Imidacloprid is a neonicotinoid insecticide used to control agricultural pests around the world. This pesticide can have adverse effects on non-target organisms, especially in aquatic environments. The present study evaluated the toxicity of an imidacloprid-based insecticide in amphibians, using Leptodactylus luctator and Physalaemus cuvieri tadpoles as study models. Spawning of both species were collected within less than 24 h of oviposition from a non-agricultural land at Erechim, Rio Grande do Sul state, Brazil. Survival, swimming activity, body size, morphological malformations, and genotoxic parameters were analyzed at laboratory conditions. A short-term assay was conducted over 168 h (7 days) with five different concentrations of imidacloprid (3–300 µg L⁻¹) being tested. The insecticide did not affect survival, although the tadpoles of both species presented reduced body size, malformed oral and intestine structures, and micronuclei and other erythrocyte nuclear abnormalities following exposure to this imidacloprid-based compound. Exposure also affected swimming activity in L. luctator, which reflected the greater sensitivity of L. luctator to imidacloprid in comparison with P. cuvieri. The swimming activity, body size, and malformations observed in L. luctator and the morphological malformations found in P. cuvieri indicated that even the lowest tested concentration of the insecticide were harmful to amphibians. At concentrations of over 3 μg L⁻¹, P. cuvieri presents a smaller body size, and both species are affected by genotoxic cell damage. This demonstrates that imidacloprid is potentially toxic for the two study species at environmentally relevant concentrations.