Anemone toxin II unmasks two conductance states in neuronal sodium channels

Abstract

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 (≈ 65 pS) state and a long-lived (s-min) low-conductance (≈ 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 states, 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 state 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.