REGULATORY VOLUME DECREASE IN A RENAL DISTAL TUBULAR CELL-LINE (A6) .2. EFFECT OF NA+ TRANSPORT RATE

A6 epithelia, a cell line originating from the distal tubular part of the kidney of Xenopsus laevis, were cultured on permeable supports and mounted in an Ussing-type chamber. Cell thickness (T-c), short-circui t current (I-x) and transepithelial conductance (G(t)) were recorded w hile tissues were bilaterally incubated in NaCl solutions and the tran sepithelial potential was clamped to zero. Effects of inhibition and s timulation of transepithelial Na+ transport on cell volume and on its regulation during a hyposmotic challenge were investi igated. Under co ntrol conditions a slow spontaneous decrease of T-c described by a lin ear baseline was recorded. The reduction of the apical osmolality from 260 to 140 mosmol/kg did not alter cell volume significantly, demonst rating a negligible water permeability of the apical barrier. The inhi bition of Naf uptake by replacing apical Na+ by N-methyl-D-glucamine ( NMDG(+)) did not affect cell volume under isotonic conditions. An incr ease of T-c by 12.1% above the control baseline was recorded after blo cking active transport with ouabain for 60 min. The activation of Natransport with insulin or oxytocin, which is known to activate the api cal water permeability in other epithelia, did not alter cell volume s ignificantly. The insensitivity of cell volume to alterations in apica l Na+ uptake or Na+ pump rate confirms the close coupling between apic al and basolateral transport processes. The blockage of basolateral K channels by 5 mM Ba2+ elicited a significant increase in T-c of 16.3% above control. Quinine, a potent blocker of volume-activated K+ chann els, did not change T-c significantly. Basolateral hypotonicity elicit ed a rapid rise in T-c followed by a regulatory volume decrease (RVD). An RVD was also recorded after blocking apical Na+ uptake as well as after stimulating apical Na+ uptake with oxytocin or insulin. Inhibiti on of active transport with ouabain as well as blocking K+ efflux at t he basolateral side with Ba2+ or quinine abolished the RVD. The inhibi tion of the RVD by ouabain seems to be caused by a depletion of cellul ar K+, whereas the effects of Ba2+ and quinine are most likely due to the blockage of the basolateral K+ pathway.