Phyla- and Subtype-Selectivity of CgNa, a Na+ Channel Toxin from the Venom of the Giant Caribbean Sea Anemone Condylactis Gigantea

Because of their prominent role in electro-excitability, voltage-gated sodium (NaV) channels have become the foremost important target of animal toxins. These toxins have developed the ability to discriminate between closely related NaV subtypes, making them powerful tools to study NaV channel function and structure. CgNa is a 47-amino acid residue type I toxin isolated from the venom of the Giant Caribbean Sea Anemone Condylactis gigantea. Previous studies showed that this toxin slows the fast inactivation of tetrodotoxin-sensitive NaV currents in rat dorsal root ganglion neurons. To illuminate the underlying NaV subtype-selectivity pattern, we have assayed the effects of CgNa on a broad range of mammalian isoforms (NaV1.2–NaV1.8) expressed in Xenopus oocytes. This study demonstrates that CgNa selectively slows the fast inactivation of rNaV1.3/β1, mNaV1.6/β1 and, to a lesser extent, hNaV1.5/β1, while the other mammalian isoforms remain unaffected. Importantly, CgNa was also examined on the insect sodium channel DmNaV1/tipE, revealing a clear phyla-selectivity in the efficacious actions of the toxin. CgNa strongly inhibits the inactivation of the insect NaV channel, resulting in a dramatic increase in peak current amplitude and complete removal of fast and steady-state inactivation. Together with the previously determined solution structure, the subtype-selective effects revealed in this study make of CgNa an interesting pharmacological probe to investigate the functional role of specific NaV channel subtypes. Moreover, further structural studies could provide important information on the molecular mechanism of NaV channel inactivation

Antinociception produced by Thalassia testudinum extract BM-21 is mediated by the inhibition of acid sensing ionic channels by the phenolic compound thalassiolin B

<p>Abstract</p> <p>Background</p> <p>Acid-sensing ion channels (ASICs) have a significant role in the sensation of pain and constitute an important target for the search of new antinociceptive drugs. In this work we studied the antinociceptive properties of the BM-21 extract, obtained from the sea grass <it>Thalassia testudinum</it>, in chemical and thermal models of nociception in mice. The action of the BM-21 extract and the major phenolic component isolated from this extract, a sulphated flavone glycoside named thalassiolin B, was studied in the chemical nociception test and in the ASIC currents of the dorsal root ganglion (DRG) neurons obtained from Wistar rats.</p> <p>Results</p> <p>Behavioral antinociceptive experiments were made on male OF-1 mice. Single oral administration of BM-21 produced a significant inhibition of chemical nociception caused by acetic acid and formalin (specifically during its second phase), and increased the reaction time in the hot plate test. Thalassiolin B reduced the licking behavior during both the phasic and tonic phases in the formalin test. It was also found that BM-21 and thalassiolin B selectively inhibited the fast desensitizing (τ < 400 ms) ASIC currents in DRG neurons obtained from Wistar rats, with a nonsignificant action on ASIC currents with a slow desensitizing time-course. The action of thalassiolin B shows no pH or voltage dependence nor is it modified by steady-state ASIC desensitization or voltage. The high concentration of thalassiolin B in the extract may account for the antinociceptive action of BM-21.</p> <p>Conclusions</p> <p>To our knowledge, this is the first report of an ASIC-current inhibitor derived of a marine-plant extract, and in a phenolic compound. The antinociceptive effects of BM-21 and thalassiolin B may be partially because of this action on the ASICs. That the active components of the extract are able to cross the blood-brain barrier gives them an additional advantage for future uses as tools to study pain mechanisms with a potential therapeutic application.</p

The sea source of new drugs

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Compounds from marine sources as tools to study the nervous system

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Neuroprotective and antioxidant effects of Thalassia testudinum extract BM-21, against acrylamide-induced neurotoxicity in mice

Context: Acrylamide (ACR) neurotoxicity is associated with the enhancement of lipid peroxidation and the reduction of the antioxidative capacity distal axon and nerve terminal regions. The aqueous ethanolic extract of the marine plant Thalassia testudinum, named BM-21, have shown antioxidant, anti-inflammatory and analgesic properties. Aims: To determine the neuroprotective and the antioxidant effects of BM-21, standardized to thalassiolin B content (5.8 ± 0.9%), on acrylamide (ACR)-induced distal axonopathy in male OF-1 mice. Methods: Animals were administered with ACR (70 mg/kg, s.c., 4 weeks), and BM-21 was co-administered p.o at the doses of 4, 40 and 400 mg/kg. The effect of BM-21 on neurobehavioral indexes (rota-rod test), compound muscle action potential (CMAP) of the sciatic nerve and oxidative stress parameters were investigated. Results: BM-21 significantly prevented the neurobehavioral sings of neurotoxicity and the alteration of CMAP amplitude and velocity. The lowest dose (4 mg/kg) failed to ameliorate these parameters whereas the highest dose (400 mg/kg) was the most active. BM-21 (400 mg/kg) significantly restored total hydroperoxides (THP) and glutathione (GSH) in the sciatic nerve as well as superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities. Additionally, the extract also modified THP, GSH and the activity of SOD in cerebellum and brain towards the standard values. Conclusions: BM-21 given at doses that prevented ACR-induced neurotoxicity also produced antioxidant effect in the sciatic nerve, cerebellum and brain. Thus, the neuroprotective activity of BM-21 in this model seems to be mediated at least partly by its antioxidative properties

