The regulation of selective and nonselective Na⁺ conductances in H441 human airway epithelial cells

Abstract

Brown SG, Gallacher M, Olver RE, Wilson SM. The regulation of selective and nonselective Na ϩ conductances in H441 human airway epithelial cells.

SGK1 activity in Na+ absorbing airway epithelial cells monitored by assaying NDRG1-Thr346/356/366 phosphorylation

“…5d) and regression analysis showed that G Na (601± 220 pS cell −1 ) accounted for 33.8±7.0% of the total membrane conductance at physiologically relevant potentials. The magnitude of this glucocorticoid-induced conductance is, therefore, essentially identical to that documented in previous studies in which H441 cells were exposed to dexamethasone for 24-48 h [11,13] and is, therefore, clear that the dexamethasone-induced increase in G Na is fully developed after only 2-3 h. Figure 5e shows currents recorded from cells that had been stimulated with dexamethasone (0.2 μM, 2-3 h) in the presence of LY294002 (50 μM). Since the effects of LY294002 upon PI3K are reversible, this compound was present in all bath solutions used in these experiments.…”

“…5a) showed that V m was normally −60.4±3.7 mV and lowering [Na + ] o had no effect upon V m ( Fig. 5a; V m =−61.0±4.1 mV), indicating that G Na is negligible under these conditions [see also 11,13]. Further analysis confirmed this conclusion by showing that I Na was negligible across the entire range of potentials tested (Fig.…”

“…Membrane currents (I m ) were recorded from single cells held under voltage clamp in the perforated patch recording configuration using bath and pipette solutions designed to maintain quasiphysiological ionic gradients [11,13]. In each experiment, the mean current evoked by driving the holding potential (V Hold ) through a series of four ramps (−113 to 87 mV in 2 s) was recorded both under standard conditions and after [Na + ] o had been lowered to 10 mM (Nmethyl-D-glucammonium substitution).…”

“…Values of membrane conductance (G Na ) were estimated by regression analysis (i.e., ΔI/ΔV Hold ). These methods are described in detail elsewhere [11,13].…”

“…In the present study, we, therefore, use this new approach to monitor the changes in SGK1 activity associated with the induction and hormonal regulation of the glucocorticoid-dependent Na + conductance (G Na ) in a human airway epithelial cell line [11,13].…”