Analgesic effects of Phα1ß toxin: a review of mechanisms of action involving pain pathways

Phα1ß is a neurotoxin purified from spider venom that acts as a high-voltage-activated (HVA) calcium channel blocker. This spider peptide has shown a high selectivity for N-type HVA calcium channels (NVACC) and an analgesic effect in several animal models of pain. Its activity was associated with a reduction in calcium transients, glutamate release, and reactive oxygen species production from the spinal cord tissue and dorsal ganglia root (DRG) in rats and mice. It has been reported that intrathecal (i.t.) administration of Phα1ß to treat chronic pain reverted opioid tolerance with a safer profile than ω-conotoxin MVIIA, a highly selective NVACC blocker. Following a recent development of recombinant Phα1ß (CTK 01512-2), a new molecular target, TRPA1, the structural arrangement of disulphide bridges, and an effect on glial plasticity have been identified. CTK 01512-2 reproduced the antinociceptive effects of the native toxin not only after the intrathecal but also after the intravenous administration. Herein, we review the Phα1ß antinociceptive activity in the most relevant pain models and its mechanisms of action, highlighting the impact of CTK 01512-2 synthesis and its potential for multimodal analgesia.

Analgesic effects of Phalfa1 beta toxin: a review of mechanisms of action involving pain pathways

Phα1ß is a neurotoxin purified from spider venom that acts as a high-voltage-activated (HVA) calcium channel blocker. This spider peptide has shown a high selectivity for N-type HVA calcium channels (NVACC) and an analgesic effect in several animal models of pain. Its activity was associated with a reduction in calcium transients, glutamate release, and reactive oxygen species production from the spinal cord tissue and dorsal ganglia root (DRG) in rats and mice. It has been reported that intrathecal (i.t.) administration of Phα1ß to treat chronic pain reverted opioid tolerance with a safer profile than ω-conotoxin MVIIA, a highly selective NVACC blocker. Following a recent development of recombinant Phα1ß (CTK 01512-2), a new molecular target, TRPA1, the structural arrangement of disulphide bridges, and an effect on glial plasticity have been identified. CTK 01512-2 reproduced the antinociceptive effects of the native toxin not only after the intrathecal but also after the intravenous administration. Herein, we review the Phα1ß antinociceptive activity in the most relevant pain models and its mechanisms of action, highlighting the impact of CTK 01512-2 synthesis and its potential for multimodal analgesia.

Effects of Cylindrospermopsis raciborskii strains (Woloszynska, 1912) Senayya & Subba Raju on the mobility of Daphnia laevis (Cladocera, Daphniidae) / Efeitos de cepas de Cylindrospermopsis raciborskii (Woloszynska, 1912), Senayya & Subba Raju na mobilidade de Daphnia laevis (Cladocera, Daphniidae)

Cylindrospermopsis raciborskii is a cyanobacterium distributed worldwide that is known to produce cyanotoxins. Some of the Brazilian strains can produce saxitoxins (STXs), which are classified as neurotoxins and can paralyze cladocerans .Daphnia laevis is a cladoceran with a wide distribution in the Americas and has been studied as a possible test-organism in toxicity bioassays. The present work tested the acute effect on D laevis mobility when fed a saxitoxin-producing (STX and neoSTX) C. raciborskii strain, CYRF-01, and compared the results with the effects of a non-toxic strain (NPCS-1). Neonates (6-24 hours after birth) were exposed to concentrations of C. raciborskii varying from 102 to 106 cells·mL–1 of each strain for up to three hours. The cladocerans were then transferred to a medium without toxic filaments for 24 hours. Only the organisms exposed to the STX-producing strain showed signs of the immobilization of swimming movements, confirming the effects of the toxins. There was a linear correlation between the time required to induce stopping the swimming movement, with a shorter time to needed to induce immobilization at a higher the concentration; this correlation was inverse to the time required to recover the swimming movements (longer at higher concentrations, p < 0.1). D. laevis is a tropical and subtropical species with great potential for use in toxicity tests for the detection of STXs, despite being native to and found in a great array of freshwater bodies. This is the first assay testing STX-producing and non-producing C. raciborskii strains on D. laevis, species that are both found in Brazilian ecosystems.

Cylindrospermopsis raciborskii é uma espécie de cianobactéria, difundida mundialmente, conhecida como produtora de cianotoxinas. Algumas linhagens brasileiras são conhecidas como produtoras de saxitoxinas (STXs), as quais são classificadas como neurotoxinas e podem induzir a paralização dos movimentos natatórios em cladóceros. Daphnia laevis é um cladócero de ampla distribuição nas Américas e tem sido estudado como um organismo teste para uso em ensaios de toxicidade. Neste sentido, o presente trabalho avaliou os efeitos agudos de C. raciborskii, de uma cepa produtora de STXs (STX e neoSTX) - CYRF-01 e outra não produtora de cianotoxinas (NPCS-1) sobre a mobilidade de D. laevis. Para tanto, neonatas entre 6-24 horas de idade foram submetidas a concentrações de C. raciborskii variando de 102 a 106 céls.mL–1 de ambas as cepas, por um período de 3 horas, e transferidos posteriormente para um meio isento de filamentos tóxicos por 24horas. Apenas os organismos expostos à cepa produtora de STXs apresentaram paralisia dos movimentos natatórios, confirmando o efeito da cianotoxina. Houve uma correlação linear entre o tempo de paralização e as concentrações, isto é, nas concentrações mais altas, os organismos paralisaram num curto período de tempo, e conseqüentemente, levaram mais tempo para se recuperar (p < 0,1). D.laevis é uma espécie de região tropical e subtropical com potencial uso em bioensaios para detecção de STXs. Trata-se dos primeiros relatos acerca da resposta de D.laevis quando submetida às cepas de C. raciborskii produtora e não produtora de STXs, ambas isoladas de ecossistemas brasileiros.

