Quantitative axial profiles of retinoic acid in the embryonic mouse spinalcord: 9-cis retinoic acid only detected after all-trans-retinoic acid levels are super-elevated experimentally

Studies using bioassays in normal mice and gene activation in transgenic re porter mice have demonstrated peaks of retinoic acid receptor (RAR) signali ng in the brachial and lumbar regions of the spinal cord. Recently, Solomin et al. (Solomin et al. [1998] Nature 395:398-402) detected a retinoid X re ceptor (RXR) signal in the same region of the developing spinal cord at a s lightly later stage than the RAR signal. This finding raises the question o f which retinoid ligands underlie PAR and RXR signaling in this part of the embryo. Quantitative measurements of regional differences in retinoid prof iles have not been reported previously due to limitation in the sensitivity and specificity of available retinoid detection methods. Here, by using a recently developed ultrasensitive HPLC technique (Sakhi et al. [1998] J. Ch romatogr. A 828:451-460), we address this question in an attempt to identif y definitively the endogenous retinoids present in different regions of the spinal cord at the stages when regional differences in RAR and RXR signali ng have been reported. We find a bimodal distribution of all-trans retinoic acid (at-RA), the ligand for RARs, and relate this to the expression of se veral retinoid-synthesizing enzymes. However, we do not detect 9-cis-retino ic acid (9-cis-RA), the putative RXR ligand, in any region of the spinal co rd unless retinoid levels are massively increased experimentally by gavage feeding pregnant mice with teratogenic doses of at-RA. This study provides for the first time quantitative profiles of endogenous retinoids along the axis of the developing spinal cord, thereby establishing a foundation for m ore definitive studies of retinoid function in the future. It sets definite limits on how much 9-cis-RA potentially is present and demonstrates that a t-RA predominates over 9-cis-RA by at least 30- to 180-fold in different sp inal cord regions. (C) 2001 Wiley-Liss, Inc.