The requirement of vitamin A (retinol) for successful completion of vertebrate embryogenesis is well established. Retinoid signaling involves a two-step metabolic event in which retinol is first converted to retinal, and then retinal is converted to the active ligand retinoic acid, which modulates the transcriptional activity of a nuclear retinoic acid receptor (RAR). During mouse embryogenesis, retinoic acid is not detected at 6.5 days of embryonic development (E6.5) when gastrulation first initiates, but it is detected at E7.5 and later. This suggests that retinoid signaling during embryogenesis may be initiated during the primitive streak stage. Here we have used whole-mount in situ hybridization to examine E6.5-E8.5 mouse embryos for expression of RARa, RARb, RARg, and two enzymes, class IV alcohol dehydrogenase (ADH-IV) and class I aldehyde dehydrogenase (ALDH-I), which have been shown to have retinol and retinal dehydrogenase activities, respectively. At E6.5, RARa mRNA was expressed ubiquitously in embryonic and extraembryonic tissues, RARg mRNA was detected throughout all embryonic tissues, but mRNAs for RARb, ADH-IV, and ALDH-I were not detected. By E7.5, RARa mRNA was still ubiquitous, RARb mRNA was now observed in presumptive hindbrain ectoderm and adjacent mesenchyme, RARg mRNA was still observed in all embryonic tissues, and ADH-IV as well as ALDH-I mRNAs were now both expressed in primitive streak mesoderm. In E8.5 embryos, RARa mRNA was still ubiquitous, RARb mRNA was present in the caudal hindbrain as well as the closed neural tube and foregut, RARg mRNA was widespread but most prevalent in caudal embryonic tissues, and mRNAs for both ADH-IV and ALDH-I were expressed in cranial mesenchyme, somites, and paraxial mesoderm. Thus, ADH-IV and ALDH-I, two metabolic enzymes able to convert retinol to retinoic acid, are both initially expressed in primitive streak mesoderm at E7.5 when retinoic acid is first detectable. On the other hand, RARa and RARg expression is widespread and present at E6.5 prior to retinoic acid detection. These results suggest that upregulation of ADH-IV and ALDH-I gene expression in primitive streak mesoderm may lead to retinoic acid synthesis and initiation of retinoid signaling during mouse embryogenesis. Dev. Dyn. 208:536-543, 1997. r 1997 Wiley-Liss, Inc.