Vasopressin-stimulated chloride transport in transimmortalized mouse cell lines derived from the distal convoluted tubule and cortical and inner medullary collecting ducts

Abstract

Background. The fine control of NaCl absorption takes place in the distal parts of the renal tubule, but the regulation of Cl- transport in this region has not been fully elucidated. We have analysed the effects of dD-arginine vasopressin (dDAVP) on Cl- fluxes in cultured mouse distal convoluted tubule (mpkDCT), cortical collecting duct (mpkCCD) and inner medullary collecting duct (mpkIMCD) cell lines. Methods. RT-PCR and Western blotting were used to detect the amiloride-sensitive sodium channel (ENaC) and cystic fibrosis transmembrane conductance regulator (CFTR) mRNAs and protein in cultured mpkDCT, mpkCCD and mpkIMCD cells. Cl- fluxes were analysed by measuring the short-circuit current (Isc) and bidirectional 36Cl- fluxes on confluent cells grown on filters. Results. All three cell lines expressed ENaC and CFTR and had Isc stimulated by dDAVP. The rise in Isc caused by dDAVP (10-8 M) was inhibited by amiloride, and to a lesser extent by 5-nitro-2 -(3-phenylpropylamino)-benzoic acid (NPPB) in all three cell lines. The dDAVP-dependent Isc measured under apical Na+-free condition was reduced by Cl- channel blockers with a profile (NPPB>gliben-clamide>DIDS), similar to that for rat CFTR. dDAVP stimulated the apical-to-basal 36Cl- flux and to a lesser extent the basal-to-apical 36Cl- flux under open-circuit condition in all three cultured cell lines. Adding NPPB to the apical side reduced the basal-to-apical 36Cl- flux but not the opposite 36Cl- flux from dDAVP-treated cells. Conclusion. These results indicate that dDAVP stimulates the bi-directional flux of Cl-, resulting in net Cl- absorption, in these cultured mouse distal and collecting duct cells. Isc experiments also suggest the presence of a minor component of electrogenic Cl- secretion, possibly mediated by CFTR.