Dissertação de Mestrado
Avaliação da liberação de L-glutamato por sinaptosomas cérebro-corticais de rato, pela ação da peçonha da serpente coral sul-americana, Micrurus lemniscatus (linnaeus, 1758), e por algumas de suas frações semi-purificadas
Alessandra de Castro Montandon
The ophidic accidents attributed to the genus Micrurus sp., despite rare, are extremely severe and can led to death by respiratory arrest. The clinical manifestations of envenoming arise mainly from venom neurotoxic activity. Glutamate, the main exitatory neurotransmitter in CNS (central nervous system), when released in excess, causes neurotoxic effects, known as excitotoxicity. These effects are related to the pathogenesis of many neurodegenerative diseases. In this work, we investigated for the first time, the effect of Micrurus lemniscatus snake venom, as well as some of its semipurified fractions, on the release of L-glu (L-glutamate) from rat brain synaptosomes. After an screening of the activity of venom semi-purified fractions, it was found that those containing PLA2s were responsible for the most part of L-glu release. Two of these fractions, 20 and 31, composed according to mass analysis by -NTxs and PLA2s, respectively, induced the release time- dependent of L-glu from synaptosomes. Between the tested fractions having no PLA2s, only fraction 20, induced increased L-glu release. This fraction, unlike the venom and fraction 31, did not cause synaptosomes lysis or liposomes rupture. Sodium and potassium channels blockers, did not change L-glu release by venom and fraction 20; however, this release was partially affected in the presence of cadmium, an inespecific voltage-dependent calcium channels blocker. Also, it was not observed changes in L-glu release induced by the venom in the presence of D-tubocurarine, atropine and carbachol. However, L-glu release induced by venom and by fraction 20, was completely abolished when calcium was replaced by strontium in the incubation medium. M. lemniscatus venom, as well as fraction 31 exhibited high phospholipasic activity when compared to C. durissus terríficus venom. The fraction 20, suggested to be a possible short chain -NTx, showed no phospholipase activity. Thus, the results of this study suggest that the central neurotoxicity exerted in vitro by M. lemniscatus venom may be related to excessive release of L-glu in the CNS, caused mainly by the activity of PLA2s present in it. The fraction 20, may be associated with the central neurotoxicity symptoms found in patients envenomed by this snake, since the PLA2s, apparently, do not cross the blood brain barrier. The increased L-glu release induced by the venom and by this fraction, seems to involve, at least in part, the contribution of voltage-dependent calcium channels, considering the partial inhibition of this effect by cadmium. Moreover, the presence of calcium ions seems to be essential in this process, considering the total inhibition of L-glu release by strontium.