We have developed a model system, consisting of rat brain synaptic vesicles and rat brain synaptic plasma membrane vesicles, to study the fusion process associated with the exocytotic release of neurotransmitters. Our results show a significant increase in the extent of fusion when the reaction takes place in cytosol compared to that obtained when fusion is carried out in buffer. This effect is mediated by cytosolic proteins, although N-ethylmaleimide-sensitive factor does not play a role in fusion. We also registered an almost complete inhibition of fusion when synaptic vesicles were pre-incubated with botulinum toxin B, indicating that synaptobrevin plays an important role in the coalescence of membrane lipids of the interacting membranes.