Inhibitory Actions of Synthesised Polyamine Spider Toxins and Their Analogues on Neuronal Voltage-Activated Calcium Currents

• Published in: Comparative biochemistry and physiology. C, Comparative pharmacology and toxicology Vol. 116; no. 1; pp. 23 - 32
• Main Authors: Sutton, Kathy G, Stapleton, Simon R, Timms, Graham, Gilmore, Jeremy, Brust, Paul F, Bleakman, David, Scott, Roderick H
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 1997
• Subjects: Cultured neurones; high voltage-activated calcium channels; polyamines; spider toxins; synthesised FTX; ω-Agatoxin IVA; spider toxins; ω-Agatoxin IVA; Cultured neurones; polyamines; high voltage-activated calcium channels; synthesised FTX
• ISSN: 0742-8413
• DOI: 10.1016/S0742-8413(96)00141-7

The whole cell variant of the patch-clamp technique was used to investigate the actions of polyamine spider toxins and their analogues on high voltage-activated Ca 2+ currents. The actions of synthesised FTX (putative natural toxin from the American funnel web spider), sFTX-3.3, Orn-FTX-3.3 and Lys-FTX-3.3 (synthetic analogues of FTX) were studied using cultured dorsal root ganglion neurones from neonatal rats, C2D7 cells (HEK293 cells stably coexpressing recombinant human N-type voltage-activated Ca 2+ channel, α 1B-1- α 2b δβ 1b subunits) and freshly isolated cerebellar Purkinje neurones. In dorsal root ganglion neurones, sFTX-3.3 (10 μM) inhibited high voltage-activated Ca 2+ currents evoked by depolarisations to 0 mV from a holding potential of −90 mV. Partial overlap in Ca 2+ current sensitivity to the polyamine sFTX-3.3 and the peptide spider toxin ω-Aga IVA was observed. However, evidence also suggests sFTX-3.3 and ω-Aga IVA do not show complete pharmacological overlap and that distinct parts of the Ca 2+ current are sensitive to one of two inhibitors. The arginine group on sFTX-3.3 appears to be important for its inhibitory action on Ca 2+ currents, because analogues where this amino acid was replaced with either ornithine (Orn-FTX-3.3) or lysine (Lys-FTX-3.3) were relatively inactive at concentrations below 1 mM. Synthesised FTX (100 μM) was inactive as an inhibitor of Ca 2+ currents recorded from dorsal root ganglion and only produced modest effects in Purkinje neurones and C2D7 cells. At a concentration of 1 mM, nonselective actions were observed that indicated that synthesised FTX and sFTX-3.3 could reversibly inhibit both N- and P-type Ca 2+ channels equally well. In conclusion, the potency of polyamines as nonselective inhibitors of Ca 2+ channels is in part determined by the presence of a terminal arginine, and this may involve an interaction between terminal guanidino groups with Ca 2+ binding sites.