Influence of temperature upon paralyzing and myotoxic effects of bothropstoxin-I on mouse neuromuscular preparations

Bothropstoxin-I (BthTX-I), from B. jararacussu venom, is a phospholipase A(2) (PLA(2)) homologue devoid of enzymatic activity. Besides inducing severe myonecrosis, BthTX-I promotes paralysis of both directly and indirectly evoked contractions in isolated neuromuscular preparations. We applied an experimental paradigm in order to characterize the steps involved in the toxic effects of BthTX-I on mouse neuromuscular junction. Myotoxicity was assessed by microscopic analysis of extensor digitorum longus muscles; paralyzing activity was evaluated through the recording of isolated contractions indirectly evoked in phrenic-diaphragm preparations. After 90 min at 35 degreesC, BthTX-I induced complete and irreversible paralysis, and damaged 30.3 +/- 2.7% of muscle fibers. In contrast, no effect was observed when tissues were incubated with BthTX-I at 10degreesC for 60 min and subsequently washed with toxin-free solution and maintained at 35 degreesC. These results indicate that the binding of BthTX-I to the cellular tissue surface is very weak at low temperature and that an additional factor is necessary. However, when tissues were submitted to BthTX-I (10degreesC for 60 min), and the temperature was elevated to 35 degreesC, omitting the washing step, it was observed muscle paralysis and damage in 39.04 +/- 4.2% of muscle fibers. These results indicate that a temperature-dependent step is necessary for BthTX-I to promote both its myotoxic and paralyzing activities. (C) 2004 Elsevier B.V.. All rights reserved

Paralyzing and myotoxic effects of a recombinant bothropstoxin-1 (BthTX-I) on mouse neuromuscular preparations

As a first step to investigate the structure-function relationship of bothropstoxin-1 (BthTX-1), a myotoxin from Bothrops jararacussu snake venom, Our group previously cloned a recombinant toxin (rBthTX-1) in Escherichia coli. The aim or this work was to characterize the biological activities of this rBthTX-1 (1.0 mu M) in both phrenic-diaphragm and extensor digitorum longus preparations in vitro, by means of myographic and morphologic techniques. Native BthTX-1 (1.0 mu M) was used as a standard. The influence of heparin (27.5 mu g/ml) upon the biological activities of both toxins was also investigated. rBthTX-1 had similar effects to the native toxin inducing blockage of both directly and indirectly evoked contractions in phrenic-diaphragm preparations, and muscle damage characterized by edema, round fibers, and cell areas devoid of myofibrils. Interestingly the paralyzing activity of rBthTX-1 was slightly more potent than the native toxin. Heparin prevented paralyzing and myotoxic effects of both the native and recombinant toxins. This work shows that rBthTX-1 was expressed in a fully active form, and presents a biological profile similar to the native toxin. (c) 2005 Elsevier GmbH All rights reserved

Antagonism of myotoxic and paralyzing activities of bothropstoxin-I by surarnin

Polyanionic substances are known to inhibit the myotoxic effects of some crotalide snake venoms. Bothropstoxin-I (BthTX-I), a basic Lys49 phospholipase (PLA(2)) homologue from Bothrops jararacussu venom, besides inducing muscle damage, also promotes the blockade of both directly and indirectly evoked contractions in mouse neuromuscular preparation. In this work, we evaluated the ability of suramin, a polysulfonated naphtylurea derivative, to antagonize the myotoxic and the paralyzing activities of BthTX-I on mice neuromuscular junction in vitro. Myotoxicity was assessed by light and electronic microscopic analysis of extensor digitorum longus (EDL) muscles; paralyzing activity was evaluated through the recording of both directly and indirectly evoked contractions of phrenic-diaphragm (PD) preparations. BthTX-I (1 muM) alone, or pre-incubated with suramin (10 muM) at 37degreesC for 15 min was added to the preparations for 120 min. BthTX-I induced histological alterations typical of myonecrosis in 14.6 +/- 1.0% of EDL muscle fibers. In addition, BthTX-I blocked 50% of both directly and indirectly evoked contractions in PD preparations in 72.1 +/- 9.1 and 21.1 +/- 2.0 min, respectively. Pre-incubation with suramin abolished both the muscle-damaging and muscle-paralyzing activities of BthTX-I. Since suramin is a polyanionic substance, we suggested that its effects result from the formation of inactive acid-base complexes with BthTX-I. (C) 2003 Elsevier Ltd. All rights reserved

O papel do esteroide anabolizante sobre a hipertrofia e força muscular em treinamentos de resistência aeróbia e de força

INTRODUÇÃO: Os efeitos dos esteroides anabolizantes (EA) sobre a massa muscular e força são controversos e dependentes do treinamento realizado e das fibras musculares recrutadas. Com isso, o objetivo deste estudo foi avaliar os efeitos da associação de EA ao treinamento de força ou aeróbio sobre a hipertrofia e força muscular. MÉTODOS: Ratos Wistar (42) foram divididos em seis grupos: sedentário (SC, n = 7), sedentário anabolizante (SA, n = 7), treinado natação controle (TNC, n = 7), treinado natação anabolizante (TNA, n = 7), treinado força controle (TFC, n = 7) e treinado força anabolizante (TFA, n = 7). O EA foi administrado duas vezes por semana (10mg/kg/semana). Os protocolos de treinamento foram realizados durante 10 semanas, cinco sessões semanais. Foram avaliadas a hipertrofia dos músculos sóleo, plantar e gastrocnêmio (massa muscular corrigida pelo comprimento da tíbia), a proteína total muscular (Bradford) e a força muscular em patas traseiras (testes de resistência à inclinação). RESULTADOS: Não foram observadas diferenças significantes na hipertrofia do músculo sóleo. Os grupos TFC e TFA apresentaram, respectivamente, hipertrofia de 18% e 31% no músculo plantar comparado ao grupo SC. A hipertrofia foi 13% maior no grupo TFA em relação ao grupo TFC. Resultados semelhantes foram encontrados no músculo gastrocnêmio. Os grupos TFC e TFA apresentaram significantes aumentos na quantidade total de proteína nos músculos plantares, sendo essa mais pronunciada no grupo TFA e positivamente correlaciona a hipertrofia muscular. Observamos aumento de força nas patas traseiras nos grupos TCF e TAF. CONCLUSÃO: A administração de EA ou sua associação ao treinamento aeróbio não aumenta a massa muscular e força. Porém, à associação ao treinamento de força leva a maior hipertrofia muscular em fibras glicolíticas. Portanto, o tipo de treinamento físico, recrutamento muscular e características das fibras musculares, parecem ter importante impacto sobre as respostas anabólicas induzidas pelo E