Regulation of free Ca2+ concentration in hair-cell stereocilia

By affecting the activity of the adaptation motor, Ca2+ entering a hair bundle through mechanoelectrical transduction channels regulates the sensitivity of the bundle to stimulation. For adaptation to set the position of mechanosensitivity of the bundle accurately, the free Ca2+ concentration in stereocilia must be tightly controlled. To define the roles of Ca2+-regulatory mechanisms and thus the factors influencing adaptation motor activity, we used confocal microscopy to detect Ca2+ entry into and clearance from individual stereocilia of hair cells dialyzed with the Ca2+ indicator fluo-3. We also developed a model of stereociliary Ca2+ homeostasis that incorporates four regulatory mechanisms: Ca2+ clearance from the bundle by free diffusion in one dimension, Ca2+ extrusion by pumps, Ca2+ binding to fixed stereociliary buffers, and Ca2+ binding to mobile buffers. To test the success of the model, we compared the predicted profiles of fluo-3 fluorescence during the response to mechanical stimulation with the fluorescence patterns measured in individual stereocilia. The results indicate that all four of the Ca2+ regulatory mechanisms must be included in the model to account for the observed rate of clearance of the ion from the hair bundle. The best fit of the model suggests that a free Ca2+ concentration of a few micromolar is attained near the adaptation motor after transduction-channel opening. The free Ca2+ concentration substantially rises only in the upper portion of the stereocilium and quickly falls toward the resting level as adaptation proceeds.