FIGURE 5.

Representative plots of activation and steady-state inactivation curves for Na_v1.4 and Na_v1.8 channels. A, pulse protocols for activation curves (top) and inactivation curves (bottom). B and C, symbols indicate data in the absence (solid lines and closed squares) and presence (dotted lines and open circles) of 0.1 μm CTX3C. Boltzmann functions (smooth lines) were fitted to each set of activation curves

or inactivation curves

In the case of ciguatoxin-modified Na_v1.8 channels, activation curves were fitted by a sum of two Boltzmann distributions

During standard double-pulse protocols for the steady-state inactivation measurement, giving conditioning pulses of –120 to –100 mV, increased the availability of Na⁺ channels in the presence of CTX3C before the transition to inactivation, especially in Na_v1.8, making a peak in the curve at around –100 mV. In this case, a fit was obtained by not fitting data points showing potentials more negative than –100 mV. The parameters obtained by the fitting procedure for all of the constructs are shown in Table 1 for the activation curve and in Table 2 for the steady-state inactivation curve. The activation curves were calculated from the peak I_Na value obtained at each membrane potential divided by the driving force. The steady-state inactivation curves were obtained by a double-pulse protocol. The I_Na evoked with a test pulse to –20 mV was preconditioned by a 500-ms clamp step to a variable membrane potential (–160 to –20 mV in 20-mV increments) with a gap of 0.25 ms, and normalized by the unconditioned current elicited with a test pulse from the holding potential (–100 mV).