Solvent effects on squid sodium channels are attributable to movements of a flexible protein structure in gating currents and to hydration in a pore

Abstract

1. Solvent effects on the time course of gating and sodium currents were analysed in squid sodium channels using four non-electrolytes of different size, glycerol, erythritol, glucose and sucrose, to separate effects of viscosity from those of osmalarity and to obtain viscosity and osmolarity parameters that were independent of molecular size. 2. The gating and sodium currents were reversibly slowed in a voltage-independent manner as the non-electrolyte concentration increased. 3. Solvent effects were analysed using a model in which the percentage change in time constant was expressed by an equation involving the viscosity parameter α and the osmolarity parameter δ: t/t(o) = α((η/η(o)) - 1 + 100α-1)exp(δΔΠ), where η/η(o) is solution viscosity and ΔΠ is increase in osmolarity Since the solution viscosity was found experimentally to be a function of the solution osmolarity solvent effects are described by an equation with one independent variable η/η(o) or ΔΠ. 4. Voltage sensor movement, reflected in gating currents, was primarily sensitive to viscosity, as its decay time constant was a function of η/η(o) with only a minor sensitivity to osmolarity (δ was 2-3 water molecules). 5. For sodium currents, it was equal to that of gating currents but δ was 2-3 times greater, suggesting that the final channel opening was primarily sensitive to osmolarity (Δδ was 5 water molecules). The relative ineffectiveness of the largest non-electrolyte, sucrose, suggested that this osmolarity-sensitive step in channel opening occurred in the narrow pore region. 6. Sodium channel inactivation was primarily sensitive to osmolarity (Δδ was 8-12 water molecules). 7. The observed viscosity dependence of the sodium current activation and inactivation processes was attributable to the viscosity-dependent process accompanying; the gating current. 8. This model explains why non-electrolytes slow sodium currents while electrolytes do not. 9. Viscosity effects on gating currents can be explained by a process in which non-electrolytes interact with the flexible hydrophilic parts of sodium channel proteins, but osmolarity effects on the final step need to be explained by a local interaction of several water molecules with fluctuating protein segments in the pore.