Short Term Effect of Aldosterone on Na,K-ATPase Cell Surface Expression in Kidney Collecting Duct Cells

Abstract

Aldosterone controls extracellular volume and blood pressure by regulating Na ؉ reabsorption, in particular by epithelia of the distal nephron. A main regulatory site of this transcellular transport is the epithelial sodium channel (ENaC) that mediates luminal Na ؉ influx. The Na,K-ATPase (Na ؉ pump) that coordinately extrudes Na ؉ across the basolateral membrane is known to be regulated by short term aldosterone as well. We now show that in the cortical collecting duct (CCD) from adrenalectomized rats, the increase in Na,K-ATPase activity (approximately 3-fold in 3 h), induced by a single aldosterone injection, can be fully accounted by the increase in Na,K-ATPase cell surface expression (؉ 497 ؎ 35%). The short term aldosterone action was further investigated in cultured mouse collecting duct principal cells mpkCCD cl4 . Within 2 h, maximal Na,K-ATPase function assessed by Na ؉ pump current (I p ) measurements and Na,K-ATPase cell surface expression were increased by 20 -50%. Aldosterone did not modify the Na ؉ dependence of the Na ؉ pumps and induced transcription-and translation-dependent actions on pump surface expression and current independently of ENaC-mediated Na ؉ influx. In summary, short term aldosterone directly increases the cell surface expression of preexisting Na ؉ pumps in kidney CCD target cells. Thus, aldosterone controls Na ؉ reabsorption in the short term not only by regulating the apical cell surface expression of ENaC (Loffing, J., Zecevic, M., Feraille, E., Kaissling, B., Asher, C., Rossier, B. C., Firestone, G. L., Pearce, D., and Verrey, F. (2001) Am. J. Physiol. 280, F675-F682) but also by coordinately acting on the basolateral cell surface expression of the Na,K-ATPase.

Early Aldosterone-Induced Gene Product Regulates the Epithelial Sodium Channel by Deubiquitylation

“…We actually first verified whether Usp2-45 is expressed and regulated by aldosterone in these cells. The relative expression of the two isoforms in mpkCCD c14 cells was similar to the observation made in microdissected CNT/CCD with a 10-fold higher expression of the Usp2-45 [35]) in mpkCCD c14 cells. Usp2-45 upregulation is slightly more delayed than that of Sgk1.…”

“…In contrast, the expression of the protease-dead mutant had no effect. The increase in Na ϩ transport that was induced by aldosterone (300 M; see Materials and Methods in the online supplementary material and reference [35]) that was given for 3 h was not modified significantly by the expression of wild-type and mutant Usp2-45, as expected considering that short-term aldosterone prevents the Nedd4-2-mediated ubiquitylation of ENaC by inducing Sgk1. These results that were obtained in CCD cells confirm that Usp2-45, a ubiquitin-specific protease that was identified because of its in vivo regulation by aldosterone, also acts on endogenous ENaC in kidney tubule cells.…”

ERK1/2 Controls Na,K-ATPase Activity and Transepithelial Sodium Transport in the Principal Cell of the Cortical Collecting Duct of the Mouse Kidney

“…Thus, we addressed the question of whether ENaC or Na,K-ATPase activity is controlled by the pERK1/2 pathway. We first tested the effect of PD98059 and two other MEK1/2 inhibitors, namely U0126 and SL327, on Na,K-ATPase transport activity using a protocol developed by Summa et al (21). In this protocol the apical membrane of mpkCCD cl4 cells is selectively permeabilized to monovalent cations with amphotericin B, thus allowing free diffusion of Na ϩ into the cell through the apical membrane.…”

“…After a 7-min stabilization period, cells were apically permeabilized with 17.5 g/ml amphotericin B for 5 min, as described (21). The Na,K-ATPase was activated by apical and basolateral addition of Na ϩ -containing buffer (60 mM Na ϩ gluconate, 56 mM K ϩ gluconate, 1.8 mM CaCl 2 , 1.6 mM MgCl 2 , 0.8 mM KH 2 PO 4 , 2 mM D-glucose, 12 mM essential amino acids, 2 mM nonessential amino acids, 0.4 mM glutamine, 25 mM HEPES, 3 mM BaCl 2 , pH 7.4) (Buffer B).…”

Hyperaldosteronemia and Activation of the Epithelial Sodium Channel Are Not Required for Sodium Retention in Puromycin-Induced Nephrosis

“…Aldosterone is the major sodium retaining hormone in the distal tubule. In CCD, aldosterone increases both ENaC and Na ϩ ,K ϩ -ATPase activities through a biphasic mode: In the short term, it increases the targeting of intracellular pools of ENaC and Na ϩ ,K ϩ -ATPase to the apical and basolateral membrane, respectively (6,7), and in a later phase, it increases the cellular amount of ENaC and Na ϩ ,K ϩ -ATPase (8,9). Because plasma aldosterone level is increased in PAN-nephrotic rats at the time of sodium retention (3,10,11), it is usually assumed that hyperaldosteronemia mediates sodium retention in PAN nephrosis.…”