There is considerable evidence that nitric oxide activity is essential for memory formation, particularly from studies using inhibitors of nitric oxide synthase. The particular stage of memory formation requiring nitric oxide activity has not, however, been systematically investigated. In the current experiments, day-old black Australorp-white Leghorn chicks were trained on a passive avoidance task. Intracranial injections of the nitric oxide synthase inhibitor, l-NG-nitroarginine methylester (0.5 mM), were found to inhibit memory formation shortly after training, when injected pre- or posttraining. This effect was replicated with a second inhibitor, l-NG-nitroarginine (1 mM), and counteracted by the NO donor, sodium nitroprusside (150 microM). These findings provide covergent evidence that nitric oxide activity plays a critical role in the consolidation of memory in the day-old chick.