[ON THE NEUROTOXINS OF 2 SPECIES OF NORTH AFRICAN SCORPIONS. II. PROPERTIES OF THE NEUROTOXINS (SCORPAMINES) OF ANDROCTONUS AUSTRALIS (L.) AND BUTHUS OCCITANUS (AM.)].
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The lytic activity of parabutoporin (PP) and opistoporin 1 (OP1) on mammalian and bacterial membranes have been described. We investigated pore-formation and ion selectivity in cardiac myocytes by measuring the whole cell leak current by means of the patch clamp technique. Pore formation was observed as the induction of leak currents. Ion selectivity of the pores was indicated by the shift of the reversal potential (E(rev)) upon substitution of intra- and extra-cellular ions. Results were compared with the effect of gramicidin A (gramA). PP and OP1 induced a fluctuating leak current and indicate non-selectivity of PP-induced pores. PP- and OP1-induced pores are between 1.38 and 1.78 nm in diameter.
In the present investigation we used native and recombinant TsNTxP to elicit antibodies in three different animal models (mouse, rabbit and sheep). Differences among anti-TsNTxP antibodies were analyzed using sets of overlapping pentadecapeptides of the TsNTxP amino acid sequence and also modified peptides to reveal key residues in antibody-peptide binding. Despite the identification of similar peptides by the antibodies in the C and N-terminal, peculiarities of each system were observed including the level of reactivity and also the number and type of key residues in the continuous epitopes of TsNTxP. In addition, in vitro neutralization assays indicated that sheep are an alternative and efficient model for the production of anti-Tityus serrulatus venom.
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The experiments were conducted to identify the toxin that produces pulmonary oedema in Mesobuthus tamulus (BT) envenomed animals. Crude BT venom was subjected to Sephadex gel filtration (G-75) and the fractions were screened for optical density (OD), neurotoxicity (prolongation of compound action potential in frog sciatic nerve) and lethality. All these parameters exhibited a peak between 54-94 ml eluates. Fractions of this peak were pooled (SP) and loaded on to carboxymethyl cellulose column. The column was then eluted with increasing buffer concentrations at constant pH and temperature. Eluates were screened for neurotoxicity and OD. Four peaks of neurotoxic activity (T1-T4) were detected. T2 and T3 were lethal whereas T1 and T4 were non-lethal. T2 exhibited mainly neurotoxicity and failed to augment phenyldiguanide (PDG)-induced reflex response or to produce pulmonary oedema. T3 was having minimal neurotoxic actions but augmented PDG-reflex and produced pulmonary oedema. The effects of T3 persisted even after dialysis with 8 kDa cut-off filter but not those of T2. The T3 effects resembled toxic manifestations of BT venom and were blocked by aprotinin pre-treatment. T3 demonstrated a band at approximately 100 kDa in SDS-PAGE. The results demonstrate the presence of a lethal, high molecular weight, pulmonary oedema producing toxin in BT venom.
Tityus serrulatus venom (Tsv) was intraperitoneally (ip) injected at doses of 75, 150 and 300mug/kg and IL-1beta (2.0 microg/kg) was given intravenously (iv) to male Wistar rats. Rectal temperature was measured by radiotelemetry. Vagotomy was performed according to Bluthe et al. [1994. Lipopolysaccharide induces sickness behaviour in rats by a vagal mediated mechanism. C R Acad. Sci. 317(6), 499-503]. Cerebrospinal fluid (CSF) and peritoneal fluid (PF) levels of bradykinin (BK) were measured by ELISA. B(1) (des-Arg(9)-[Leu(8)]-BK; DALBK) and B(2) kinin receptor (icatibant) antagonists (1.0 mg/kg each), the induced nitric oxide synthase inhibitor aminoguanidine (50.0 mg/kg), the neuronal nitric oxide synthase inhibitor 7-nitroindazole (30.0 mg/kg), the dual cyclooxygenase inhibitor ibuprofen (10.0 mg/kg), the selective interleukin-1 receptor antagonist IL-ra (2.0 mg/kg) and dipyrone (120 mg/kg) were given ip. Celecoxib (5 mg/kg) was given per os (po). Tsv at doses of 75 microg/kg evoked no change in rectal temperature while at doses of 150 and 300 microg/kg it promoted long-lasting fever (2 degrees C+/-0.1). Tsv (150 microg/kg) increased by nearly 3 and 5 times, respectively BK concentration in the CSF and in the PF. Subdiaphragmatic vagotomy or 7-nitroindazole reduced, icatibant, DALBK, IL-1ra, aminoguanidine and dipyrone abolished, while ibuprofen and celecoxib failed to affect Tsv-induced fever. These results suggest that PGs do not play a relevant role, whereas, kinins via their B(1) and B(2) receptors, IL-1, nitric oxide and vagal neurotransmission are involved in Tsv-induced fever.
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