The expression of fimbrin in the developing rat cochlea was analyzed using an immunohistochemical technique with fimbrin antibody. The cochlea displayed temporal and lateral-longitudinal gradients for fimbrin expression during development. Fimbrin immunoreactivity first appeared in the inner hair cell stereocilia of the basal turn on the first gestational day studied (day 18). At birth, both inner (IHC) and outer hair cell (OHC) stereocilia of the basal turn showed positive labeling with fimbrin antibody. The progression of appearance was always from IHCs to OHCs and fimbrin immunostaining appeared in the apical hair cells by postnatal day 6. Immunostaining was restricted to stereocilia and the cuticular plate, and no immunoreactivity was observed in neighboring structures of the epithelium. Double labeling using both fimbrin antibody and phalloidin binding revealed similar chronological expression from the earliest stage studied. Increasing fimbrin immunoreactivity was observed in hair cells until late postnatal and adult stages. This study suggests that fimbrin is expressed with F-actin during development and fimbrin together with actin may constitute the two basic molecules that participate in stereocilia formation. We speculate that fimbrin may help maintain the parallel growth of actin filaments within the stereocilia. These data additionally support previous findings that hair cell maturation occurs from the base to the apex and from IHCs to OHCs.