Removal of the superior colliculus (SC) in neonatal Wistar rats results in a rapid loss of retinal ganglion cells (RGCs). There is an early twofold increase in RGC death 4–8 hr postlesion (PL) followed by a later 10–11-fold increase in pyknosis about 24 hr PL. We have now used neurotrophic factors (BDNF, NT-4/5, NT-3, NGF, LIF), glutamate receptor antagonists (MK-801, DNQX, CNQX), an antioxidant (N-ace-tyl-L- cysteine), and an NOS inhibitor (L-NAME) to determine whether the early and late phases of lesion-induced RGC death involved similar or different mechanisms. Normal and pyknotic nuclei of tectally projecting RGCs were visualized by injecting the left s.c. of 2 d old rats with diamidino yellow (DY). Two days later the injection site was removed. In most rats, right eyes were injected with factors immediately after the s.c. ablation. Rats were perfused either 6 or 24 hr PL. In the latter group a second intravitreal injection of the appropriate factor was sometimes made 12 hr PL. NT- 4/5 and BDNF significantly decreased RGC pyknosis 6 and 24 hr PL, whereas NT-3 was only protective 6 hr PL. LIF slightly reduced RGC death 24 hr PL, but NGF had no influence on RGC survival at either time point. NT-4/5 also reduced the rate of naturally occurring RGC death. MK-801, DNQX, CNQX, N-acetylcysteine, and L-NAME all prevented the early lesion-induced increase in RGC death but had no significant effect on RGC death measured 24 hr PL; none of these factors significantly reduced the rate of naturally occuring RGC death. Cycloheximide, shown previously to reduce RGC pyknosis 24 hr PL, did not prevent RGC death 6 hr PL. The data indicate that there are at least two mechanisms involved in RGC death after neonatal target ablation. The early increase is related to excitotoxic effects mediated by glutamate receptors and involves NOS and the production of free radicals. We found no evidence that RGC death measured 24 hr PL is dependent on these processes, but the later death does require protein synthesis and, most likely, the activation of an endogenous suicide program. NT-4/5 and BDNF protected RGCs from both types of lesion- induced death.