AbstractOptic axons were labelled with horseradish peroxidase to establish the presence of side branches and examine their distribution and morphology in the developing optic nerve of the quokka wallaby, Setonix brachyurus, the cat and rat at stages when axon numbers are at their peak. In each species, three quarters of the axons were essentially straight and lacked side branches. The remaining axons took significantly longer paths and bore side branches, mostly at points where axons undulated or changed direction. Side branches occurred at intervals of 2843 m, had lengths of 23 m and were usually simple rather than branched. A minority (1%) of the axons crossed diagonally between fascicles and two thirds of these had more side branches (interval: 1018 m) on the interfascicular portion than were found on the forward-directed axons. A small number of axons (0.01%) doubled back to grow retrogradely towards the eye, these axons also bore relatively more side branches (interval: 822 m), especially at points where the axons changed direction. Ultrastructural reconstruction showed that side branches resembled small axonal profiles and constituted 2% of the total axon number. It is suggested that side branches are involved in the fine-tuning of growth cone navigation. Most side branches are lost by adulthood, indicating their transient nature. The absence of retrogradely-directed axons from adults suggests that cells with such axons are removed by naturally occurring cell death.