The leonardo gene encodes a conserved member of the 14-3-3 protein family, which plays a role in Drosophila learning. Immunological localization of the protein shows that it is expressed at synaptic connections and enriched in presynaptic boutons of the neuromuscular junction (NMJ). Null leonardo mutants die as mature embryos. Electrophysiological assays of the mutant NMJ demonstrate that basal synaptic transmission is reduced by 30% and that transmission amplitude, fidelity, and fatigue resistance properties are reduced at elevated stimulation frequencies and in low external [Ca2+]. Moreover, transmission augmentation and post-tetanic potentiation (PTP) are disrupted in the mutant. These results suggest that Leonardo plays a role in the regulation of synaptic vesicle dynamics, a function which may underlie synaptic modulation properties enabling learning.