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In 1995, we discovered a new pathway in the cat, which originates from the nucleus retroambiguus (NRA) and terminates in a distinct set of lumbosacral hindlimb, axial, and pelvic floor motoneuronal cell groups [VanderHorst VG.JM, Holstege G (1995) Caudal medullary pathways to lumbosacral motoneuronal cell groups in the cat: evidence for direct projections possibly representing the final common pathway for lordosis. J Comp Neurol 359:457-475]. The NRA is a compact group of interneurons located laterally in the caudal medulla oblongata. Its projection to lumbosacral moloneurons is thought to represent the final common pathway for male mounting and for female receptive or lordosis behavior. However, females only display lordosis behavior when they are in estrus, which suggests that the NRA-lumbosacral pathway is only active during estrus. This raised the question of whether estrogen affects this pathway. The effect of estrogen on the NRA-lumbosacral projection was studied light microscopically, using wheat-germ agglutinin horseradish peroxidase (WGA-HRP) as a tracer. The rubrospinal pathway served as control. The density of labeled NRA fibers in their target hindlimb motoneuronal cell groups appeared abundant in estrous and very weak in nonestrous cats. Such differences were not found in the rubrospinal pathway. For electron microscopical study, the NRA projection to the semimembranosus motoneuronal cell group was selected. In this cell group, an almost ninefold increase of labeled profiles was found in estrous versus nonestrous cats. Moreover, the semimembranous motoneuronal cell group contained labeled growth cones in estrous, but not in nonestrous, cats. The present study is the first to show that estrogen induces axonal outgrowth of a precisely identified pathway in the adult mammalian central nervous system. The possible mechanisms underlying this outgrowth are discussed.