Figure 1.

NS5A interferes with K⁺ channel function and is neuroprotective. A, Top, Whole-cell K⁺ currents in rat cortical neurons in culture, evoked by 10 mV incremental steps (−80 to +50 mV). Neurons were transfected with vector (VEC) or NS5A (genotype 1b) and exposed to vehicle or AMG. Scales are 1 nA and 5 ms. Bottom, Mean (± SEM; n = 7–20 cells per group) current densities (at +10 mV) from cells such as those shown above. ANOVA/Bonferroni, all p < 0.001; ***VEC versus NS5A, ^†VEC versus VEC/AMG, ^‡NS5A versus NS5A/AMG and VEC/AMG versus NS5A/AMG. B, Resting membrane voltage (Vm) and input resistance were measured under current clamp (R_in) and threshold synaptic conductance (Thresh-g_syn) was measured under dynamic clamp in untransfected (UT) and vector-expressing cells (VEC) or NS5A1b-transfected neurons (mean ± SEM; n = 5–8 cells per group). Scales are 20 mV and 500 ms. (C) Repetitive firing was recorded for VEC and NS5A1b transfected neurons using 1 s depolarizing current steps of increased amplitudes. Top, Traces are representative of neurons in each group at three depolarizing currents (n = 8). Bottom, Frequency-current relationships were fit to nonlinear regressions and compared using an F test. NS5A1b transfected neurons have a significantly higher repetitive firing frequency as a function of current when compared with VEC transfected neurons (p < 0.01). D, Rat cortical neurons expressing NS5A1b, but not empty vector (VEC), are protected from exposure to AMG. Shown are mean (±SD) luciferase values (counts per second, CPS) of a representative experiment, performed in quadruplicate; *p < 0.05, ANOVA/Dunnet. Inset, mean ± SEM of viability for a total of 7 independent experiments (each performed in quadruplicate), expressed as a percentage of control; **p < 0.01, paired t test.