Valproic acid-induced somite teratogenesis in the chick embryo: relationship with Pax-1 gene expression

The repeated pattern of the axial skeleton results from the segmentation and re-segmentation of the mesodermally derived somites. During these early events of somite development, the vertebrate embryonic axial skeleton is most susceptible to the teratogenic effects of a variety of pharmaceutical and environmental agents. One example is the anticonvulsant drug valproic acid (VPA), which has been shown to cause craniofacial and minor and major skeletal defects in human and animal embryos. We hypothesize that a candidate set of molecular targets of teratogens are the Pax family of pattern-forming genes, specifically Pax-1, which has been previously demonstrated to be an important regulator of axial skeletal patterning at the somite level. In this study, early developmental stage chick embryos were treated with VPA and dose-dependent malformations in somite development were observed. Two classes of anomalies were evident: class I included discrete sites of somitic fusions or mis-segmentation, and Class II included large areas of disorganized somite patterning. Northern blot analysis revealed a decreased level of Pax-1 expression in VPA-treated embryos. Whole mount in situ hybridization analysis showed that somite anomalies correlate spatially with regions of decreased Pax-1 expression. Finally, comparison of the VPA-induced somitic anomalies with those caused by gene-specific perturbation of Pax-1 gene expression through the use of an antisense oligonucleotide revealed significant similarities. Taken together, these results support the hypothesis that Pax-1 is a molecular target in VPA axial skeletal teratogenicity.