Abstract Patients with neurofibromatosis type 1 (NF1) have mutations in the NF1 tumor-suppressor gene that functions as a GTPase activating protein (GAP) for p21ras. Individuals with NF1 display a variety of skeletal defects including lytic bone lesions. However, the effect of loss of NF1 on osteoclasts or osteoblasts, the two principal cells involved in bone remodeling, is not understood. Osteoclasts are specialized myeloid cells that adhere to bone matrix, and secrete lytic enzymes that degrade bone. Given that we have previously identified haploinsufficient phenotypes in Nf1 +/− mast cells, we defined the role of neurofibromin in regulating various osteoclast functions in vitro and in vivo in Nf1 +/− mice. Strikingly, histologic sections from femurs of Nf1 +/− mice, revealed large numbers of multinucleated osteoclasts. This phenotype is reminiscent of the osteoclasts in patients with Paget’s disease and of osteoclasts isolated from SHIP deficient mice, which have elevated levels of PI-3K activity. Additionally, osteoclast progenitors from Nf1 +/− mice were hyperresponsive to limiting concentrations of M-CSF and RANKL and formed higher numbers of OCL progenitors compared to +/+ controls (P<0.05). M-CSF binding to its receptor (c-fms) causes a rapid increase in p21ras activity. Nf1 +/− osteoclasts had higher basal and M-CSF-stimulated p21ras-GTP levels compared to wild type cells. The Ras-PI3K- pathway has been implicated in controlling numerous osteoclast functions. An increase in basal and M-CSF stimulated Akt activation, a sensitive indicator of PI3-K activity, was observed in Nf1 +/− osteoclasts. Since a key initiating component of the resorptive process of osteoclasts is their ability to bind to αv B3 on the bone surface, we next examined whether haploinsufficiency of Nf1 altered adhesion by culturing osteoclasts on αv B3 coated plates. Nf1 +/− osteoclasts had a significant increase in adhesion to αv B3 compared to wildtype controls. Osteoclasts were then cultured onto bone slices and the number and size of the resorption pits were scored. Nf1 +/− osteoclasts have a higher frequency of forming pits as compared to wildtype controls and additionally the area of each pit was significantly greater. Further, a solid phase immunofixed-enzyme activity assay that measures bone resorption revealed that Nf1 +/− mice had a 2 fold increase in activity as compared to WT controls. Consistent with this activity, Nf1 +/− femurs required less stress to fracture in biomechanical testing. Further, utilizing a genetic intercross between Nf1+/− and Class1A PI3K deficient mice (p85α), we demonstrate that the increased number of osteoclast progenitors, as well as the increased hypersensitivity of Nf1 +/− osteoclast progenitors to M-CSF and RANKL were corrected to WT levels in progenitors that are mutant at both the Nf1 and the p85α loci. Additionally, the gain in function in the proliferation, migration, adhesion, and lytic activity of Nf1 +/− osteoclasts is mediated via the Ras-PI3K signaling axis. Collectively, these studies provide novel insights into the role of neurofibromin in regulating a lineage that may contribute to skeletal abnormalities in individuals with NF1.