We have isolated and characterized a new excitatory toxin from the venom of the sea anemone Bunodosoma caissarum, named Bc2. We investigated the mechanism of action of the toxin on Ca(2+)-regulated exocytosis in single bovine adrenal chromaffin cells, monitoring simultaneously fura-2 fluorescence measurements and electrochemical recordings using a carbon fiber microelectrode. Bc2 induced quantal release of catecholamines in a calcium-dependent manner. This release was associated with a sustained rise in cytosolic Ca(2+) and displayed two different patterns of response: a continuous discharge of prolonged duration that changed to a transient burst as the toxin concentration (or incubation time) increased. Continuous secretion was dependent on the activity of native voltage-dependent Ca(2+) channels and showed a pattern similar to that of alpha-latrotoxin; however, its kinetics adjusted better to that of continuous cell depolarization with high K(+) concentration. In contrast, transient secretion was independent of Ca(2+) entry through native voltage-dependent Ca(2+) channels and showed inhibition of late vesicle fusion that was accompanied by "freezing" of F-actin disassembly. These new features make Bc2 a promising new tool for studying the machinery of neurotransmitter release.