Previous studies with the isolated perfused rat lung showed that both clathrin- and actin-mediated pathways are responsible for endocytosis of dipalmitoylphosphatidylcholine (DPPC)-labeled liposomes by granular pneumocytes in the intact lung. Using surfactant protein-A (SP-A) gene-targeted mice we examined the uptake of [³H]-DPPC liposomes by isolated mouse lungs under basal and secretagogue-stimulated conditions. Unilamellar liposomes composed of [³H]-DPPC: PC: cholesterol: PG (10:5:3:2 mol fraction) were instilled into the trachea of anesthetized mice and the lungs were perfused (2h). Uptake was calculated as % of instilled dpm in the post- lavaged lung. Amantadine, an inhibitor of clathrin and, thus, receptor-mediated endocytosis via clathrin-coated pits, decreased basal [³H]-DPPC uptake by 70% in SP-A+/+ but only by 20% in SP-A -/- lung, data compatible with an SP-A/receptor-regulated lipid clearance pathway in the SP-A+/+ mice. The non-clathrin, actin dependent process was low in the SP-A +/+ lung but accounted for 55% of liposome endocytosis in the SP-A -/- mouse. With secretagogue (8-Br-cAMP) treatment, both clathrin- and actin-dependent lipid clearance were elevated in the SP-A +/+ lungs while neither pathway responded in the SP-A -/- lungs. Binding of iodinated SP-A to type II cells isolated from both genotypes of mice was similar indicating a normal SP-A receptor status in the SP-A -/- lung. Inclusion of SP-A with instilled liposomes served to "rescue" the SP-A -/- lungs by re-establishing secretagogue-dependent enhancement of liposome uptake. These data are compatible with a major role for receptor-mediated endocytosis of DPPC by granular pneumocytes, a process critically dependent on SP-A.