Neuroprotective effects of Thalassia testudinum leaf extract BM-21 on focal ischemia in rats

Context: The extract from the marine plant Thalassia testudinum BM-21, standardized to thalassiolin B content (5.8 ± 0.3%), possesses antioxidant, anti-inflammatory and neuroprotective effects on acrylamide-induced neurotoxicity in mice and global ischemia in Mongolian gerbils. Aims: To determine whether or not BM-21 possesses neuroprotective effects against cerebral ischemia induced by permanent middle cerebral artery occlusion (pMCAo), a clinically relevant model of stroke. Methods: BM-21 was administered orally (400 mg/kg, once-a–day/10 days) prior to ischemia. Twenty-four hours after occlusion, we studied neurological signs, infarct volume, cerebral edema, histological damage and oxidative stress in cortex and striatum. In addition, brain susceptibility to in vitro lipid peroxidation induced by kainic acid and 2,2′-azobis(2-amidinopropane) dihydrochloride was studied after the BM-21 administration. Results: BM-21 prevented behavioral deficit; reduced infarct volume and cerebral edema; markedly decreased neuronal damage in striatum and cortex region. After occlusion, there was a significant increase of oxidative stress in cortex and striatum. Treatment of ischemic rats with BM-21 (400 mg/kg) prevented lipid peroxidation and protein damage and increased the antioxidant enzymatic activities and glutathione. BM-21 also inhibited the in vitro lipid peroxidation in total brain homogenates. Conclusions: Oral pre-treatment of BM-21 protects rats against pMCAo ischemia-induced damage in the striatum and cortex. Results suggest that the protection of BM-21 involve at least partially, the increase resistance to oxidative stress

Peptide fingerprinting of the neurotoxic fractions isolated from the secretions of sea anemones Stichodactyla helianthus and Bunodosoma granulifera. New members of the APETx-like family identified by a 454 pyrosequencing approach

Sea anemones are known to contain a wide diversity of biologically active peptides, mostly unexplored according to recent peptidomic and transcriptomic studies. In the present work, the neurotoxic fractions from the exudates of Stichodactyla helianthus and Bunodosoma granulifera were analyzed by reversed-phase chromatography and mass spectrometry. The first peptide fingerprints of these sea anemones were assessed, revealing the largest number of peptide components (156) so far found in sea anemone species, as well as the richer peptide diversity of B. granulifera in relation to S. helianthus. The transcriptomic analysis of B. granulifera, performed by massive cDNA sequencing with 454 pyrosequencing approach allowed the discovery of five new APETx-like peptides (U-AITX-Bg1a-e - including the full sequences of their precursors for four of them), which together with type 1 sea anemone sodium channel toxins constitute a very distinguishable feature of studied sea anemone species belonging to genus Bunodosoma. The molecular modeling of these new APETx-like peptides showed a distribution of positively charged and aromatic residues in putative contact surfaces as observed in other animal toxins. On the other hand, they also showed variable electrostatic potentials, thus suggesting a docking onto their targeted channels in different spatial orientations. Moreover several crab paralyzing toxins (other than U-AITX-Bg1a-e), which induce a variety of symptoms in crabs, were isolated. Some of them presumably belong to new classes of crab-paralyzing peptide toxins, especially those with molecular masses below 2 kDa, which represent the smallest peptide toxins found in sea anemones. (C) 2011 Elsevier Inc. All rights reserved.CNPqCITMA (Brazil)CNPq-CITMA (Brazil) [490194/2007-9]FAPESP [07/56525-3]FAPESPFAPEMIGFAPEMIGINCTTOXINCTTOXCAPESCAPESCNPqCNPqScience and Technology Development Fund of Macau SARScience and Technology Development Fund of Macau SAR [058/2009]Research Committee, University of Macau [UL017/09-Y1]Research Committee, University of MacauInternational Foundation for Science [F/4082-1, F/4082-2]International Foundation for ScienceThird World Academy of Sciences [06344-2007]Third World Academy of Science

CgNa, a type I toxin from the giant Caribbean sea anemone Condylactis gigantea shows structural similarities to both type I and II toxins, as well as distinctive structural and functional properties1

CgNa (Condylactis gigantea neurotoxin) is a 47-amino-acid- residue toxin from the giant Caribbean sea anemone Condylactis gigantea. The structure of CgNa, which was solved by 1H-NMR spectroscopy, is somewhat atypical and displays significant homology with both type I and II anemone toxins. CgNa also displays a considerable number of exceptions to the canonical structural elements that are thought to be essential for the activity of this group of toxins. Furthermore, unique residues in CgNa define a characteristic structure with strong negatively charged surface patches. These patches disrupt a surface-exposed cluster of hydrophobic residues present in all anemone-derived toxins described to date. A thorough characterization by patch–clamp analysis using rat DRG (dorsal root ganglion) neurons indicated that CgNa preferentially binds to TTX-S (tetrodotoxin-sensitive) voltage-gated sodium channels in the resting state. This association increased the inactivation time constant and the rate of recovery from inactivation, inducing a significant shift in the steady state of inactivation curve to the left. The specific structural features of CgNa may explain its weaker inhibitory capacity when compared with the other type I and II anemone toxins