Síntese e aplicações estratégicas de neurotoxinas produzidas por cianobactérias / Synthesis and strategic applications of some neurotoxins produce for cyanobacteria

As cianobactérias apresentam, distribuição variada e podem ocorrer desde as regiões frias do ártico até os trópicos, em corpos d'água doce e no ambiente marinho. Algumas espécies de cianobactérias produzem compostos com conhecida toxidade, podendo causar efeitos deletérios em seres humanos e animais. Entre estes compostos estão os com atividade neurotóxica devido aos mais variados mecanismos de ação. O objetivo geral deste projeto é a obtenção por via sintética de algumas neurotoxinas e variantes, entre elas a β-N-metil-L-alanina (L-BMAA), anatoxina-a e anatoxina-a(s), bem como algumas aplicações. Para a L-BMAA, foi buscada a determinação de rotas sintéticas viáveis para a obtenção deste aminoácido modificado sob a forma racêmica e enantiomericamente pura, além de um possível produto cíclico que pode ser gerado naturalmente a partir da L-BMAA. Algumas rotas estratégicas foram determinadas com êxito para a síntese destes compostos. Entre as aplicações dos produtos obtidos podem ser citados: (i) a determinação de um método analítico para a determinação deste aminoácido por RMN de 1H; (ii) um método analítico por LC-MS utilizando D3-L-BMAA como padrão interno e (iii) a indicação de um possível mecanismo de neurotoxidade ligado a neurodegeneração via um intermediário produzido naturalmente do equilíbrio entre L-BMAA e íons bicarbonato...

Development of an immunodetection test for a botulinum-like neurotoxin produced by Clostridium sp. RKD.

BACKGROUND & OBJECTIVES: Clostridial neurotoxins are among the most toxic substances known and cause severe illnesses in both humans and animals. A neurotoxigenic Clostridium sp. (strain RKD) isolated from intestine of decaying fish produced a novel, botulinum type B like neurotoxin as suggested by mouse bioassay, protection with anti-botulinum antibodies and PCR. The aim of the present investigation was to develop a laboratory based detection assay as an alternative to the mouse bioassay without compromising sensitivity and specificity. METHODS: Growth and toxin production were carried out in trypticase peptone yeast-extract glucose (TPYG) broth. Toxicity was estimated in terms of minimum lethal dose (MLD) by mouse bioassay. The toxin was partially purified by acid precipitation. It was used for toxoid preparation by formaldehyde treatment. This purified IgG was used for detection of neurotoxin using indirect ELISA. The culture supernatant was concentrated using a stirred cell with a 50 kDa cut-off membrane at 4 degrees C. Further purification was carried out using Prep cell. Fractions showing toxicity and sufficient purity were pooled, concentrated and analyzed on sodium dodecyl sulphatepolyacrylamide gel electrophoresis (SDS-PAGE). RESULTS: The toxin was purified with a recovery of 8.56 per cent. Polyclonal antiserum was raised in mice using partially purified toxin with a titre of 1: 80000. A detection assay with sensitivity of approximately 15 and 300 ng/ml for partially purified and crude toxins, respectively were achieved using an indirect ELISA method. INTERPRETATION & CONCLUSION: The Clostridium sp. RKD produced a potent neurotoxin earlier shown to have novelties. A specific detection assay for the neurotoxin has been developed that may be useful both from food safety and clinical point of view.

Neurodepressive action of a piscicidal glycoside of plant, Aesculus indica (Colebr.) in fish.

Sublethal concentration (2.6 mg/l) of a triterpene based piscicidal glycoside of A. indica damaged the neurons, fibre tracts and central correlation sites for gustatory, tactile and visceral sensory impulses in medulla oblongata of fish after prolonged poisoning. The fear, sinking to bottom, lack of schooling and non discriminatory pattern in treated fish were because of neurodepression. The jerky movement, ventilatory inefficiency, swallowing air bubbles and light pinkness of gills were due to the degradation of respiratory centres in the vagal lobes of toxified fish.

Isolation and purification of a lethal scorpion toxin with neuromuscular blocking activity.

Toxin-L a lethal neuromuscular blocking agent was isolated from the venom of the scorpion, Lychas laevifrons (Pocock), by the CM-cellulose ion-exchange chromatography. It was a homogenous, thermolabile and low molecular weight protein. The toxin produced irreversible blockade of indirect stimulation induced twitch responses on innervated rat phrenic nerve-diaphragm and chick biventer cervicis preparation. The toxin did not produce any contractile response on toad rectus abdominis muscle preparation. On chronically denervated rat diaphragm, the toxin failed to alter the responses induced by direct stimulation, exogenous acetylcholine, potassium chloride and caffeine. Acetylcholine and carbachol induced contractions on isolated chick biventer cervicis remained unaltered by the toxin. Neostigmine failed to alter toxin induced neuromuscular blockade on innervated rat diaphragm. The toxin released a significant amount of acetylcholine from innervated rat diaphragm. It may be concluded that the toxin acts presynaptically through the release of acetylcholine, thereby producing neuromuscular blockade.