Isotretinoin (ITR), a teratogen in many species, is associated with increased oxidative stress. Metallothionein (MT) is an important tissue antioxidant whose concentrations are induced by zinc. To study the role of supplemental Zn as an inducer of embryonic MT, we injected pregnant CD-1 mice subcutaneously with saline vehicle, or 20 or 40 mg/kg Zn on gestational day (GD) 6.5. After 48 h, embryonic MT concentrations increased in a dose-related manner (r = 0.64, P < 0.05) with Zn treatment. The possible protective role of Zn pretreatment against ITR teratogenicity was investigated in vivo and in vitro. CD-1 mice were pretreated with saline or Zn (20 and 40 mg/kg) on GD 8.5 and 9.5. ITR was administered to both groups of mice via three intragastric intubations of 100 mg ITR/kg at 4 h intervals on GD 10.5. On GD 18.5, Zn pre-treated mice demonstrated decreased ITR-mediated growth retardation, cleft palates and postpartum mortality. A reduction in embryonic MT concentrations was observed in mice exposed to ITR. Mouse embryos cultured on GD 8.5 with an addition of 15 micromol/L Zn for 48 h had a sixfold greater MT concentration (688 microg/g protein) than controls. The Zn pretreatment of cultured embryos prevented malformations and lessened growth retardation caused by 24 h exposure to 17 micromol/L ITR. These results suggest that Zn-mediated induction of MT in mouse embryos could protect against ITR teratogenicity.