Disruption of the mdr2 gene in mice leads to a complete absence of phospholipid from bile (Smit, J. J. M., et al. 1993. Cell. 75:451-462). We have investigated the control of both mdr2 P-glycoprotein (Pgp) expression and bile salt secretion on biliary lipid secretion in the mouse. Lipid secretion was monitored at various bile salt output rates in wild-type mice (+/+), heterozygotes (+/-), and homozygotes (-/-) for mdr2 gene disruption. In (-/-) mice, phospholipid secretion was negligible at all bile salt output rates. In (+/-) mice, a curvilinear relation between bile salt and phospholipid secretion was observed similar to that in (+/+) mice; however, at all bile salt secretion rates phospholipid secretion was reduced compared to (+/+) mice, indicating that mdr2 Pgp exerts a strong control over secretion. Infusion of increasing amounts of taurocholate up to maximal secretory rate led to a decline in the phospholipid and cholesterol secretion in both (+/+) and (+/-) mice in accordance to what has been observed in other species. In contrast, in (-/-) mice cholesterol secretion increased under these conditions while phospholipid output remained extremely low. The increased cholesterol secretion may represent extraction of cholesterol from the canalicular plasma membrane by taurocholate micelles as opposed to the concomitant secretion of both phospholipid and cholesterol in the presence of a functional mdr2 Pgp. Increased bile flow in (-/-) mice could be attributed completely to an increase in the bile salt-independent fraction and may therefore be caused by the bile duct proliferation in these mice.