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

Characteristics of a rat cortical collecting duct cell line that maintains high transepithelial resistance. This study describes the establishment of a rat kidney cortical collecting duct (CCD) clonal cell line (RCCD 1 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, RCCD 1 cells are organized as monolayers of cuboid cells separated by tight junctions and form domes. Grown on permeable filters, confluent RCCD 1 cells exhibit high transepithelial resistance (R t : 2390 ± 140 Ω · cm 2 ), transepithelial potential difference (PD) of -10.5 ± 1.2mV lumen negative, an associated short-circuit current (Isc) of 4.3 ± 0.5 μA/cm 2 , and generated significant Na + , K + , H + and HCO 3 − gradients, reflecting Na + and H + reabsorption and K + and HCO 3 − secretion. RCCD 1 cells exhibit features of both principal (PC) and intercalated (IC) cells. Consistent 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 epithelial Na + channel and α 1 and β 1 subunits of Na + -K + -ATPase. Moreover, Na + -K + -ATPase was immunolocalized at the basolateral side of the cells. Arginine vasopressin (AVP) induced a significant increase in both cellular cAMP content and Isc. Amiloride decreased in a dose-dependent manner Isc from untreated and AVP-treated RCCD 1 cells. In addition, a barium-sensitive K + conductance was evidenced in the apical side of the cells. Consistent with IC phenotype, isoproterenol (ISO) provoked a large increase in cellular cAMP and stimulated Isc. The effect of ISO on Isc was blocked by 5 × 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 RCCD 1 cell line has maintained many of the original properties of rat CCD from which they were derived.