Corticosteroid-dependent sodium transport in a novel immortalized mouse collecting duct principal cell line

The final control of sodium balance takes place in the cortical collecting duct (CCD) of the nephron, where corticosteroid hormones regulate sodium re absorption by acting through mineralocorticoid (MR) and/or glucocorticoid ( GR) receptors. A clone of principal CCD cells (mpkCCD(c14)) has been establ ished that is derived from a transgenic mouse (SV40 large T antigen under t he control of the SV40 enhancer/L-type pyruvate kinase promoter). Cells gro wn on filters form polarized monolayers with high electrical transepithelia l resistance (R-T approximately 4700 Ohm x cm(2)) and potential difference (P-D approximately -50 mV) and have an amiloride-sensitive electrogenic sod ium transport, as assessed by the short-circuit current method (I-SC approx imately 11 mu A/cm(2)). Reverse transcription-PCR experiments using rat MR primers, [H-3]aldosterone, and [H-3]dexamethasone binding and competition s tudies indicated that the mpkCCD(c14) cells exhibit specific MR and GR, Ald osterone increased I-SC in a dose- (10(-10) to 10(-6) M) and time-dependent (2 to 72 h) manner, whereas corticosterone only transiently increased I-SC (2 to 6 h). Consistent with the expression of 11 beta-hydroxysteroid dehyd rogenase type 2, which metabolizes glucocorticoids to inactive 11-dehydrode rivates, carbenoxolone potentiated the corticosterone-stimulated I-SC. Aldo sterone (5 x 10(-7) M)-induced I-SC (fourfold) was associated with a three- to fivefold increase in alpha-ENaC mRNA (but not in those for beta- or gam ma-ENaC) and three- to 10-fold increases in alpha-ENaC protein synthesis. I n conclusion, this new immortalized mammalian CCD clonal cell line has reta ined a high level of epithelial differentiation and sodium transport stimul ated by aldosterone and therefore represents a useful mammalian cell system for identifying the genes controlled by aldosterone.