Spatial distribution of omega-agatoxin IVA binding sites in mouse brain slices

A peptide toxin derived from funnel-web spider venom, omega-agatoxin IVA, blocks voltage-sensitive calcium channels. Many pharmacological and electrophysiological studies have shown that these channels are widely distributed in both the central nervous system (CNS) and neuromuscular junctions. However, a direct morphological demonstration of the binding sites of this toxin is still lacking. To identify which cells have the binding sites, a biologically active, biotin-conjugated omega-agatoxin IVA was applied to mouse cerebellar and hippocampal slices. Confocal microscopy revealed that omega-agatoxin IVA binding sites were distributed on the somata of Purkinje cells, cerebellar granule cells and interneurons, as well as on the dendrites of Purkinje cells. In the hippocampus, the binding sites were localized on the somata of pyramidal cells of the CA1-CA4 region and on the somata of granule cells in the dentate gyrus. A sequential competitive reaction confirmed the specificity of the binding in the cerebellum and CA1 pyramidal cells, and also suggested a difference in the binding affinity between CA1 and CA3 pyramidal cells. Since a high concentration of omega-agatoxin IVA (2 microM) was needed for the present study, the omega-agatoxin IVA binding sites presented in this study may represent "P-type" and "Q-type" calcium channels.