Conservation of pH sensitivity in the epithelial sodium channel (ENaC) with Liddle's syndrome mutation

Gain-of-function mutations of the epithelial Na+ channel (ENaC) cause a rare form of hereditary hypertension, Liddle's syndrome. How these mutations lead to increased channel activity is not yet fully understood. Since wild-type ENaC (wt-ENaC) is highly pH-sensitive, we wondered whether an altered pH-sensitivity of ENaC might contribute to the hyperactivity of ENaC with Liddle's syndrome mutation (Liddle-ENaC). Using Xenopus laevis oocytes as an expression system, we compared the pH-sensitivity of wt-ENaC (alpha beta gamma rENaC) and Liddle-ENaC (alpha beta (R564stop)gamma rENaC). Oocytes were assayed fur an amiloride-sensitive (2 muM) inward current (DeltaI(ami)) at -60 mV holding potential and cytosolic pH was altered by changing the extracellular pH in the presence of 60 mM sodium acetate. Alternatively, cytosolic acidification was achieved by proton loading the cells using a proton-coupled oligopeptide transporter (PepT-1) co-expressed ill the oocytes together with ENaC, Cytosolic but not extracellular acidification substantially reduced DeltaI(ami) while cytosolic alkalinisation had a stimulatory effect. This pH-sensitivity was largely preserved in oocytes expressing Liddle-ENaC. The inhibition of wt-ENaC and Liddle-ENaC by cytosolic acidification was independent of so-called sodium-feedback inhibition, since it was not associated with a concomitant increase in intracellular Na+ concentration estimated from the reversal potential of DeltaI(ami). In addition C-terminal deletions in the alpha or gamma subunits or in all three subunits of ENaC did not abolish the inhibitory effect of cytosolic acidification. We conclude that ENaC's pH-sensitivity is not mediated by its cytoplasmic C-termini and that an altered pi-I-sensitivity of ENaC does not contribute to the pathophysiology of Liddle's syndrome.