Mechanistic insights into lipid regulation of the acid-sensing ion channels.

Abstract

Acid-sensing ion channels are primary sensors of acidosis in the central and peripheral nervous systems. In this respect, ASICs have been implicated in a number of pathologic conditions where tissue acidosis occurs, including pain sensing during inflammation. Interestingly, although ASICs are activated by protons, the prolonged acidosis present in these conditions should render the channels inactive. One potential solution to this paradox is that, in addition to elevated concentration of protons, there are elevated endogenous inflammatory molecules that further potentiate the channel. Among the molecules elevated in inflammatory conditions are single acyl chain lipids like the polyunsaturated fatty acids (PUFAs). We have previously demonstrated a number of these lipids can stabilize the open state of ASICs, leading to a prolonged non-desensitizing activation, which in some cases can occur even at neutral pH. We found that this effect was dependent on a negatively charged head interaction with the outer transmembrane domain of the channel. However, a detailed understanding of the residues involved in this interaction and the mechanism by which lipids potentiate ASICs remain unknown. Here, we explore these open questions using patch-clamp electrophysiology. We find that lipid potentiation of the channel occurs in a state-dependent manner. We highlight isoform-dependent effects of lipid action on ASICs and use site-directed mutagenesis to narrow down key residues important in lipid binding. We examine lipid effects on the single channel parameters to determine how potentiation manifests at the whole-cell level. Finally, we test several potential mechanisms by which lipids potentiate, including the potential alteration of ionic concentrations at activation sites. Together, we build a model of the binding and mechanism of potentiation for an endogenous regulator of ASICs and provide a basis for future therapeutic development.