P-Glycoprotein (P-gp), the multidrug resistance (MDR) gene product, has previously been shown to be functional and overexpressed in adult rat hepatocytes during primary culture. In the present study, we examined P-gp expression in human, mouse and hamster hepatocytes cultured in conditions similar to those used for rat liver cells. Northern blotting and doxorubicin P-gp-mediated efflux analyses revealed that liver parenchymal cells from these three species exhibited only limited increased MDR mRNA levels with no intracellular decreased drug retention, thus suggesting that the marked functional P-gp induction observed in rat hepatocytes could reflect a species-specific response of these cells to an unfamiliar environment. An 8 h exposure to cycloheximide, which strongly induced MDR mRNAs in rat liver cells, also increased, although to a far lesser extent, MDR mRNA levels in human, mouse and hamster hepatocytes, indicating that P-gp expression in normal liver parenchymal cells could be at least partly negatively regulated by a labile protein factor.