The presynaptic active zone is the critical region of a chemical synapse at which Ca2+ entry provokes neurotransmitter release by exocytotic fusion of synaptic vesicles. To facilitate investigations of synaptic function, we have identified a group of fluorescent substances that label individual active zones in living hair cells. The Ca2+ indicator fluo-3, the compound studied in most detail, binds to the presynaptic dense bodies that are characteristic of active zones in hair cells and other cells that tonically release transmitter. The indicator's binding is reversible, with a dissociation constant of approximately 350 microM. Because fluo-3 that is bound to a presynaptic dense body continues to detect Ca2+ with an unaltered dissociation constant, the binding of this substance provides a valuable tool for exploration of the Ca2+ concentration at the site of vesicle fusion.