Recent studies have suggested that myosin I beta mediates the adaptation of mechanoelectrical transduction in vestibular hair cells. An important prediction of this hypothesis is that myosin I beta should be found in the side insertional plaque, an osmiophilic hair bundle structure that anchors tip links and is thought to house the adaptation motor. To determine whether myosin I beta was situated properly to perform adaptation, we used immunofluorescence and immunoelectron microscopy with the monoclonal antibody mT2 to examine the distribution of myosin I beta in hair bundles of the bullfrog utricle. Although utricular hair cells differ in their rates and extent of adaptation [Baird RA (1994) Comparative transduction mechanisms of hair cells in the bullfrog utriculus. II. Sensitivity and response dynamics to hair bundle displacement. J Neurophysiol 71:685-705.], myosin I beta was present in all hair bundles, regardless of adaptation kinetics. Confirming that, nevertheless, it was positioned properly to mediate adaptation, myosin I beta was found at significantly higher levels in the side insertional plaque. Myosin I beta was also present at elevated levels at the second tip link anchor of a hair bundle, the tip insertional plaque, found at the tip of a stereocilium. These data support myosin I beta as the adaptation motor and are consistent with the suggestion that the motor serves to restore tension applied to transduction channels to an optimal level, albeit with different kinetics in different cell types.