Characteristics of a rat cortical collecting duct cell line that maintains high transepithelial resistance

BLOT-CHABAUD, MARCEL; LAPLACE, MONIQUE; CLUZEAUD, FRANÇOISE; CAPURRO, CLAUDIA; CASSINGÉNA, ROLAND; VANDEWALLE, ALAIN; FARMAN, NICOLETTE; BONVALET, JEAN PIERRE

KIDNEY INTERNATIONAL

NATURE PUBLISHING GROUP

Año: 1996 vol. 50 p. 367 - 376

0085-2538

This study describes the establishment of a rat kidney cortical collecting duct (CCD) clonal cell line (RCCD1 cells) that maintains high transepithelial resistance and specific hormonal sensitivities. Immortalized cells were obtained by infection of primary cultured CCD cells with the wild-type simian virus 40. Grown on Petri dishes, RCCD1 cells are organized as monolayers of cuboid cells separated by tight junctions and form domes. Grown on permeable filters, confluent RCCD1 cells exhibit high transepithelial resistance (R(t) 2390 ± 340 Ω · cm2), transepithelial potential difference (PD) of - 105 ± 1.2 mV lumen negative, an associated short-circuit current (Isc) of 4.3 ± 0.5 μA/cm2, and generated significant Na+, K+, H+ and HCO3 gradients, reflecting Na+ and H+ reabsorption and K+ and HCO3 secretion. RCCD1 cells exhibit features of both principal (PC) and intercalated (IC) cells. Consistently with PC phenotype, about 50% of the cells were positively stained by a PC=specific agglutinin. In situ hybridization studies revealed the presence of α, β and γ subunit mRNAs of the amiloride-sensitive Na+ channel and α1 and β1 subunits of Na+-K+-ATPase. Moreover, Na+-K+-ATPase was immunolocalized at the basolateral of the cells. Arginine vasopressin (AVP) induced a significant increase in both cellular cAMP content and Isc. Amiloride decreased in a dose-dependent manner be from untreated and AVP- treated RCCD1 cells. In addition, a barium-sensitive K+ conductance was evidenced in the apical side of the cells. Consistent with K+ phenotype, isoproterenol (ISO) provoked a large increase in cellular cAMP and stimulated Isc. The effect of ISO on Isc was blocked 5 x 10-3 M DPC, a chloride channel blocker. Finally, AVP plus ISO had additive effect on Isc. Taken together, these results provide evidence that the RCCD1 cell has maintained many of the original properties of rat CCD from which they were derived.