Do protons block Na⁺ channels by binding to a site outside the pore?

Abstract

In a detailed structural model of the Na⁺ channel which has evolved over the past decade, the selectivity of the channel for cations^1,2, the voltage-dependence of channel block by protons³ and other small cations², and block by tetrodotoxin (TTX)^4,5 all depend on the presence of a negatively charged site inside the channel that is part of a constriction called the ‘selectivity filter’. Much of the detail inherent in the model arises from Woodhull's quantitative desciption of the voltage dependence of Na⁺ channel block by protons³, in which the apparent relief of block observed at positive potentials occurs because the blocking site is located inside the sodium channel ∼25% of the way through the membrane voltage drop. I report here that proton block of skeletal muscle Na⁺ channels, determined from tail currents, is voltage-independent, and that the apparent voltage-dependent block of peak Na⁺ currents is instead an effect of low pH on channel kinetics. These results suggest that the proton binding site lies effectively outside the membrane field and thus at or outside the channel mouth. As a consequence, present models of the channel interior will need to be re-examined and substantially modified.