Initiation of phenytoin teratogenesis: pharmacologically induced embryonic bradycardia and arrhythmia resulting in hypoxia and possible free radical damage at reoxygenation.

The aim of this study was to investigate if phenytoin has the capacity to induce embryonic hypoxia mediated via adverse effects on the embryonic heart. Mouse embryos of different strains (CD-1, C57B1/6J and A/J) as well as Sprague Dawley (SD) rat embryos were cultured in vitro (in 75-80% rat serum) by the whole embryo technique. Effects on the heart were examined on gestational day 10 for mouse embryos and days 11 and 13 for rat embryos. Phenytoin was dissolved in water to give concentrations of 50-800 microM. In the mouse embryo studies, phenytoin caused a concentration-dependent decrease in embryonic heart rate in all three strains, with a slight decrease at 100 microM (2-7%) and a more pronounced effect at 200 microM (approximately 20%). Temporary or permanent cardiac arrest occurred in 86% of the CD-1 embryos at 500 microM, in 67% of the C57B1/6JM at 400 microM, and in all A/J embryos at 300 microM. Arrhythmias was observed in 8% in CD-1 embryos at 200 microM, in 18% at 150 microM in C57B1/6J embryos, and in 67% of the A/J embryos at 100 microM (lowest tested concentrations where arrhythmias occurred). In rat embryos, a concentration-dependent decrease in heart rate was observed on both days 11 and 13 at similar concentrations as in the mouse embryo studies. In a separate experiment, the effects on the heart rate of free phenytoin (not serum protein bound) were examined in rat embryos cultured in serum-free medium. Already at 12 microM a significant decrease in heart rate was observed. Altogether, the results support the hypothesis that phenytoin teratogenicity is initiated by pharmacologically induced embryonic hypoxia. A genetic susceptibility to the adverse effects of phenytoin on the embryonic heart may be of importance to explain strain and species differences in phenytoin teratogenicity.