Red spruce (Picearubens Sarg.) seedlings were treated with one of four concentrations of NH4NO3 (0, 300, 1500, and 3000 kg N•ha−1•year−1) applied to the soil, with and without triple superphosphate, during early, mid-, or late summer. Laboratory freezing assessments indicated that cold tolerance of treated seedlings generally increased with increasing nitrogen (N) uptake, with the exception of the highest N treatment. Seedlings receiving 1500 kg N•ha−1•year−1 were most cold tolerant on most sample dates. In general, these seedlings were hardier than those receiving 300 kg N•ha−1•year−1, which were hardier than unfertilized control seedlings. Seedlings receiving supplemental N also acclimated to cold more rapidly in autumn and deacclimated more slowly in spring than unfertilized controls. Supplemental phosphorus (P) had no influence on cold tolerance, and there was no evidence of a N × P interaction. Significant differences in cold tolerance associated with time of N application (early, mid-, and late summer) were detected in autumn and winter, but not in spring. In general, seedlings receiving N in mid- or late summer were as hardy or hardier than seedlings fertilized in early summer, regardless of the concentration of fertilizer. Significant interactions between N and timing of treatments occurred primarily because N applied in early summer resulted in only a slight increase in cold tolerance, whereas mid- and late summer N application resulted in a substantial increase in cold tolerance. Combined results suggest that it is highly unlikely that either the amount or timing of atmospheric N deposition is responsible for the winter injury frequently observed in red spruce.