Please use this identifier to cite or link to this item: http://repositorio.unicamp.br/jspui/handle/REPOSIP/242180
Type: Artigo
Title: The neuromuscular activity of Bothriopsis bilineata smaragdina (forest viper) venom and its toxin Bbil-tx (asp49 phospholipase A(2)) on isolated mouse nerve-muscle preparations
Author: Floriano, Rafael Stuani
Rocha, Thalita
Carregari, Victor Corasolla
Marangoni, Sérgio
Cruz-Höfling, Maria Alice da
Hyslop, Stephen
Rodrigues-Simioni, Léa
Rowan, Edward G.
Abstract: The presynaptic action of Bothriopsis bilineata smaragdina (forest viper) venom and Bbil-TX, an Asp49 PLA(2) from this venom, was examined in detail in mouse phrenic nerve-muscle (PND) preparations in vitro and in a neuroblastoma cell line (SK-N-SH) in order to gain a better insight into the mechanism of action of the venom and associated Asp49 PLA(2). In low Ca2+ solution, venom (3 mu g/ml) caused a quadriphasic response in PND twitch height whilst at 10 mu g/ml the venom additionally induced an abrupt and marked initial contracture followed by neuromuscular facilitation, rhythmic oscillations of nerve-evoked twitches, alterations in baseline and progressive blockade. The venom slowed the relaxation phase of muscle twitches. In low Ca2+, Bbil-TX [210 nM (3 mu g/ml)] caused a progressive increase in PND twitch amplitude but no change in the decay time constant. Venom (10 mu g/ml) and Bbil-TX (210 nM) caused minor changes in the compound action potential (CAP) amplitude recorded from sciatic nerve preparations, with no significant effect on rise time and latency; tetrodotoxin (3.1 nM) blocked the CAP at the end of the experiments. In mouse triangularis sterni nerve-muscle (TSn-m) preparations, venom (10 mu g/ml) and Bbil-TX (210 nM) significantly reduced the perineural waveform associated with the outward K+ current while the amplitude of the inward Na+ current was not significantly affected. Bbil-TX (210 nM) caused a progressive increase in the quantal content of TSn-m preparations maintained in low Ca2+ solution. Venom (3 mu g/ml) and toxin (210 nM) increased the calcium fluorescence in SK-N-SH neuroblastoma cells loaded with Fluo3 AM and maintained in low or normal Ca2+ solution. In normal Ca2+, the increase in fluorescence amplitude was accompanied by irregular and frequent calcium transients. In TSn-m preparations loaded with Fluo4 AM, venom (10 mu g/ml) caused an immediate increase in intracellular Ca2+ followed by oscillations in fluorescence and muscle contracture; Bbil-TX did not change the calcium fluorescence in TSn-m preparations. Immunohistochemical analysis of toxin-treated PND preparations revealed labeling of junctional ACh receptors but a loss of the presynaptic proteins synaptophysin and SNAP25. Together, these data confirm the presynaptic action of Bbil-TX and show that it involves modulation of K+ channel activity and presynaptic protein expression. (C) 2015 Elsevier Ltd. All rights reserved.
The presynaptic action of Bothriopsis bilineata smaragdina (forest viper) venom and Bbil-TX, an Asp49 PLA(2) from this venom, was examined in detail in mouse phrenic nerve-muscle (PND) preparations in vitro and in a neuroblastoma cell line (SK-N-SH) in order to gain a better insight into the mechanism of action of the venom and associated Asp49 PLA(2). In low Ca2+ solution, venom (3 mu g/ml) caused a quadriphasic response in PND twitch height whilst at 10 mu g/ml the venom additionally induced an abrupt and marked initial contracture followed by neuromuscular facilitation, rhythmic oscillations of nerve-evoked twitches, alterations in baseline and progressive blockade. The venom slowed the relaxation phase of muscle twitches. In low Ca2+, Bbil-TX [210 nM (3 mu g/ml)] caused a progressive increase in PND twitch amplitude but no change in the decay time constant. Venom (10 mu g/ml) and Bbil-TX (210 nM) caused minor changes in the compound action potential (CAP) amplitude recorded from sciatic nerve preparations, with no significant effect on rise time and latency; tetrodotoxin (3.1 nM) blocked the CAP at the end of the experiments. In mouse triangularis sterni nerve-muscle (TSn-m) preparations, venom (10 mu g/ml) and Bbil-TX (210 nM) significantly reduced the perineural waveform associated with the outward K+ current while the amplitude of the inward Na+ current was not significantly affected. Bbil-TX (210 nM) caused a progressive increase in the quantal content of TSn-m preparations maintained in low Ca2+ solution. Venom (3 mu g/ml) and toxin (210 nM) increased the calcium fluorescence in SK-N-SH neuroblastoma cells loaded with Fluo3 AM and maintained in low or normal Ca2+ solution. In normal Ca2+, the increase in fluorescence amplitude was accompanied by irregular and frequent calcium transients. In TSn-m preparations loaded with Fluo4 AM, venom (10 mu g/ml) caused an immediate increase in intracellular Ca2+ followed by oscillations in fluorescence and muscle contracture; Bbil-TX did not change the calcium fluorescence in TSn-m preparations. Immunohistochemical analysis of toxin-treated PND preparations revealed labeling of junctional ACh receptors but a loss of the presynaptic proteins synaptophysin and SNAP25. Together, these data confirm the presynaptic action of Bbil-TX and show that it involves modulation of K+ channel activity and presynaptic protein expression
Subject: Bloqueio neuromuscular
Canais iônicos
Cálcio - Metabolismo
Venenos de serpentes
Sinaptofisina
Country: Reino Unido
Editor: Elsevier
Citation: The Neuromuscular Activity Of Bothriopsis Bilineata Smaragdina (forest Viper) Venom And Its Toxin Bbil-tx (asp49 Phospholipase A(2)) On Isolated Mouse Nerve-muscle Preparations. Pergamon-elsevier Science Ltd, v. 96, p. 24-37 MAR-2015.
Rights: fechado
Identifier DOI: 10.1016/j.toxicon.2015.01.001
Address: http://www.sciencedirect.com/science/article/pii/S0041010115000045
Date Issue: 2015
Appears in Collections:IB - Artigos e Outros Documentos
FCM - Artigos e Outros Documentos

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