Dopaminergic neurons have the capacity to release dopamine not only from their axon terminals, but also from their somatodendritic compartment. The actual mechanism of somatodendritic dopamine release has remained controversial. Here we established for the first time a rat primary neuron culture model to investigate this phenomenon and use it to study the mechanism under conditions of non-stimulated spontaneous firing (1-2 Hz). We found that we can selectively measure somatodendritic dopamine release by lowering extracellular calcium to 0.5 mm, thus confirming the previously established differential calcium sensitivity of somatodendritic and terminal release. Dopamine release measured under these conditions was dependent on firing activity and independent of reverse transport through the plasma membrane. We found that treatment with botulinum neurotoxins A and B strongly reduced somatodendritic dopamine release, thus demonstrating the requirement for SNARE proteins SNAP-25 and synaptobrevin. Our work is the first to provide such direct and unambiguous evidence for the involvement of an exocytotic mechanism in basal spontaneous somatodendritic dopamine release.