Effects of ionic strength on proton block of TRPV1 channels. (A) Representative single-channel currents from TRPV1 channels recorded in symmetrical 1,000-mM KCl solutions at different voltages and extracellular pHs as indicated above traces. (B and C) Dose-dependent relationships of the unitary current amplitudes versus [K⁺] at −60 mV (B) and +60 mV (C), respectively. Note that the dose–response curves were saturated at different maximal current amplitudes with different pHs. Dashed lines represent predictions of the Gouy-Chapman-Stern model based on simultaneous fitting of the data at multiple extracellular pHs. The model assumes that protons exert their effects by one-to-one binding to negative surface charges on the membrane to lower local H⁺ and K⁺ concentrations. At high [K⁺], it predicted a common maximal current amplitude independent of extracellular pH. The fits correspond to the following parameter values: σ = 0.2e/nm², pKa = 7 for protonation of surface charges, K_S = 372 mM for the dissociation constant of K⁺ ion binding, n = 1.5 for the apparent Hill coefficient, and i_max = 10 pA for the maximum current amplitude (−60 mV); σ = 0.1e/nm², pKa = 7.2, K_S = 160 mM, n = 1.1, and i_max = 17 (+60 mV). The surface potentials Ψ₀ at 2 M K⁺ for pHs 8.5, 7.4, and 5.5 were, respectively (in mV): −18, −15, and −0.6 (−60 mV), and −17, −12, and −0.4 (+60 mV). (D and E) Fits to a competitive inhibition model in which protons bind to the same sites occupied by K⁺ to block ion flows. Data at different pHs were fit simultaneously. The fit yields the following model parameters: K_S = 219 mM for the dissociation constant of K⁺ binding, pK_a = 6.5 for proton binding, n = 0.5 for the pseudo Hill coefficient, and i_max = 15pA (Vh = −60 mV); Ks = 70 mM, pKa = 5.8, n = 0.7, and i_max = 19 pA (Vh = +60 mV). (F and G) Effects of [K⁺] on proton inhibition at the indicated membrane potentials. The ratios of the unitary current amplitudes at pHs 7.4 and 5.5 to the amplitudes at pH 8.5 were plotted.