Novel insecticidal toxins from the venom of the spider Segestria florentina.

Three insecticidal polypeptide toxins (F5.5, F5.6, F5.7) with molecular masses 4973, 4993 and 5159Da were isolated from the venom of the central Asian spider Segestria florentina. These toxins caused the complete flaccid paralysis of Heliothis virescens (Lepidoptera: Noctuidae) larvae (LD(50) 4-10 microg/g), whereas they were inactive upon intravenous injections into mice. On the basis of N-terminal amino acid sequences a family of eight genes encoding highly homologues polypeptides (SFI1-SFI8) was revealed, some of which encode polypeptides actually demonstrated to be present in S. florentina venom. All deduced polypeptides consist of 46 amino acids residues. Comparison of primary structures of SFI1-SFI8 with other spider toxins suggests that this family might share structural and functional relationships with other small spider neurotoxins, several of which are known to be highly selective agonists/antagonists of different voltage-dependent Ca(2+) channels.

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 alpha-helix bundle. At concentrations of 0.05-0.1 microm, 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 (ICa) 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 ICa after a latency of few seconds. ICa was decreased twofold by 10 nm ectatomin. However, the most prominent effect of ectatomin was observed after stimulation of ICa by isoproterenol, an agonist of beta-adrenoreceptors, or forskolin, a stimulator of adenylate cyclase. At a concentration of 1 nm, ectatomin abolished the isoproterenol- and forskolin-sensitive components of ICa. The inhibitory effect of ectatomin was partially reversed by subsequent application of 2 microm of forskolin, whereas subsequent isoproterenol application did not produce the same effect.

Some properties of sodium channels in neuroblastoma cells modified with scorpion toxin and chloramine-T. Single channel measurements.

Currents through sodium channels of neuroblastoma cells were measured using patch technique in outside-out configuration. Scorpion toxin (ScTX) produced 3 to 4 fold prolongation of mean open time and increased number of reopenings. The mean open times showed slow fluctuations around some average values. The distribution of channel open times for ScTX-modified channels required more than one exponential to be fitted. Chloramine-T (ChT) produced qualitatively similar, though weaker, prolongation of open times.

[Effect of several m-cholinoblockaders on the functioning of sodium potential-dependent channels of neuroblastoma cells]. / Vliianie nekotorykh m-kholinoblokatorov na funktsionirovanie natrievykh potentsialzavisimykh kanalov kletok neiroblastomy.

Effects of two central antimuscarinic drugs, namely N-methyl-4-piperidinol benzylate and tropine phenylcyclopentylglycolate on Na currents in internally dialyzed mouse neuroblastoma cells were studied using the suction-pipette voltage-clamp technique. Like amine local anesthetics, the both compounds produce two phenomenologically different types of sodium current inhibition: a steady block without conditioning and a cumulative (use-dependent) block which develops during repetitive membrane pulsing. The ability of these m-cholinoblockers to modulate voltage-dependent ionic permeability of the neuronal membranes is supposed to be one of possible components in the mechanism of their both main and accessory (non-synaptic) effects.

[Effect of blood plasma and serum and its fractions on the activity of the sodium channels in neuroblastoma cells]. / Vliianie plazmy, syvorotki krovi i ee fraktsii na aktivnost' natrievykh kanalov neiroblastomy.

Blood plasma, serum and its fractions containing components of different molecular masses obtained by dialysis and gradual ultrafiltration were tested for their action on the sodium currents of voltage clamped internally perfused neuroblastoma cells. Blood plasma free of platelet factor 4, serum and its fractions containing components of molecular masses more than 50 kD produced an increase in sodium channel currents and shifts in activation and inactivation curves along the voltage axis towards more negative or positive potentials, respectively. Moreover, the serum deficient in components with the molecular masses less than 50 kD produced a more prominent effect on all the parameters tested. Thus, the low-molecular fractions were suggested to exert an inhibitory activity on the stimulatory effects produced by the high-molecular components of the serum.

[Effect of ruthenium red on the sodium channel inactivation of neuroblastoma cells]. / Vliianie rutenievogo krasnogo na inaktivatsiiu natrievykh kanalov kletok neiroblastomy.

Currents through sodium channels in the neuroblastoma cell membrane were measured after internal and external application of the ruthenium red. The treatment of the membrane by the dye resulted in a slowdown of inactivation kinetics of sodium currents and in the changes of activation and inactivation stationary parameters, which were more pronounced after internal application of the ruthenium red.

[Action of chloramine-T on the processes of sodium channel activation and inactivation in neuroblastoma cells]. / Issledovanie deistviia khloramina-T na protsessy aktivatsii i inaktivatsii natrievykh kanalov kletok neiroblastomy.

Effect of chloramine-T, the specific reagent on methionine residues, on gating of sodium channels was studied in neuroblastoma cell membrane. After the chloramine treatment the inactivation became slower, incomplete and the steepness of its voltage dependence considerably decreased; the inactivation curve was shifted towards depolarization. Time course of activation did not change. The activation curve shifted towards negative potentials, its slope being insignificantly decreased. Effective charge of activation as determined from the limiting logarithmic slope of the activation decreased by a factor of 1.17. Possible explanations of the phenomena observed are discussed.