SNX482 selectively blocks P/Q Ca 2+ channels and delays the inactivation of Na + channels of chromaffin cells

Abstract

The effects of the toxin SXN482 on Ca 2+ channel currents ( I Ca), Na + currents ( I Na), and K + currents ( I K) have been studied in bovine adrenal medullary chromaffin cells voltage-clamped at −80 mV. Currents were elicited by depolarising pulses to 0–10 mV ( I Ca and I Na) or to +60 mV ( I K). SNX482 blocked I Ca in a concentration-dependent manner. The inhibition curve exhibited two phases. The first high-affinity phase comprised 28% of the whole-cell current and exhibited an IC 50 of 30.2 nM. The second low-affinity phase comprised over 70% of I Ca and had an IC 50 of 758.6 nM. Blockade was rapid and fully reversible upon washout of the toxin. Occlusion experiments showed additivity of blockade exerted by nifedipine plus SNX482 (0.3 μM) and by ω-conotoxin GVIA plus SNX482. In contrast, blockade exerted by combined ω-agatoxin IVA plus SNX482 (about 50% of the whole cell) did not show additivity. At 0.3 μM and higher concentrations, SNX482 delayed the inactivation of I Na. The time constant ( τ) for inactivation of I Na in control conditions doubled in the presence of 0.5 μM SNX482. At 0.3 μM, SNX482 did not affect I K. Our data demonstrate that: (i) SNX482 selectively blocks P/Q Ca 2+ channels at submicromolar concentrations; (ii) the toxin partially blocks Na + channels; (iii) SNX482 delays the inactivation of Na + channels. These results reveal novel properties of SNX482 and cast doubts on the claimed selectivity and specificity of the toxin to block the R-type Ca 2+ channel.