Characterization of stretch-activated ion channels in Xenopus oocytes

Abstract

1. The gating and permeation properties of endogenous stretch-activated (SA) ion channels in Xenopus oocytes have been studied using the patch-clamp single channel recording technique. 2. As estimated from the probability of being open (Po), SA channels were equally sensitive to suction or pressure. The Po was also weakly sensitive to voltage, increasing with depolarization. Channel activation did not require Ca2+. 3. Kinetic analysis of single-channel records indicated that there are three closed states and one open state. Among three closed-time distributions, the longest was the most sensitive to both pipette pressure and membrane voltage. The open time was independent of both pressure and voltage under a wide variety of ionic conditions, but was sensitive to the species of extracellular ion as follows: Na+ greater than Cs+ greater than K+ greater than Rb+ greater than Li+. The open time had a monotonic mole fraction relationship in mixtures of Li+ and K+. 4. The SA channels were cation-selective inward rectifiers. The selectivity for permeation, based on slope conductance, was: K+ greater than NH4+ greater than Cs+ greater than Rb+ greater than Na+ greater than Li+ greater than Ca2+. 5. Tetraethylammonium (TEA+) was impermeable but was not a channel blocker. 6.Open-channel current amplitude saturated with increasing extracellular K+, and was a monotonic function of the mole fraction of Li+ and K+ in mixtures of the two ions. 7. The channel has at least two separate ion binding sites: an intra-channel site suggested by the permeation data, and an allosteric site suggested by the voltage-independent effects of permeant ions on open time. A symmetric two-barrier, one-site model can quantitatively describe the permeation data. A kinetic model is proposed to quantify the gating kinetics and the effect of ion binding at the allosteric site.