Figure 5. Newts possess Na_v channel mutations that confer physiological resistance to TTX.

(A) Predicted topology of Na_v1.6 with mutations in domains I, III, and IV. Sequence alignment of Na_v1.6 pore-loop motifs revealed three amino acid differences in newts from Oregon or Idaho populations. (B) Representative currents from wild-type mouse Na_v1.6 or Na_v1.6 with newt substitutions Y371A, V1407I, and I1699V treated with 1 µM (blue) or 10 µM (orange) TTX. (C) Current-voltage (I–V) relationships showing normalized currents for wild-type (n = 21) and mutant Na_v1.6 (n = 20) channels. Wild-type Na_v1.6 was blocked by TTX (Tukey’s multiple comparisons test with Bonferroni correction: control vs. 1 µM, p<0.0001; control vs. 10 µM, p<0.0001), while mutated Na_v1.6 was unaffected (repeated measures ANOVA, p=0.879). (D) Dose-response curves showing the proportion of Na⁺ current elicited during a step depolarization from −100 to −20 mV for wild-type, individual mutants, and triple-mutant Na_v1.6 channels exposed to increasing concentrations of TTX. Sample sizes are provided in Table 2. Data were fit with a Hill equation to estimate IC₅₀ values.

Figure 5—figure supplement 1. Newt Na_v1.6 mutations increase TTX resistance in the orthologous mouse Na_v1.6.

Current-voltage curves showing normalized peak Na⁺ currents measured for wild-type mouse Na_v1.6 and mutant constructs DI Y371A, DIII V1407I, and DIV I1699V containing one of three mutations in the presence of the control and wash solutions (black), 1 µM TTX (blue), and 10 µM TTX (red). Each construct was coexpressed with rat β1 and β2 subunits. Data are shown as peak currents normalized to the maximum peak current recorded for each oocyte. Dotted lines indicate current after washing TTX from the oocyte. The construct containing the DI mutation was largely unaffected by TTX at 1 or 10 µM (n = 17); those containing the DIII (n = 13) and DIV (n = 15) mutations were more sensitive, but each channel shows reduced sensitivity relative to the wild-type channel (n = 21).