Phosphatidylinositol 3'(OH)-kinase (PI 3-kinase) is activated on stimulation of macrophages with colony-stimulating factor 1 (CSF-1). We studied its potential role in the internalization and degradation of CSF-1 and its receptor in two primary populations of murine macrophages, namely bone marrow-derived macrophages (BMM) and resident peritoneal macrophages (RPM). Even though CSF-1 induced PI 3-kinase activity in both BMM and RPM, wortmannin, a potent inhibitor of PI 3-kinase activity, at concentrations that inhibited PI 3-kinase activity by 90% in these cells, had little or no effect on receptor internalization and degradation in either BMM or RPM or on CSF-1 degradation by BMM. Strong (more than 90%) inhibition was, however, observed for CSF-1 degradation by RPM. These findings suggest that both wortmannin-sensitive and wortmannin-insensitive pathways of ligand degradation exist in macrophages and that, although CSF-1 and CSF-1 receptor share the same endocytic pathway initially, they might be targeted to different compartments at later stages of degradation.