Inhibition of neuronal high-voltage activated calcium channels by the ω- Phoneutria nigriventer Tx3-3 peptide toxin

Abstract

We have investigated the effect of ω-PnTx3-3 (referred to in previous papers simply as Tx3-3), a peptide toxin from the venom of the spider Phoneutria nigriventer, on neuronal high-voltage activated (HVA) Ca 2+ channels, using whole-cell patch-clamp. ω-PnTx3-3 (120 nM) blocked 74±8% of the total HVA Ca 2+ currents of cerebellar granule neurones, without affecting the low-voltage activated (LVA) current. P/Q/R-type currents in cerebellar granule neurones, isolated using 4 μM nicardipine and 100 nM ω-conotoxin GVIA, were markedly (79±6%) inhibited by 60 nM ω-PnTx3-3. R-type currents, isolated either by additional application of 0.5–1 μM of ω-agatoxin IVA or by pre-incubation with 5 μM ω-conotoxin MVIIC were inhibited almost totally by 120 nM of ω-PnTx3-3. ω-PnTx3-3 reversibly altered the kinetics of the P/Q/R current, increasing the degree of inactivation that occurred during a 50 ms pulse from 20% to 40%. N-type currents, recorded from neuroblastoma N18 cells, were partially (34±2%) inhibited by 320 nM ω-PnTx3-3. L-type currents, recorded from GH3 cells, were partially (45±12%) inhibited by 80 nM ω-PnTx3-3. We conclude that ω-PnTx3-3 inhibits all known HVA Ca 2+ channels, and most effectively the P/Q- and R-type currents.