We have previously demonstrated that both peripheral and central neurons from 8 day embryonic chick and newborn mouse can be maintained in a serum-free medium using the N1 supplement consisting of insulin, transferrin, putrescine, progesterone, and selenite. In the present studies we show that dorsal root ganglionic (DRG) neurons from embryonic chick (E7-E15) and neonatal mouse can be cultured in a serum-free environment with only the addition of insulin and transferrin, plus Nerve Growth Factor (NGF). Chick DRG from E10-E15 contain a population of neurons sensitive to a chick embryo eye-derived ganglionic neuronotrophic factor (GNTF), which is distinct from the neuronal subset dependent upon NGF. The GNTF-dependent chick neurons can also be maintained in culture with insulin and transferrin supplements. Neonatal mouse DRG neurons, whether supported by NGF or eye-derived GNTF, likewise survive in serum-free medium with only insulin and transferrin. Limited numbers of neurons survive for the first 24 hours in a serum-free medium lacking insulin or transferrin, but failed to display neurite outgrowth even in the presence of added trophic factor.