Quantitative axial profiles of retinoic acid in the embryonic mouse spinal cord: 9‐Cis retinoic acid only detected after all‐trans‐retinoic acid levels are super‐elevated experimentally

Abstract

Studies using bioassays in nor-We find a bimodal distribution of all-trans retinoic acid (at-RA), the ligand for RARs, and relate this to the expression of several retinoid-synthesizing enzymes. However, we do not detect 9-cis-retinoic acid (9-cis-RA), the putative RXR ligand, in any region of the spinal cord 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 more definitive studies of retinoid function in the future. It sets definite limits on how much 9-cis-RA potentially is present and demonstrates that at-RA predominates over 9-cis-RA by at least 30-to 180-fold in different spinal cord regions.

Spatiotemporal retinoid-X receptor activation detected in live vertebrate embryos

“…CYP26 expression may explain the lack of gRXR activation in the brain and the inability of exogenous retinoic acid isomers to induce reporter gene expression anterior to the spinal cord, whereas the synthetic ligand LG268 is able to do so. Although endogenous concentrations of retinoids have been measured in early X. laevis embryos (49), other studies in frogs and mice have failed to show significant levels of 9-cis RA in the developing embryo (50,51). These findings raise questions about a physiological role for endogenous 9-cis RA as a physiological RXR ligand.…”

Cyanobacteria blooms produce teratogenic retinoic acids

“…Of the 39 cyanobacteria and algae species, 9cRA/9c-4-oxo-RA were detected in 14 species with intracellular concentrations from 1.4 to 5.6 × 10 2 ng/g DW ( Table 2). Although the physiological function for the normal existence and role of 9cRA is of great interest in vertebrates (25), 9cRA was only identified in Xenopus embryos (38), in humans, or in mice after excessive amounts of retinol or atRA (39,40). A recent study determined 9cRA to be a naturally occurring RA with concentrations of 3-12 ng/g (10-40 pmol/g) in mouse pancreas, which confirms that 9cRA contributes to the biological activity of vitamin A in vivo (41).…”

“…During the culture time, the culture was shaken three times every day. The cyanobacteria and medium were taken for analysis of RAs and 4-oxo-RAs on days 10,30,35,40,45,50,55,60,70, and 90. The cell counting was carried out (n = 6), and the number of cells (cells per liter) was calculated at each sampling time.…”

Methods for detecting and identifying retinoids in tissue

“…SPE coupled on-line via an automated diversion valve was originally described in 1981 by Roth et al for extraction of drugs in bio-fluids. Variants of this technique were later adapted by Kraft et al (1988), researchers at Hoffman-La Roche in Basel, and several others for the analysis of retinoids in biological samples (Wyss andBucheli, 1988b, 1992;Gundersen et al, 1997;Wyss et al, 1998;Sakhi et al, 1998;Gundersen and Blomhoff, 1999;Ulven et al, 2000Ulven et al, , 2001Hartman et al, 2001;Otto et al, 2003;Ruhl and Schweigert, 2003;Moren et al, 2005;Ruhl et al, 2005;Berggren et al, 2005). A drawback of this technique is a high degree of complexity requiring a notable level of technical expertise by the user as well as a somewhat higher initial cost compared to regular HPLC systems.…”

“…Reproduced with permission from Gundersen and Blomhoff (2001). for RA, RAL, and ROH of 23 pg, 1.0 ng, and 0.5 ng, respectively, whereas APCI gave mass limits of detection of 0.670 pmol atROH and 0.720 pmol atretinyl palmitate on-column. Ulven et al (2001) used RP-LC-ESI-MS (single stage) in combination with diode array detection for identification of ROH and RA in dissected segments of the embryonic mouse spinal cord (Fig. 8).…”