PRELIMINARY EVIDENCE OF PHENYTOIN-INDUCED ALTERATIONS IN EMBRYONIC GENE-EXPRESSION IN A MOUSE MODEL

SWV mouse embryos collected on gestational days (GD) 9:12 and 10:00 fo llowing chronic in utero exposure to teratogenic concentrations of phe nytoin were utilized for in situ transcription studies of gene express ion. The substrate cDNA obtained from the frozen embryo sections was a mplified into radiolabelled antisense RNA (RT/aRNA) and used as a prob e to screen a panel of 20 cDNA clones representing genes that are impo rtant regulators of craniofacial and neural development. The magnitude of alteration in gene expression following phenytoin treatment was de termined densitometrically by changes in the hybridization intensity o f the aRNA probes to the cDNA clones immobilized to the slot blots. We found that both Wnt-1 and the calcium channel gene were developmental ly regulated, as their level of expression decreased significantly bet ween the two collection times. Phenytoin treatment produced a signific ant downregulation in the level of expression for 25% of the genes exa mined in the GD 9:12 embryos, including the growth factors TGF-beta an d NT3, the proto-oncogene Wnt-1, the nicotinic receptor, and the volta ge sensitive calcium channel gene. Additional changes in the coordinat e expression of several of the growth and transcription factors were o bserved at both gestational timepoints. The application of RT/aRNA tec hnology has extended our appreciation of the normal patterns of gene e xpression during craniofacial and neural development, and provided the first demonstration of multiple coordinate changes in transcription p atterns following teratogenic insult.