Analysis of ectatomin action on cell membranes

Ectatomin (m = 7928 Da) is a toxic component from the Ectatomma tuberculatum ant venom containing two homologous polypeptide chains (37 and 34 residues) linked to each other by a disulfide bond. In aqueous solution it forms a four α-helix bundle. At concentrations of 0.05-0.1 μM, ectatomin forms channels in cellular and artificial bilayer membranes. Immunochemical analysis of the intracellular distribution of ectatomin showed that the toxin gets efficiently inserted into the plasma membrane at a concentration of 5 x 10^-7 M and does not penetrate inside the cell. The effect of ectatomin on cardiac L-type calcium current was studied. Calcium currents (I(Ca)) in isolated rat cardiac ventricular myocytes were measured using the whole-cell perforated patch-clamp technique. It was shown that ectatomin at concentrations of 0.01-10 nM inhibited I(Ca) after a latency of few seconds. I(Ca) was decreased twofold by 10 nM ectatomin. However, the most prominent effect of ectatomin was observed after stimulation of I(Ca) by isoproterenol, an agonist of β-adrenoreceptors, or forskolin, a stimulator of adenylate cyclase. At a concentration of 1 nm, ectatomin abolished the isoproterenol- and forskolin-sensitive components of I(Ca). The inhibitory effect of ectatomin was partially reversed by subsequent application of 2 μM of forskolin, whereas subsequent isoproterenol application did not produce the same effect.