Extracellular anion modulation of ASIC3

Abstract

Acid‐sensing ion channels (ASICs) are non‐voltage cation channels gated by extracellular protons. ASICs have chloride‐binding sites in the extracellular domain that are highly conserved between ASIC isoforms. However, the significance of the Cl binding is unknown. We investigated the effect of different anions on heterologously expressed ASIC3 currents recorded by whole‐cell patch‐clamp. Replacement of extracellular Cl with the impermeable and inert anion, methanesulfonate (MS), had no effect upon peak current amplitude, however at the threshold of ASIC3 activation there was a slight potentiation of pH 7‐evoked current in MS. The mechanism involved a shift in steady‐state desensitization whereby Cl caused more desensitization at a given holding pH. In addition, ASIC3 currents desensitized at a faster rate in Cl compared to MS. Other anions, including bromide and thiocyanate, produced the opposite effect of MS; they decreased proton sensitivity and increased kinetic rates. The effects of various anions followed the lytropic series: thiocyanate > Br > Cl > MS. Mutation of the purported Cl binding sites in ASIC3 abolished the modulatory effects of anions. In addition to sensing pH changes, modulation of ASICs by anion fluxes associated with ischemia further positions the channels to sense changes in the extracellular environment.