Two populations of primary osteoblasts and the cloned murine osteoblast cell line MC3T3 were activated with osteotropic agents and the conditioned media tested for the presence of macrophage colony stimulating factor (M-CSF). Differences in ability of the three populations to secrete the cytokine as well as in the efficacy of the activating agents used to induce it were observed. The ability of these agents to modulate M-CSF mRNA levels was also examined. The data indicate that osteoblasts do not respond uniformly and the reasons for this are discussed. Osteoblasts may exert major regulatory influences on hematopoiesis via CSF secretion as well as function in their more traditional role as the cells responsible for new bone formation.