Figure 5. Blockade of wild type-Shaker by CTX in low ionic strength.

(A) Potassium currents from the same oocyte recorded in external solutions in which the main cation is ~ 10 mM Na⁺ (10 Na⁺; left) or ~ 10 mM K⁺ (10 K⁺; left). (B) Same oocytes as in A, in the presence of 5 nM CTX. The time course of the currents appears as a delayed channel activation (C). Relative conductance (G/Gmax) vs. Voltage in low ionic strength. Data points are average of 5 different experiments in 10-Na⁺ (open circles) and 10-K⁺ (filled circles). The solid lines are fits to a Boltzmann distribution. The dashed lines indicate the approximate voltages at which G/Gmax is 0.5 (−30 and −22 mV, for 10-K⁺ and 10-Na⁺, respectively). (D) Point-by-point quotient ratios of the CTX current traces divided by the control current traces at their corresponding voltages. The blue lines are mono-exponential fits to the ratios. Note that all relaxations converge to the same point at the beginning of the pulse, pointing to the CTX inhibition at resting. (E) Fitting parameters of the traces in C. The average time constants (filled symbols) and the asymptotic, or equilibrium, inhibition at each voltage (open symbols) from five experiments each were used to calculate the association and dissociation rates. Lines have no theoretical meaning. (F) Rates constants of CTX binding to the channel calculated according to Equation 3a and 3b. The continuous lines are mono-exponential fits of the function $k=k_{(V=0)}{e}^{\frac{z\delta FV}{RT}}$, where F, R and T have their usual meaning. V is the applied voltage and zδ is the effective valence of the voltage dependence. For five measurements in each condition, the mean (± SEM) were: in 10 Na⁺, k_off(V=0)=3.02 ± 0.02 s⁻¹ with zδ=0.47 ± 0.02; k_on(V=0)=812 ± 26 μM⁻¹s⁻¹ with zδ=0.106 ± 0.026; meanwhile in 10 K⁺, k_off(V=0)=5.6 ± 0.16 s⁻¹ with zδ=0.4 ± 0.016 and k_on(V=0)=3572 ± 96 μM⁻¹s⁻¹ with zδ=0.133 ± 0.023.