Antagonism of synaptosomal calcium channels by subtypes of omega-agatoxins.
Venom of the funnel web spider Agelenopsis aperta inhibits the binding of 125I-omega-conotoxin GVIA (omega-CgTx) to calcium channels in chick brain synaptosomal membranes. Fractionation of the venom by liquid chromatography shows that this inhibitory activity is associated primarily with a diverse class of peptide toxins called omega-agatoxins (omega-Aga). Using binding inhibition as an assay, we purified and identified the novel, 76-amino acid toxin, omega-Aga-IIIA. Inhibition of 125I-omega-CgTx binding to chick synaptosomal membranes by omega-Aga-IIIA and omega-Aga-IIA is correlated with block of potassium-stimulated 45Ca entry into synaptosomes; omega-Aga-IA neither inhibits 125I-omega-CgTx binding nor 45Ca entry under identical conditions. omega-Aga-IIA and omega-Aga-IIIA are 20-30-fold more potent than omega-CgTx as antagonists of synaptosomal calcium channels. However, whereas omega-CgTx completely blocks 45Ca entry into synaptosomes at saturating concentrations, the omega-agatoxins maximally block only 60-70% of 45Ca entry. Pretreatment of synaptosomes with omega-Aga-IIIA occludes block of 45Ca entry by omega-CgTx. The results indicate that, while the omega-agatoxins bind to the entire population of omega-CgTx-sensitive calcium channels in chick synaptosomal membranes, they exert only a partial block of 45Ca flux. Such block could occur via two distinct mechanisms. Toxin binding may alter the kinetics of a homogeneous population of channels, resulting in lower overall conductance upon depolarization. Alternatively, the omega-agatoxins may bind to two distinct channel subtypes, only one of which is blocked as a result of toxin occupation.
Venema
V J
VJ
Department of Entomology, University of California, Riverside 92521.
Swiderek
K M
KM
Lee
T D
TD
Hathaway
G M
GM
Adams
M E
ME
D060848:Agatoxins; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000302:isolation & purification / Q000494:pharmacology*; D015220:Calcium Channels / Q000187:drug effects*; D002645:Chickens; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D013111:Spider Venoms / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*; D013574:Synaptosomes / Q000187:drug effects* / Q000378:metabolism
D060848:Agatoxins; D000596:Amino Acids; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D013111:Spider Venoms; C070132:omega-agatoxin III