title: Does extracellular calcium determine what pool of GABA is the target for alpha-latrotoxin?
abstract: Presynaptic neurotoxin alpha-latrotoxin, from the venom of Latrodectus mactans tredecimguttatus, causes massive [(3)H]GABA release from rat brain synaptosomes, irrespective of calcium presence in the extracellular medium. Whether the binding of alpha-latrotoxin to Ca(2+)-dependent (neurexin 1 alpha) or to Ca(2+)-independent (latrophilin) receptor triggers [(3)H]GABA release by the same mechanisms or different ones, inducing either exocytotic process or outflow by mobile membrane GABA transporter, is unknown. We examined alpha-latrotoxin-evoked [(3)H]GABA release from synaptosomes which cytosolic [(3)H]GABA pool was depleted either by applying competitive inhibitors of the GABA transporter, nipecotic acid and 2,4-diaminobutyric acid, or by permeation with digitonin. We also compared the effect of the GABA transporter inhibitors on depolarisation-evoked and alpha-latrotoxin-evoked [(3)H]GABA release using as depolarising agents 4-aminopyridine and high KCl in the Ca(2+)-containing and in Ca(2+)-free medium, respectively. Incubation of synaptosomes with nipecotic acid induced the essential acceleration of unstimulated [(3)H]GABA release and deep inhibition of high KCl-evoked Ca(2+)-independent [(3)H]GABA release. In contrast, at the similar conditions the effect of alpha-latrotoxin was greatly augmented with respect to the control response. Another way to assay what GABA pool was involved in alpha-latrotoxin-induced release lays in an analysis of the effects of depolarisation and alpha-latrotoxin in consecutive order. The preliminary 4-aminopyridine-stimulated [(3)H]GABA release attenuated the toxin effect. But when depolarisation occurred in Ca(2+)-free medium, no influence on alpha-latrotoxin effect was revealed. Employing digitonin-permeated synaptosomes, we have shown that alpha-latrotoxin could stimulate [3H]GABA release in the medium with 1mM EGTA, this effect of the toxin was blocked by concanavalin A and was ATP-dependent. The latter suggests that alpha-latrotoxin-released neurotransmitter has the vesicular nature. We assume that the type of the toxin membrane receptor does not determine the mechanisms of [(3)H]GABA release evoked by alpha-latrotoxin.
authors.affiliation: Department of Neurochemistry, Palladin Institute of Biochemistry, National Academy of Science of Ukraine, St. Leontovich 9, Kiev 01601, Ukraine.
authors.forename: L G
authors.initials: LG
authors.lastname: Storchak
authors.forename: M V
authors.initials: MV
authors.lastname: Linetska
authors.forename: N H
authors.initials: NH
authors.lastname: Himmelreich
mesh_terms: D000613:Aminobutyrates / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000502:physiology*; D002462:Cell Membrane / Q000502:physiology; D003600:Cytosol / Q000378:metabolism; D004594:Electrophysiology; D005110:Extracellular Space / Q000378:metabolism*; D008297:Male; D009498:Neurotoxins / Q000494:pharmacology*; D009557:Nipecotic Acids / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013111:Spider Venoms / Q000494:pharmacology*; D013574:Synaptosomes / Q000378:metabolism; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D000613:Aminobutyrates; D009498:Neurotoxins; D009557:Nipecotic Acids; D013111:Spider Venoms; C030278:nipecotic acid; C005959:2,4-diaminobutyric acid; D005680:gamma-Aminobutyric Acid; C020763:alpha-latrotoxin; D002118:Calcium