Although it is known that mammalian otoconia consist of calcium bicarbonate and organic materials, none of the protein components have been identified in mammals at the molecular level, and the mechanism of morphogenesis and calcification of the otoconia remains unclear. In the present study, we demonstrated the presence of osteopontin (OPN) in rat otoconia by using immunohistochemistry, and detected OPNmRNA in the sensory hair cells using a non-radioisotopic in situ hybridization technique. These results indicate that OPN is one of the protein components in rat otoconia and suggest that sensory hair cells are involved in the production of otoconia.