Figure 4. Current- and voltage-clamp experiments on TTX-S voltage-gated sodium channels in the presence of CgNa.

(A) CgNa increases the length of the action potential. Action potentials of a DRG neuron recorded under control conditions, in the presence of CgNa (10 μM) and after washout (5 min). In this particular cell, a small depolarization of approx. 2 mV was observed. The dotted line in all panels indicates the zero voltage level. Inset: data for the entire experimental group in which current-clamp experiments were carried out. Exposure to the toxin increased the duration of action potentials measured at 50% of its amplitude by 463% (n=3). (B) CgNa increased the inactivation time course of TTX-S I_Na. Inset: representative experiment showing the effect of CgNa (10 μM) 1 min after commencing perfusion. Note the slowing of the inactivation process. In both cases, the current level returned to zero after the end of the depolarization pulse. The main plot shows the dose–response curve for CgNa (n=41) on τ_h. Data were fitted (continuous line) by a dose–response function with an IC₅₀ of 1.34±0.4 μM (n=41) and a Hill slope coefficient of 0.6±0.2 (n=41). (C) Effect of CgNa on the current-density–voltage curve. Top: I_Na produced at different potentials in the presence and absence of CgNa (10 μM, for clarity only the first 5 ms of each record are shown). Currents were produced by voltage pulses to the potentials indicated from a holding potential of −100 mV. Note that CgNa affects the currents at all the voltages tested. Bottom: current-density–voltage relationships (n=6). Perfusion with CgNa (10 μM, closed circles) did not produce significant changes either on the voltage at which the maximum current density was reached or on the reversal potential. (D) Effect of CgNa on steady-state inactivation. Top: representative experiment from which the curves were obtained. Bottom: steady-state inactivation of I_Na (n=6). The steady-state inactivation parameter (h_∞) was determined using the two-pulse protocol shown in the inset. Data obtained at the test pulse were plotted as a function of the pre-pulse potential and fitted to a Boltzmann function (continuous lines). CgNa (10 μM, closed circles) caused a significant 8 mV hyperpolarizing shift in the V_½,inact. The slope factor was also significantly changed at 3.5 mV.