Question: 

The appropriate regulation of the amiloride-sensitive epithelial sodium channel (ENaC) in the aldosterone-sensitive distal nephron is essential for the maintenance of body sodium balance and the long term regulation of arterial blood pressure. Recently, we showed in the mouse cortical collecting duct cell line mCCD_cl1 that prostaglandin E₂ (PGE₂) can stimulate ENaC probably via basolateral EP4 receptors (Niklas et al. 2010 Acta Physiologica 198, Supplement 677). In the present study we further investigated the underlying mechanisms and a potential autocrine release of PGE₂ by mCCD_cl1 cells.

Methodology: 

We used the recently established mCCD_cl1 cell line, a model of principal cells of the cortical collecting duct, grown on permeable supports. Transepithelial ion transport was quantified by equivalent short circuit current (I_SC) measurements in Ussing chambers. A biotinylation assay and Western Blotting were used to detect changes of whole cell or apical surface expression of ENaC. Extracellular PGE₂ concentrations were analysed using ELISA.

Result: 

Exposure of the mCCD_cl1 cells to basolateral PGE₂ and to forskolin, an activator of the adenylyl cyclase (AC), had similar but non-additive stimulatory effects on ENaC. This indicates that the AC / protein kinase A signalling pathway is involved in PGE₂-dependent ENaC stimulation. Biotin surface labelling and Western Blotting revealed that basolateral PGE₂ enhanced the abundance of ENaC channels in the apical plasma membrane of the mCCD_cl1 cells. Interestingly, application of arachidonic acid (AA) also increased the amiloride-sensitive I_SC. This effect was prevented by pre-treating the cells with the cycloxygenase-2 (COX-2) selective inhibitor NS-398 but not with a selective COX-1 inhibitor. Basolateral exposure of the cells to EP4 receptor antagonists (AH23848, GW627368X) largely prevented the AA-mediated response. Finally, we detected elevated PGE₂ in the basolateral medium upon AA-stimulation.

Conclusion: 

Arachidonic acid, via an autocrine release of COX-2 derived PGE₂, enhances ENaC activity in mCCD_cl1 cortical collecting duct cells by increasing its surface expression. These findings indicate that PGE₂ may be an important (patho-) physiological modulator of renal ENaC activity. It is tempting to speculate that enhanced local production or autocrine release of PGE₂ may contribute to sodium retention in inflammatory renal disease.