Retinoic acid (RA), an active metabolite of vitamin A, is a diffusible molecule that regulates the expression of several families of genes, playing a key role in specification processes during chordate development. With the aim of defining its possible role in the developing chick inner ear, we obtained in this work a detailed spatiotemporal distribution of the enzymes involved in its synthesis, the retinaldehyde dehydrogenases (RALH1–4). Our results showed that, in contrast to the mouse inner ear, Raldh3 expression was the only Raldh gene detected in the developing chick inner ear, where it appears as early as stage 18. During inner ear morphogenesis, Raldh3 expression was predominantly observed in the endolymphatic system. The Raldh3 expression pattern delimited totally or partially the Bmp4-positive presumptive territories of vestibular sensory epithelia by stage 24 and the basilar papilla at stage 34, suggesting a possible involvement of RA in their specification. In addition, several vestibular sensory areas showed some Raldh3-expressing cells close to the Raldh3-positive domain. These results suggest that the RA signaling pathway may play a role in the initial patterning of the otic epithelium and cell differentiation therein, providing local positional information. Having in mind this Raldh3 expression pattern, we discuss the regulatory interactions among the RA, bone morphogenetic protein, and fibroblast growth factor signaling pathways in the specification of otic sensory elements. Our investigation may underpin further experimental studies aimed at understanding the possible role of signaling pathways in patterning of the developing chick inner ear. J. Comp. Neurol. 514:49–65, 2009. © 2009 Wiley-Liss, Inc.