Lactoferrin induces osteoblast proliferation and survival in vitro and is anabolic to bone in vivo. The molecular mechanisms by which lactoferrin exerts these biological actions are not known, but lactoferrin is known to bind to two members of the low density lipoprotein receptor family, low density lipoprotein-receptor-related proteins 1 (LRP1) and 2 (LRP2). We have examined the role(s) of these receptors in the actions of lactoferrin on osteoblasts. We show that lactoferrin binds to cultured osteoblastic cells, and that LRP1 and LRP2 are expressed in several osteoblastic cell types. In primary rat osteoblastic cells, the LRP1/2 inhibitor receptor associated protein (RAP) blocks endocytosis of lactoferrin and abrogates lactoferrin-induced p42/44 MAP kinase signaling and mitogenesis. Lactoferrin-induced mitogenesis is also inhibited by an antibody to LRP1. Lactoferrin also induces RAP-sensitive activation of p42/44 MAP kinase signaling and proliferation in osteoblastic human SaOS-2 cells, which express LRP1 but not LRP2. The mitogenic response of LRP1-null fibroblastic cells to lactoferrin is substantially reduced compared with that of cells expressing wild-type LRP1. The endocytic and signaling functions of LRP1 are independent of each other, since lactoferrin can activate mitogenic signaling in conditions in which endocytosis is inhibited. Taken together, these results (a) suggest that mitogenic signaling through LRP1 to p42/44 MAP kinases contributes to the anabolic skeletal actions of lactoferrin, (b) demonstrate growth-promoting actions of a third LRP family member in osteoblasts, and (c) provide further evidence that LRP1 functions as a signaling receptor in addition to its recognized role in ligand endocytosis.