Endogenous retinoids are potential regulators of vertebrate embryogenesis that have been implicated in early anterior-posterior patterning and limb-bud development. We have characterized the temporal and spatial distribution of 9-cis-retinoic acid in the Xenopus embryo and compared it to two other retinoids, afl-tmns-retinoic acid and alI-frns-retinoyl-3glucuronide. 9-cis-Retinoic acid is first detected after the midblastula transition and by the end of gastruation is localz primarily within the anterior and posterior dorsal regions ofthe embryo. Since 9-cis-retinoic acid is a 6-fold more potent dysmorphogen than trans-retinoic acid, we suggest that it is involved in the early specification of the Xenopus anterior-posterior aids.The organization of the vertebrate body plan is dependent upon communication between groups of cells in the early embryo. It has been proposed that all-trans-retinoic acid (trans-RA), a metabolite of retinol (vitamin A) and a member of a class of compounds known as retinoids, functions as one of the molecular signals in early vertebrate development (1-9). In support of this proposal, experiments have shown that trans-RA affects cellular differentiation (10) and that embryonic exposure to trans-RA causes severe malformations in a number ofvertebrate species (11-13). Furthermore, retinoid deficiency during embryonic development also produces malformations (14,15). The cellular effects of RA isomers appear to be mediated by two distinct classes of receptors within the steroid-thyroid nuclear receptor superfamily, the RA receptors (RARs) and the retinoid X receptors (RXRs), which presumably function by altering the transcription of specific target genes (16). In addition, cellular RA and retinol binding proteins (CRABPs and CRBPs) appear to modulate the localization, availability, and metabolism of these retinoids within the organism (17).Despite the ability of exogenous all-trans-RA to cause major alterations to the anterior-posterior (A-P) axis of mice (2, 3) and frogs (1, 4-6), evidence that endogenous retinoids are important modulators of early development has been difficult to obtain. Indirect evidence that retinoids are present in restricted regions of the embryo has been obtained from in vivo studies with transgenic mice (18-21) and in vitro studies with rat (22) and chicken (23) tissue explants. Chicken embryos also exhibit regional differences in the capacity to convert retinol to trans-RA (24), with the maximum activity found in Hensen's node. Since these studies have not directly identified the specific retinoids in the early embryo or measured their spatial distribution during early embryonic stages, we have addressed these questions using the amphibian embryo. The results from these studies indicate that a number of retinoids were present in the early amphibian embryo and that the levels of three of these retinoids vary both spatially and temporally in early development. Notably, 9-cis-RA was first detected after the midblastula transition and at the end of gastrulation ...

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It has been reported that fractionated doses of 13-cis-retinoic acid are disproportionately more embryotoxic in pregnant mice than is the same dose given in a single bolus. Here, we examined limited pharmacokinetic profiles of a single (100 mg/kg dose given to NMRI mice on day 11 of gestation) versus multiple (3 x 100 mg/kg, 4 h apart) doses in an effort to assess the relative contribution to teratogenicity made by the drug and/or its metabolites. The major plasma metabolite of 13-cis-retinoic acid in the mouse was 13-cis-retinoyl-beta-glucuronide, followed by the 4-oxo metabolites and all-trans-retinoic acid. Transfer to the mouse embryo was very efficient for all-trans-retinoic acid, whereas, it was tenfold less efficient for 13-cis-retinoic acid and 100-fold less efficient for 13-cis-retinoyl-beta-glucuronide. The isomer all-trans-retinoic acid was found in the placenta at concentrations two- to three-fold higher than in the plasma, suggesting placental accumulation as well as placental cis/trans isomerization. Since 13-cis-retinoyl-beta-glucuronide and 13-cis- and all-trans-retinoic acid were detected in the embryo after this multiple dosing schedule, any of the three or their combinations may have been involved in the induction of malformations, but all-trans-retinoic acid, a well-known potent teratogen detected at concentrations of between 590 and 80 ng/g for 10 critical hours during gestation, could have been the major component.

Recently, the temporal and spatial distribution patterns of the patterns of all-trans-retinol, didehydroretinol and all-transretinoid receptor ligands 9-cis-retinoic acid and all-trans-retinoic retinal did not coincide with that observed for 9-cis-retinoic acid were described in Xenopus embryos during early develacid but, in certain regards, were similar to the patterns delineated opment [Creech Kraft, Schuh, Juchau and Kimelman (1994) for all-trans-retinoic acid and all-trans-retinoyl fl-glucuronide.Proc. Natl. Acad. Sci. U.S.A., in the press]. The present study Evidence is presented that 9-cis-retinoic acid can be synthesized demonstrates the presence and distribution of their likely prefrom both all-trans-retinoic acid and all-trans-retinol in Xenopus cursors, all-trans-retinol, didehydroretinol, didehydroretinal and embryos, suggesting that the difference between the distributions all-trans-retinal, as well as the occurrence of 4-oxo metabolites, of 9-cis-retinoic acid and the other retinoids may be caused by in Xenopus embryos. The temporal and spatial distribution selective synthesis and/or protein binding of the 9-cis isomer.