The effect of Zn-induced metallothionein (MT) on the toxicity, uptake, and subcellular distribution of cadmium (Cd) was examined in rat primary hepatocyte cultures and compared to results obtained earlier in this laboratory from intact animals. Hepatocytes were isolated and grown in monolayer culture for 22 h and subsequently treated with ZnCl2 (100 microM) for 24 h, which increased MT concentration about 15-fold. After Zn pretreatment, hepatocytes were exposed to Cd for 24 h. Cytotoxicity was assessed by enzyme leakage, intracellular potassium loss, and cellular glutathione content. The toxicity of Cd was much less in Zn-pretreated cells than in control cells, similar to that previously demonstrated in the intact animal. Zn pretreatment had no appreciable effect on the hepatocellular uptake of 109Cd, but markedly altered its subcellular distribution, with more Cd accumulating in the cytosol and less in the nuclear, mitochondrial, and microsomal fractions. In the cytosol of Zn-pretreated cells, Cd was associated mainly with MT; in contrast, cytosolic Cd in control cells was mainly associated with non-MT macromolecules. Zn-induced changes in the subcellular distribution of Cd in vitro are identical to those observed in vivo in Zn-pretreated rats challenged with Cd. In summary, Zn pretreatment of rat primary hepatocyte cultures protects cells against Cd toxicity. Protection seems to be due to MT-promoted sequestration of Cd and reduction of the amount of Cd associated with critical organelles and proteins. These observations are similar to those noted in the whole animal. These results indicate that cultured hepatocytes are an ideal model for examining MT-induced tolerance to Cd hepatotoxicity.