Morphological and physiological characteristics of the two major motor axons supplying the commonly studied ventral longitudinal muscle fibers (6 and 7) of third-instar Drosophila melanogaster larvae were investigated. The innervating terminals of the two motor axons differ in the size of their synapse-bearing varicosities. The terminal with the larger varicosities also fluoresces more brightly when stained with the vital fluorescent dye 4-(4-diethylaminostyryl)-N-methylpyridinium iodide (4-Di-2-Asp) and occupies a larger total contact area on the muscle fiber. Through selective simultaneous recording of synaptic currents from identified boutons in living preparations during elicitation of synaptic potentials, it was shown that the axon with the smaller varicosities generates a large excitatory junction potential (EJP) in muscle 6 and that the axon with the larger varicosities generates a smaller EJP. Short-term facilitation is more pronounced for the smaller EJP. In preparations treated with 4-Di-2-Asp, the fluorescence of smaller varicosities increases with stimulation that elicits the large EJPs, indicating an activity-dependent entry of calcium that enhances mitochondrial fluorescence. The differences in morphology and physiology of the two axons are similar to, though less pronounced than, those observed in "phasic" and "tonic" motor axons of crustaceans.