CHARACTERISTICS OF A RAT CORTICAL COLLECTING DUCT CELL-LINE THAT MAINTAINS HIGH TRANSEPITHELIAL RESISTANCE

This study describes the establishment of a rat kidney cortical collec ting duct (CCD) clonal cell line (RCCD(1) cells) that maintains high t ransepithelial resistance and specific hormonal sensitivities. Immorta lized cells were obtained by infection of primary cultured CCD cells w ith the wild-type simian virus 40. Grown on Petri dishes, RCCD(1) cell s are organized as monolayers of cuboid cells separated by tight junct ions and form domes. Grown on permeable filters, confluent RCCD(1) cel ls exhibit high transepithelial resistance (R(t): 2390 +/- 140 Ohm . c m(2)), transepithelial potential difference (PD) of -10.5 +/- 1.2 mV l umen negative, an associated short-circuit current (Isc) of 4.3 +/- 0. 5 mu A/cm(2), and generated significant Na+, K+, H+ and HCO3- gradient s, reflecting Na+ and H+ reabsorption and K+ and HCO3- secretion. RCCD (1) cells exhibit features of both principal (PC) and intercalated (IC ) cells. Consistent with PC phenotype, about 50% of the cells were pos itively stained by a PC-specific agglutinin. In situ hybridization stu dies revealed the presence of alpha, beta and gamma subunit mRNAs of t he amiloride-sensitive epithelial Na+ channel and alpha(1) and beta(1) subunits of Na+-K+-ATPase. Moreover, Na+-K+-ATPase was immunolocalize d at the basolateral side of the cells. Arginine vasopressin (AVP) ind uced a significant increase in both cellular cAMP content and Isc. Ami loride 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 phe notype, isoproterenol (ISO) provoked a large increase in cellular cAMP and stimulated Isc. The effect of ISO on Isc was blocked by 5 x 10(-3 ) M DPC, a chloride channel blocker. Finally, AVP plus ISO had additiv e effect on Isc. Taken together, these results provide evidence that t he RCCD(1) cell line has maintained many of the original properties of rat CCD from which they were derived.