Evidence for multiple effects of ProTxII on activation gating in Na(V)1.5

The peptide toxin ProTxII, recently isolated from the venom of the tarantula spider Thrixopelma pruriens, modifies gating in voltage-gated Na(+) and Ca(2+) channels. ProTxII is distinct from other known Na(+) channel gating modifier toxins in that it affects activation, but not inactivation. It shifts activation gating positively and decreases current magnitude such that the dose-dependence of toxin action measured at a single potential reflects both effects. To test the extent to which these effects were independent, we tracked several different measures of current amplitude, voltage-dependent activation, and current kinetics in Na(V)1.5 in a range of toxin concentrations. Changes in voltage dependence and a decrease in G(max) appeared at relatively low concentrations (40-100nM) while a positive shift in the voltage range of activation was apparent at higher toxin concentrations (>/=500nM). Because ProTxII carries a net +4 charge we tested whether electrostatic interactions contributed to toxin action. We examined the effects of ProTxII in the presence of high extracellular Ba(2+), known to screen and/or bind to surface charge. Some, but not all aspects of ProTxII modification were sensitive to the presence of Ba(2+) indicating the contribution of an electrostatic, surface charge-like mechanism and supporting the idea of a multi-faceted toxin-channel interaction.