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-circuit current (I-x) and transepithelial conductance (G(t)) were recorded while tissues were bilaterally incubated in NaCl solutions and the transepithelial potential was clamped to zero. Effects of inhibition and stimulation of transepithelial Na+ transport on cell volume and on its regulation during a hyposmotic challenge were investi igated. Under control conditions a slow spontaneous decrease of T-c described by a linear baseline was recorded. The reduction of the apical osmolality from 260 to 140 mosmol/kg did not alter cell volume significantly, demonstrating a negligible water permeability of the apical barrier. The inhibition of Naf uptake by replacing apical Na+ by N-methyl-D-glucamine (NMDG(+)) did not affect cell volume under isotonic conditions. An increase of T-c by 12.1% above the control baseline was recorded after blocking active transport with ouabain for 60 min. The activation of Na+ transport with insulin or oxytocin, which is known to activate the apical water permeability in other epithelia, did not alter cell volume significantly. The insensitivity of cell volume to alterations in apical Na+ uptake or Na+ pump rate confirms the close coupling between apical 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+ channels, did not change T-c significantly. Basolateral hypotonicity elicited 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. Inhibition of active transport with ouabain as well as blocking K+ efflux at the basolateral side with Ba2+ or quinine abolished the RVD. The inhibition of the RVD by ouabain seems to be caused by a depletion of cellular K+, whereas the effects of Ba2+ and quinine are most likely due to the blockage of the basolateral K+ pathway.