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Conotoxins: Biochemical probes for ion channels and receptors
9th World Congress
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caeruleus venom, Bungarus multicinctus venom and its neurotoxic components ~t-bungarotoxin and fl-bungarotoxin, Crotalus durissus terrificus venom and its neurotoxic component crotoxin, and Oxyuranus scutellatus venom and its neurotoxic component taipoxin, all of which have phospholipase A 2 activity associated with them. Venom ('venom' will hereinafter denote 'venom' and 'toxin') was administered i.p., followed immediately by an i.p. injection of drug. Lethality was measured 24 hr later. The optimal protective dose of each drug with respect to each venom was first determined. Second, the drug-induced change in the LDSO of venom was measured. Third, the effect on lethality of the relative time of injection of drug with respect to the injection of venom was determined. Chloroquine increased the LDsoSof B. caeruleus venom, B. multicinctus venom and fl-bungarotoxin by 16, 5.0 and 17-fold, respectively, while having no effect on the LDsoS of the other venoms. Chlorpromazine also increased the LDsoS of only the same three venoms by 8.7, 2.6 and 3.8-fold, respectively. At high doses dexamethasone caused a 3.5-fold increase in the LDso of O. scutellatus venom and a 4.0-fold increase in the LD5o of taipoxin while having no effect on the LOsoS of the other venoms. Nicergoline increased the LDso of B. multicinctus venom by 4.5-fold and the LDso of fl-bungarotoxin by 3.9-fold while having no effect on the LDsoS of the other venoms, with the exception of B. caeruleus venom which has yet to be tested. Piracetam had no effect on any of the venoms. Primaquine increased the LDsoS of B. caeruleus venom, B. multicinctus venom and fl-bungarotoxin by 2.9, 6.0 and 3.9-fold, respectively while having no effect on the LOsoS of the other venoms. Quinacrine increased the LDso of B. multicinctus venom by 10.5-fold and the LDso of fl-bungarotoxin by 8.6-fold while having no effect on the LDsoSof the other venoms with the exception of B. caeruleus venom which has yet to be tested. The relationship between lethality and the time of injection of drug relative to the time of injection of venom was similar for all the drug-venom combinations tested. Protection from lethality was maximal when the drugs were administered immediately after the injection of a 100% fatal dose of venom. Protection was still present, although significantly reduced, when drugs were injected 15 min either before or after intoxication. A 30 min pre- or postintoxication drug treatment offered virtually no protection from lethality. I found that selected drugs significantly reduced the lethality in mice of some snake venoms and their presynaptic neurotoxic components if the drugs were administered soon before or after intoxication. Conotoxins: biochemical probes for ion channels and receptors. LOURDESJ. CRUZ (Department of Biochemistry, College of Medicine, University of the Philippines, Manila, and Dept. of Biology, University of Utah, Salt Lake City, UT 84112, U.S.A.). CONOTOXINS are a group of neurotoxic peptides found in the venom of marine snails of the Conus genus. Currently in use as tools in neurobiology are the major paralytic toxins from fish-hunting species: o~-conotoxins which block voltage-sensitive calcium channels at nerve endings, ct-conotoxins which inhibit the acetycholine receptor at the postsynaptic terminus, and p-conotoxins which block muscle voltage-sensitive sodium channels. Representatives of each class have been chemically synthesized and radioiodinated for use in examining properties of the target molecules. Although ~t-conotoxins act like the ct-neurotoxins from elapid snake venoms, /~-conotoxins are the only peptides known to compete with the guanidinium neurotoxins (TTX and STX), thus blocking sodium channels, and ¢o-conotoxins are the first peptide toxins shown to block calcium channels. In the past several years ta-conotoxin GVIA has been used extensively in studying neuronal calcium channels. GVIA is particularly attractive as a biochemical probe because it has a very high affinity for the receptor (Ko< 10-12M) and it has a high degree of tissue specificity. Current biochemical, physiological and pharmacological data obtained in various laboratories indicate interaction of ~o-conotoxins with at least two subtypes of neuronal voltage sensitive calcium channels. Our recent data suggest that the various forms and derivatives of ~o-conotoxins isolated from C. geographus, C. magus and C. striatus will be useful in differentiating Ca channel subtypes.
Cloning o f cDNAs encoding snake toxins: sequence analysis and expression studies. FRI~DI~RIC DUCANCEL, GILBERTE GUIGNERY-FRELAT,TORU TAMIYA,JEAN-CLAUDEBOULAINand ANDP-d/Ml~NEZ(Service de Biochimie, D6partement de Biologie, C.E.N. Saclay, 91191 Gif/Yvette Cedex, France). Toxic PROTEINSpresent in most venoms of Elapidae and Hydrophiidae can be divided into two major structural classes. First, there are the non-enzymatic toxins with polypeptide chains folded into three loops emerging as broad fronds from a globular core rich in disulfide bonds. Acetylcholine receptor (AcChr) binding toxins, cardiotoxins and presumably anticholinesterase toxins have such an architecture. The second category comprises toxins with a phospholipase A 2 (PLA2) structure. Toxins which block the release of acetylcholine from nerve endings, myotoxins, etc., have a PLA 2 structure, with one or more subunits. We have cloned and sequenced cDNAs encoding the precursors of snake venom gland proteins belonging to the two structural classes. One of them codes for the precursor of an AcchR binding toxin. The other code for the precursors of PLA 2.
