Basolateral translocation by vasopressin of the aldosterone-induced pool of latent Na-K-ATPases is accompanied by alpha 1 subunit dephosphorylation: Study in a new aldosterone-sensitive rat cortical collecting duct cell line

The regulation of plasma membrane Na+-K+-ATPases (NKA) expression by aldost erone and arginin vasopressin (AVP) in the cortical collecting duct (CCD) h as been examined in a new rat CCD cell line, designated as RCCD2. This cell line has maintained many characteristics of the CCD-in particular, the exp ression of the mineralocorticoid receptor. Mineralocorticoid receptor is ex pressed at the protein level and binds H-3-aldosterone (approximately 15 to 20 fmol/mg protein). Short-circuit current (Isc) experiments showed approx imately a twofold increase in Isc associated with a decrease in transepithe lial resistance when cells were treated with aldosterone concentrations as low as 10(-9) M. This effect on Isc was significant 2 h after aldosterone a ddition and was still present after 24 h. It was accompanied by an increase in the amount of mRNA encoding for the a subunit of the epithelial sodium channel (sixfold) and the alpha1 subunit of NKA (fourfold) after 24 h of ho rmone treatment. In addition, mRNA expression of the serum- and glucocortic oid-induced kinase (Sgk) was increased by 10(-9) M aldosterone treatment as early as 45 min after hormone addition. As had already been documented in native CCD obtained by microdissection, incubation of RCCD2 cells for 24 It with aldosterone resulted in the constitution of a latent pool of NKA that could be rapidly recruited by AVP (15 min). NKA biotinylation experiments and preparation of membrane fractions show that this latent pool of NKA is present in the intracellular compartment of the cells and is recruited by A VP in the basolateral membrane through a translocation process. This mechan ism is accompanied by dephosphorylation of the alpha (1) catalytic subunit of NKA.