration and stopped-flow light scattering techniques. Efflux of tracer solute was measured by loading packed vesicles or cells with 14C solute, diluting into nonradioactive buffer and filtering rapidly at varying incubation times. In BBMV at 23 degrees C, [14C-urea] decreased exponentially with time constant 3.2 +/- 0.3 s (S.D., n = 5) corresponding to a permeability coefficient (Purea) of 1.6 X 10(-6) cm/s, assuming a BBMV surface-to-volume ratio of 2 X 10(5) cm-1. Purea decreased to 7 X 10(-7) cm/s in the presence of 20 mM phenylurea. Tracer efflux determinations of BBMV Purea (1.6 X 10(-6) cm/s) and Pglycerol (0.6 X 10(-6) cm/s), and BLMV Purea (1.8 X 10(-6) cm/s) and Pthiourea (2.5 X 10(-6) cm/s) were in excellent agreement with Ps values determined by stopped-flow light scattering, where the time course of vesicle volume (linearly related to scattered light intensity) was measured in response to 100 mM outwardly directed solute gradients. These results establish accurate Ps value in brush border and basolateral membranes and support the application of light scattering to measure Ps in vesicles. In PCT cells however, there were systematic differences in urea and thiourea transport measured by tracer efflux and light scattering, indicating the potential difficulties in applying light scattering to Ps measurements in complex cell systems.