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caeruleus venom, Bungarus multicinctus venom and its neurotoxic components ~t-bungarotoxin and fl-bungarotoxin, Crotalus durissus terrificus venom and its neurotoxic component crotoxin, and Oxyuranus scutellatus venom and its neurotoxic component taipoxin, all of which have phospholipase A 2 activity associated with them. Venom ('venom' will hereinafter denote 'venom' and 'toxin') was administered i.p., followed immediately by an i.p. injection of drug. Lethality was measured 24 hr later. The optimal protective dose of each drug with respect to each venom was first determined. Second, the drug-induced change in the LDSO of venom was measured. Third, the effect on lethality of the relative time of injection of drug with respect to the injection of venom was determined. Chloroquine increased the LDsoSof B. caeruleus venom, B. multicinctus venom and fl-bungarotoxin by 16, 5.0 and 17-fold, respectively, while having no effect on the LDsoS of the other venoms. Chlorpromazine also increased the LDsoS of only the same three venoms by 8.7, 2.6 and 3.8-fold, respectively. At high doses dexamethasone caused a 3.5-fold increase in the LDso of O. scutellatus venom and a 4.0-fold increase in the LD5o of taipoxin while having no effect on the LOsoS of the other venoms. Nicergoline increased the LDso of B. multicinctus venom by 4.5-fold and the LDso of fl-bungarotoxin by 3.9-fold while having no effect on the LDsoS of the other venoms, with the exception of B. caeruleus venom which has yet to be tested. Piracetam had no effect on any of the venoms. Primaquine increased the LDsoS of B. caeruleus venom, B. multicinctus venom and fl-bungarotoxin by 2.9, 6.0 and 3.9-fold, respectively while having no effect on the LOsoS of the other venoms. Quinacrine increased the LDso of B. multicinctus venom by 10.5-fold and the LDso of fl-bungarotoxin by 8.6-fold while having no effect on the LDsoSof the other venoms with the exception of B. caeruleus venom which has yet to be tested. The relationship between lethality and the time of injection of drug relative to the time of injection of venom was similar for all the drug-venom combinations tested. Protection from lethality was maximal when the drugs were administered immediately after the injection of a 100% fatal dose of venom. Protection was still present, although significantly reduced, when drugs were injected 15 min either before or after intoxication. A 30 min pre- or postintoxication drug treatment offered virtually no protection from lethality. I found that selected drugs significantly reduced the lethality in mice of some snake venoms and their presynaptic neurotoxic components if the drugs were administered soon before or after intoxication. Conotoxins: biochemical probes for ion channels and receptors. LOURDESJ. CRUZ (Department of Biochemistry, College of Medicine, University of the Philippines, Manila, and Dept. of Biology, University of Utah, Salt Lake City, UT 84112, U.S.A.). CONOTOXINS are a group of neurotoxic peptides found in the venom of marine snails of the Conus genus. Currently in use as tools in neurobiology are the major paralytic toxins from fish-hunting species: o~-conotoxins which block voltage-sensitive calcium channels at nerve endings, ct-conotoxins which inhibit the acetycholine receptor at the postsynaptic terminus, and p-conotoxins which block muscle voltage-sensitive sodium channels. Representatives of each class have been chemically synthesized and radioiodinated for use in examining properties of the target molecules. Although ~t-conotoxins act like the ct-neurotoxins from elapid snake venoms, /~-conotoxins are the only peptides known to compete with the guanidinium neurotoxins (TTX and STX), thus blocking sodium channels, and ¢o-conotoxins are the first peptide toxins shown to block calcium channels. In the past several years ta-conotoxin GVIA has been used extensively in studying neuronal calcium channels. GVIA is particularly attractive as a biochemical probe because it has a very high affinity for the receptor (Ko< 10-12M) and it has a high degree of tissue specificity. Current biochemical, physiological and pharmacological data obtained in various laboratories indicate interaction of ~o-conotoxins with at least two subtypes of neuronal voltage sensitive calcium channels. Our recent data suggest that the various forms and derivatives of ~o-conotoxins isolated from C. geographus, C. magus and C. striatus will be useful in differentiating Ca channel subtypes.
Cloning o f cDNAs encoding snake toxins: sequence analysis and expression studies. FRI~DI~RIC DUCANCEL, GILBERTE GUIGNERY-FRELAT,TORU TAMIYA,JEAN-CLAUDEBOULAINand ANDP-d/Ml~NEZ(Service de Biochimie, D6partement de Biologie, C.E.N. Saclay, 91191 Gif/Yvette Cedex, France). Toxic PROTEINSpresent in most venoms of Elapidae and Hydrophiidae can be divided into two major structural classes. First, there are the non-enzymatic toxins with polypeptide chains folded into three loops emerging as broad fronds from a globular core rich in disulfide bonds. Acetylcholine receptor (AcChr) binding toxins, cardiotoxins and presumably anticholinesterase toxins have such an architecture. The second category comprises toxins with a phospholipase A 2 (PLA2) structure. Toxins which block the release of acetylcholine from nerve endings, myotoxins, etc., have a PLA 2 structure, with one or more subunits. We have cloned and sequenced cDNAs encoding the precursors of snake venom gland proteins belonging to the two structural classes. One of them codes for the precursor of an AcchR binding toxin. The other code for the precursors of PLA 2.