A late phase of exocytosis from synaptosomes induced by elevated [Ca2+](i) is not blocked by Clostridial neurotoxins

Abstract

Treatment of rat cerebrocortical synaptosomes with botulinum toxin types E and C1 or tetanus toxin removed the majority of intact SNAP-25, syntaxin 1A/1B, and synaptobrevin and diminished Ca2+-dependent K+ depolarization- induced noradrenaline secretion. With botulinum toxin type E, <10% of intact SNAP-25 remained and K+-evoked release of glutamate and GABA was inhibited. The large component of noradrenaline release evoked within 120 s by inclusion of the Ca2+ ionophore A23187 with the K+ stimulus was also attenuated by these toxins; additionally, botulinium neurotoxin type E blocked the first 60 s of ionophore-induced GABA and glutamate exocytosis. However, exposure to A23187 for longer periods induced a phase of secretion nonsusceptible to any of these toxins (>120 s for noradrenaline; >60 s for glutamate or GABA). Most of this late phase of release represented exocytosis because of its Ca2+ de- pendency, ATP requirement, and sensitivity to a phosphatidylinositol 4- kinase inhibitor. Based on these collective findings, we suggest that the ionophore-induced elevation of [Ca2+](i) culminates in the disassembly of complexes containing nonproteolyzed SNAP receptors protected from the toxins that can then contribute to neuroexocytosis.