To accommodate developing tooth germ in the alveolar bone, active bone resorption and the recruitment of numerous osteoclasts are essential. Recently, the signaling of receptor activator of nuclear factor-KappaB (RANK) and its ligand (RANKL) was reported to play a pivotal role in osteoclast formation and activation. The aim of this study was to examine the expression of RANKL and the contribution of RANK-RANKL signaling to the process of tooth germ and alveolar bone development. In situ hybridization showed RANKL was expressed in dental follicle cells and osteoblasts on the alveolar bone surface surrounding developing tooth germs. To elucidate the function of RANKL, mouse mandibular explants on embryonic day 14 were subjected to organ culture with osteoprotegerin (OPG), an inhibitor of RANK-RANKL signaling as a decoy receptor of RANKL. Many tooth germs were compressed with the surrounding bone tissue in the OPG-treated explants, whereas these abnormalities were not seen in untreated explants. The numbers of tartrate-resistant acid phosphatase (TRAP)-positive osteoclastic cells aligning on the alveolar bone surface were significantly decreased in OPG-treated explants compared with untreated explants. Moreover, TRAP-positive osteoclastic cells were not observed along the alveolar bone surfaces depressing tooth germs. These observations suggest that osteoclastogenesis in the alveolar bone, which is essential for the accommodation of normal tooth development, is mediated by RANK-RANKL signaling.