Bisphenol A (BPA), a chemical widely used to manufacture plastics, is estrogenic and capable of disrupting sex differentiation. However, recent in vitro studies have shown that BPA can also antagonize T(3) activation of the T(3) receptor. The difficulty in studying uterus-enclosed mammalian embryos has hampered the analysis on the direct effects of BPA during vertebrate development. This study proposed to identify critical T(3) pathways that may be disrupted by BPA based on molecular analysis in vivo. Because amphibian metamorphosis requires T(3) and encompasses the postembryonic period in mammals when T(3) action is most critical, we used this unique model for studying the effect of BPA on T(3)-dependent vertebrate development at both the morphological and molecular levels. After 4 d of exposure, BPA inhibited T(3)-induced intestinal remodeling in premetamorphic Xenopus laevis tadpoles. Importantly, microarray analysis revealed that BPA antagonized the regulation of most T(3)-response genes, thereby explaining the inhibitory effect of BPA on metamorphosis. Surprisingly, most of the genes affected by BPA in the presence of T(3) were T(3)-response genes, suggesting that BPA predominantly affected T(3)-signaling pathways during metamorphosis. Our finding that this endocrine disruptor, well known for its estrogenic activity in vitro, functions to inhibit T(3) pathways to affect vertebrate development in vivo and thus not only provides a mechanism for the likely deleterious effects of BPA on human development but also demonstrates the importance of studying endocrine disruption in a developmental context in vivo.