Insulin Stimulates Transepithelial Sodium Transport by Activation of a Protein Phosphatase That Increases Na-K Atpase Activity in Endometrial Epithelial Cells

Abstract

The objective of this study was to investigate the effects of insulin and insulin-like growth factor I on transepithelial Na+ transport across porcine glandular endometrial epithelial cells grown in primary culture. Insulin and insulin-like growth factor I acutely stimulated Na+ transport two- to threefold by increasing Na+-K+ ATPase transport activity and basolateral membrane K+ conductance without increasing the apical membrane amiloride-sensitive Na+ conductance. Long-term exposure to insulin for 4 d resulted in enhanced Na+ absorption with a further increase in Na+-K+ ATPase transport activity and an increase in apical membrane amiloride-sensitive Na+ conductance. The effect of insulin on the Na+-K+ ATPase was the result of an increase in Vmax for extracellular K+ and intracellular Na+, and an increase in affinity of the pump for Na+. Immunohistochemical localization along with Western blot analysis of cultured porcine endometrial epithelial cells revealed the presence of α-1 and α-2 isoforms, but not the α-3 isoform of Na+-K+ ATPase, which did not change in the presence of insulin. Insulin-stimulated Na+ transport was inhibited by hydroxy-2-naphthalenylmethylphosphonic acid tris-acetoxymethyl ester [HNMPA-(AM)3], a specific inhibitor of insulin receptor tyrosine kinase activity, suggesting that the regulation of Na+ transport by insulin involves receptor autophosphorylation. Pretreatment with wortmannin, a specific inhibitor of phosphatidylinositol 3–kinase as well as okadaic acid and calyculin A, inhibitors of protein phosphatase activity, also blocked the insulin-stimulated increase in short circuit and pump currents, suggesting that activation of phosphatidylinositol 3–kinase and subsequent stimulation of a protein phosphatase mediates the action of insulin on Na+-K+ ATPase activation.

Sodium and anion transport across the avian uterine (shell gland)epithelium

“…The observation that amiloride and benzamil produced inhibition of the basal I sc and that this decrease in current was associated with inhibition of the apical-tobasolateral and net Na + flux strongly suggested that ENaC Na + channels were involved in Na + absorption across this epithelium. This result was consistent with previous studies of Na + transport across the porcine, murine and human endometrial epithelium (Vetter and O'Grady, 1996;Deachapunya et al, 1999;Palmer-Densmore et al, 2002;Chan et al, 2001;Matthews et al, 1998). In native porcine endometrial tissues mounted in Ussing chambers, the surface epithelial cells exhibited a basal I sc that was blocked by amiloride with an IC 50 value (0.8·µmol·l -1 ) identical to that reported for the shell gland epithelium in this study (Vetter and O'Grady, 1996).…”

“…In native porcine endometrial tissues mounted in Ussing chambers, the surface epithelial cells exhibited a basal I sc that was blocked by amiloride with an IC 50 value (0.8·µmol·l -1 ) identical to that reported for the shell gland epithelium in this study (Vetter and O'Grady, 1996). Similar effects of amiloride and benzamil on basal I sc were reported for cultured porcine glandular endometrial epithelial cells maintained under serum-free conditions (Deachapunya et al, 1999). Analysis of tissue morphology (Fig.·1) indicated the presence of lamellae that were clearly distinct from the mucosal morphology of mammalian endometrium (Vetter and O'Grady, 1996).…”

Insulin regulates alveolar epithelial function by inducing Na+/K+-ATPase translocation to the plasma membrane in a process mediated by the action of Akt

“…2A shows that pre-incubation of ATII cells with genistein, a broad tyrosine-kinase inhibitor, completely prevented insulin-induced Na + /K + -ATPase recruitment without affecting the basal levels. Previous studies reported the participation of the PI3K-signaling system in the activation of Na + /K + -ATPase in fibroblast and kidney epithelial cells (Deachapunya et al, 1999;Ragolia et al, 1997;Sweeney et al, 2001). Incubation with 100 nM wortmannin or LY 294002, two PI3K inhibitors, markedly prevented insulin-induced Na + /K + -ATPase translocation (Fig.…”

“…The insulin receptor tyrosine kinase is the first protein activated in the insulin-signaling network (Deachapunya et al, 1999). Fig.…”

Regulation of electrolyte transport across cultured endometrial epithelial cells by prolactin

“…In the primary culture of mouse and porcine endometrial epithelial cells, prostaglandins (PGs) especially PGE2, adrenaline, ATP, and UTP were found to activate anion secretion (Chan et al 1997, Fong et al 1998, Deachapunya & O'Grady 1998, Palmer-Densmore et al 2002. These epithelial cells also exhibited Na C transport that was activated by insulin and insulin-like growth factor, and inhibited by epidermal growth factor (Deachapunya et al 1999, Deachapunya & O'Grady 2001.…”