Bothriopsis venoms contain L-amino acid oxidase, esterase, peptidase, phosphodiesterase, phospholipase A2 (PLA2) and proteolytic activities, as well as coagulant, hemorrhagic and myotoxic activities ( Kuch et al., 1996, Porto et al., 2007 and Furtado et al., 2010), in addition to causing

neutrophil migration into the mouse peritoneal cavity ( Porto et al., 2007); other biological activities of these venoms have been poorly studied. In this work, we investigated the neuromuscular activity of venom from the Amazonian forest viper Bothriopsis bilineata smargadina. Male Swiss mice (25–30 g) obtained from the Multidisciplinary Center for Biological Investigation Alectinib molecular weight (CEMIB/UNICAMP) were housed 10/cage at 23 °C on a 12 h light/dark cycle with lights on at 6 check details a.m. Male chicks (4-8 days old) were provided by Granja Ito S/A (Campinas, SP) and housed in metal cages with a sawdust substrate. The mice and chicks had free access to food and water. This study was approved by the institutional Committee for Ethics in Animal Experimentation (CEEA/UNICAMP, protocol no. 2267-1). Bothriopsis

b. smargadina venom was a pool obtained from adult snakes of both sexes captured in the Amazon region. The venom was desiccated and stored at −20 °C until used. When required, the venom was dissolved in 0.9% NaCl prior to use. Chick biventer cervicis nerve-muscle preparations were obtained and mounted (resting tension: 0.5 g) in Krebs solution at 37 °C and allowed to stabilize for 20 min prior to use, as described elsewhere (Borja-Oliveira et al., 2003 and Rodrigues-Simioni et al., 2004). Muscle responses to exogenous acetylcholine (ACh, 110 μM) and KCl (40 mM) were obtained before and after incubation with venom (0.1–30 μg/ml) to screen for postsynaptic

neurotoxicity and myotoxicity (Harvey et al., 1994). Creatine kinase (CK) release was measured for one venom concentration (10 μg/ml) in preparations incubated at 37 °C; activity was assayed using commercial kits ADAMTS5 (CK-NAC, LaborLab, São Paulo, SP, Brazil). The influence of temperature on venom-induced neuromuscular blockade was examined by doing some experiments at 22 °C. Mouse phrenic nerve-diaphragm preparations were mounted in Tyrode solution (composition, in mM: NaCl 137, KCl 2.7, CaCl2 1.8, MgCl2 0.49, NaH2PO4 0.42, NaHCO3 11.9 and glucose 11.1, pH 7.0 at 37 °C after equilibration with 95% O2/5% CO2), as described by Oshima-Franco et al. (2004). After stabilization for 20 min, the preparations were incubated with different venom concentrations (1, 10 and 30 μg/ml, one concentration per preparation) for 120 min and the changes in twitch-tension recorded. To examine the influence of temperature on neuromuscular blockade, some experiments were initially done at 22 °C and the temperature then returned to 37 °C for the rest of the incubation.