Anemone toxin II unmasks two conductance states in neuronal sodium channels

Anemone toxin II (ATX)-modified voltage-dependent neuronal sodium channels were studied in planar lipid bilayers. ATX-modified channels displayed two predominant conducting states: a short-lived (ms-s) high-conductance (approximate to 65 pS) state and a long-lived (s-min) low-conductance (approximate to 10 pS) state. The high-conductance state underwent brief closures (ms) and the low-conductance state underwent long closures (s). The probability of detecting these states was time- and voltage-dependent. The channel's fractional open time (f(o)) due to the high-conductance state increased with depolarization and had a midpoint potential (V-a) of -36 mV and an apparent gating charge (z(a)) of 2.8. The channel's f(o) due to the low-conductance state increased with depolarization and had a V-a of +13 mV and a z(a) of 1.4. At positive potentials, ATX-modified channels slowly (minutes) entered an absorbing non-conducting state. The permeability ratio of Na+/K+ was 2 and 4 for the low- and high-conductance stares, respectively. The saxitoxin analog C3 blocked ATX-modified sodium channels with high affinity (K-d(60-90 mV) = 410 nM, 0.5 M NaCl). The data suggest that upon a depolarization step, ATX-modified channels enter rapidly (ms) into a high-conductance slate and more slowly (s-min) into a low-conductance state. Also as the membrane potential becomes more positive, the equilibrium is shifted from the high- to the low-conductance state and from the conducting states to an absorbing non-conducting state.