In contrast, philanthotoxins (from the digger wasp Philanthus triangulum) and orb web spider polyamines can cause neuromuscular blockade by blocking nicotinic channels ( Rozental et al., 1989). We have previously reported the general properties and composition of venom from the Brazilian theraphosid spider Vitalius dubius Mello-Leitão 1923 ( Rocha-e-Silva et al., 2009a, b). In this work, we describe the neuromuscular activity of this venom and of a low molecular mass component capable of blocking vertebrate motor endplate cholinergic
nicotinic receptors. Male Swiss GSK2126458 price white mice (25–30 g) were obtained from the Multidisciplinary Center for Biological Investigation (CEMIB) at UNICAMP and male HY-LINE W36 chicks (4–8 days old) were supplied by Granja Globo Aves Agrovícola Ltda (Mogi Mirim, SP, Brazil). The animals were housed at 23 ± 2 °C on a 12 h light/dark cycle with access to food and water ad libitum. The animal experiments described here were approved by an institutional Committee for Ethics in Animal
Use (CEUA/UNICAMP, protocol no. 1587-1) and were in agreement with the Ethical Principles for Animal Research established by the Brazilian Society of Laboratory Animal Science (SBCAL). Venom was collected from male and female V. dubius by electrical stimulation ( Rocha-e-Silva et al., 2009b) into Eppendorf tubes covered with Parafilm® to avoid contamination with saliva. All reagents and salts for organ bath solutions were purchased from JT Baker (Center Valley, PA, USA) and drugs Enzalutamide price were from Sigma Chemical Co. (St. Louis, MO, USA) or JT Baker. Male chicks were killed with isoflurane
inhalation and the biventer cervicis muscles were removed (Ginsborg and Warriner, 1960) and mounted under a tension of 1 g/cm in a 5 mL organ bath containing warmed (37 °C), aerated (95% O2 + 5% CO2) Krebs solution of the following composition (in mM): NaCl 118.7, KCl 4.7, CaCl2 1.8, NaHCO3 25, MgSO4 Obatoclax Mesylate (GX15-070) 1.17, KH2PO4 1.17 and glucose 11.65, pH 7.5. A bipolar platinum ring electrode was placed around the tendon within which runs the nerve trunk supplying the muscle. Field stimulation was done with a Grass S48 stimulator (0.1 Hz, 0.2 ms, 4–6 V). Muscle contractions and contractures were recorded isometrically via a force-displacement transducer coupled to a PowerLab ML866/P digital myographic system (ADInstruments Pty. Ltd., Sydney, Australia). Contractures to exogenously applied acetylcholine (ACh, 110 μM) and KCl (20 mM) were obtained in the absence of field stimulation prior to treatments and at the end of the experiment, as a test for the presence of myotoxic and neurotoxic activities (Harvey et al., 1994). The preparations were allowed to stabilize for at least 20 min before the addition of ACh or KCl. Venom was added to the bath and changes in the contractile responses were monitored for 60 min for crude venom and up to 120 min for fractions.