Gene expression domains as markers in developmental toxicity studies using mammalian embryo culture

We are examining the hypothesis that expression domains of developmental control genes may be informative markers in mammalian embryo culture studies of developmental toxicity. Expression domains might be altered directly by chemical exposure, or might reflect developmental abnormality prior to any overt morphological defect. Whole-mount in situ hybridization using digoxygenin-labeled RNA probes was used to monitor the regions of expression of Hoxb-4, Pax-3 and Emx-2. These genes were selected because of their different restrictions within the developing CNS; Hoxb-4 for its anterior margin in the hindbrain, Pax-3 for its dorso-ventral pattern in the spinal cord, and Emx-2 for being restricted to a portion of the forebrain. Valproic acid was used as a prototype developmental toxicant because of its known actions on neural tube closure and on segmentation. For patterns of expression, we made three comparisons, between: rat in vivo developed embryos and published descriptions for mouse; rat cultured and in vivo; control and valproate exposed. For these genes, there were no differences between domains of expression in rat and mouse, nor between rat cultured and in vivo embryos. In valproate-exposed embryos, some domains were spatially abnormal, for example Pax-3 in the neural crest, but this was coincident with structural defects induced by the treatment. There was no indication, for these three genes, and this teratogen, that treatment caused any shifts in boundaries of expression, nor induced any ectopic domains, even though exposures induced overt malformation.