Endocrine Abstracts

Regulation of the epithelial sodium channel (ENaC) trafficking in cells of the distal nephron is of major importance in the control of sodium reabsorption. Mutations in the genes compromising the molecular pathway by which corticosteroids regulate ENaC sodium reabsorption cause several hypertensive syndromes in humans. The molecular events underpinning ENaC exo- and endocytosis and the hormonal effects on ENaC trafficking are poorly understood. To start elucidating these pathways we have generated stable mouse collecting duct cell (M-1) lines that express an N-terminally fused EGFP-ENaC beta subunit. The cell lines grow normally, express easily visualized quantities of the EGFP fused ENaC subunit and form confluent epithelia with high transepithelial resistance when grown on semi-permeable filters. Imaging by confocal microscopy revealed that ENaC was confined to the endoplasmic reticulum (ER) in unpolarized M-1 cells. Activation of the vasopressin pathway results in ENaC exodus out of ER and into intracellular vesicles; this effect depends upon pre-stimulation by corticosteroids. When cells are fully polarized, ENaC largely exits the ER and Golgi apparatus. Corticosteroids (>100nM dexamethasone) exert a profound impact upon the trafficking of ENaC proteins in polarized M-1 cells by promoting ENaC translocation to the apical membrane.
In conclusion polarization, corticosteroids and vasopressin modulate the trafficking of ENaC in rather distinctive manners such that vasopressin or polarization fosters ENaC entering intracellular vesicles whereas corticosteroids play a role further downstream in the cascade of ENaC trafficking events by increasing the exocytic delivery of ENaC to the apical membrane. This system shows corticosteroids' influences on ENaC trafficking in mammalian collecting duct cells and will be used to gain novel molecular insight into this important pathway.