ACTIVATION OF EPITHELIAL Na + CHANNEL ACTIVITY BY POLYUNSATURATED n‐3 FATTY ACIDS: POSSIBLE INVOLVEMENT OF AN A‐KINASE ANCHORING PROTEIN

Abstract

Essential fatty acids (PUFAs) have been shown to modulate enzymes, channels and transporters, to interact with lipid bilayers and to affect metabolic pathways. We have previously shown that eicosapentanoic acid (EPA, C20:5, n-3) activates epithelial sodium channels (ENaCs) in a cAMP-dependent manner involving stimulation of PKA (Mies et al, 2004). In the present study, we explored further the mechanism of EPA stimulation of ENaC in A6 cells. Consistent with our previous studies, EPA had no further stimulatory effect on amiloride-sensitive transepithelial current (INa) in the presence of 8Br-cAMP. Thus, we investigated the effect of EPA on cellular pathways which produce cAMP. EPA did not modify either GTPase or adenylate cyclase activities. There was no measurable increase in total cellular cAMP accumulation in the presence of EPA. HT31, an inhibitor of the binding between PKA and A-Kinase Anchoring Proteins (AKAPs) as well as cerulenin, an inhibitor of myristoylation and palmitoylation, prevented the stimulatory effect of EPA and 8Br-cAMP on INa. Preliminary experiments with Fluorescence Resonance Energy Transfer were carried out with fluorescently labelled regulatory and catalytic subunits of PKA. The data suggest compartmentalization of AKAP-activated PKA with its substrates.