Differentiated oligodendrocytes express neurite growth inhibitory proteins at a time when these cells are involved in the myelination of recently formed fiber pathways. As the process of myelination follows the completion of neurite outgrowth and is concurrent with the stabilization of fiber numbers in a pathway, we set out to determine whether myelination and fiber tract stability could be causally related. Myelin formation was prevented in the rat retinofugal pathway by x-irradiating the optic nerves during oligodendrocyte proliferation. Electron microscopic and immunohistochemical analysis of irradiated optic nerves at P15 showed that oligodendrocytes and myelin were virtually absent. Optic fiber numbers were determined at 2 weeks of age throughout the length of normal and x-irradiated nerves. In some cases, normal or irradiation-treated pups were intraocularly injected with FGF 5 d prior to the fiber count in order to promote neurite outgrowth. Axon counts showed that the total fiber number in a myelin-free optic nerve was 10–30% higher than that of a myelinated nerve. Further, fiber numbers fluctuated by as much as 20% along the length of a myelin-free nerve but were relatively constant throughout the length of a myelinated nerve. Treatment of myelinated nerves with fibroblast growth factor (FGF) had no effect on either total fiber numbers or fiber number fluctuation. Conversely, fiber numbers in myelin-free/FGF- treated optic nerves were as much as 40% higher than in normals. Furthermore, total fiber numbers along the length of these nerves fluctuated by up to 34%. These results indicate that, in the absence of myelination, optic fibers are able to form sprouts.(ABSTRACT TRUNCATED AT 250 WORDS)