deamino-arginine-vasopressin and 5-nitro-2-(3-phenylpropylamino)benzoic-acid

The role of the cystic fibrosis transmembrane conductance regulator (CFTR) in the renal cortical collecting duct (CCD) has not yet been fully elucidated. Here, we investigated the effects of deamino-8-D-arginine vasopressin (dDAVP) and isoproterenol (ISO) on NaCl transport in primary cultured CCDs microdissected from normal [CFTR(+/+)] and CFTR-knockout [CFTR(-/-)] mice. dDAVP stimulated the benzamyl amiloride (BAm)-sensitive transport of Na(+) assessed by the short-circuit current (I(sc)) method in both CFTR(+/+) and CFTR(-/-) CCDs to a very similar degree. Apical addition of 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) or glibenclamide partially inhibited the rise in I(sc) induced by dDAVP and ISO in BAm-treated CFTR(+/+) CCDs, whereas dDAVP, ISO, and NPPB did not alter I(sc) in BAm-treated CFTR(-/-) CCDs. dDAVP stimulated the apical-to-basal flux and, to a lesser extent, the basal-to-apical flux of (36)Cl(-) in CFTR(+/+) CCDs. dDAVP also increased the apical-to-basal (36)Cl(-) flux in CFTR(-/-) CCDs but not the basal-to-apical (36)Cl(-) flux. These results demonstrate that CFTR mediates the cAMP-stimulated component of secreted Cl(-) in mouse CCD.

Topics: Amiloride; Animals; Cells, Cultured; Chloride Channels; Chlorides; Crosses, Genetic; Cyclic AMP; Cystic Fibrosis Transmembrane Conductance Regulator; Deamino Arginine Vasopressin; Epithelial Sodium Channels; Glyburide; Homozygote; Isoproterenol; Kidney Cortex; Kidney Tubules, Collecting; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitrobenzoates; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sodium Channel Blockers; Sodium Channels; Sodium Chloride; Transcription, Genetic

The effects of aldosterone and vasopressin on Cl- transport were investigated in a mouse cortical collecting duct (mpkCCD) cell line derived from a transgenic mouse carrying the SV40 large T antigen driven by the proximal regulatory sequences of the L-pyruvate kinase gene. The cells had features of a tight epithelium and expressed the amiloride-sensitive sodium channel and the cystic fibrosis transmembrane conductance regulator (CFTR) genes. dD-arginine vasopressin (dDAVP) caused a rapid, dose-dependent, increase in short-circuit current (Isc). Experiments with ion channel blockers and apical ion substitution showed that the current represented amiloride-sensitive Na+ and 5-nitro-2-(3-phenylpropylamino)benzoate-sensitive and glibenclamide-sensitive Cl- fluxes. Aldosterone (5 x 10(-7)M for 3 or 24 hr) stimulated Isc and apical-to-basal 22Na+ flux by 3-fold. 36Cl- flux studies showed that dDAVP and aldosterone stimulated net Cl- reabsorption and that dDAVP potentiated the action of aldosterone on Cl- transport. Whereas aldosterone affected only the apical-to-basal 36Cl- flux, dDAVP mainly increased the apical-to-basal Cl- flux and the basal-to-apical flux of Cl- to a lesser extent. These results suggest that the discrete dDAVP-elicited Cl- secretion involves the CFTR and that dDAVP and aldosterone may affect in different ways the observed increased Cl- reabsorption in this model of mouse cultured cortical collecting duct cells.

Topics: Aldosterone; Amiloride; Animals; Cell Line, Transformed; Chloride Channels; Cystic Fibrosis Transmembrane Conductance Regulator; Deamino Arginine Vasopressin; Epithelial Sodium Channels; Gene Expression; Kidney Tubules, Collecting; Male; Mice; Mice, Transgenic; Nitrobenzoates; Sodium Channels; Sodium Radioisotopes; Vasopressins