Most species of pygmy-owls are partially diurnal, and so species that occur in low or open woodlands often are easy to see. In contrast, although the Amazonian Pygmy-Owl frequently is active during the day, it inhabits the canopy of tall humid lowland forest, and so is heard far more often than it is seen. The song of most species of Glaucidium consists of a series of hollow hooting notes, but the song of Amazonian is a short rapid series of whistled notes, forming almost a trill. This species overlaps geographically with Ferruginous Pygmy-Owl (Glaucidium brasilianum), but the two species are separated by habitat, with Amazonian in forest interior and Ferruginous at the edge of forest and in lower, more open river-edge forest. As is typical of Glaucidium, the Amazonian Pygmy-Owl hunts for invertebrates and small vertebrates, but there is very little information available about its biology.