Effects of extracellular Ca(2+) concentration on hair-bundle stiffness and gating-spring integrity in hair cells

When a hair cell is stimulated by positive deflection of its hair bundle, increased tension in gating springs opens transduction channels, permitting cations to enter stereocilia and depolarize the cell. Ca(2+) is thought to be required in mechanoelectrical transduction, for exposure of hair bundles to Ca(2+) chelators eliminates responsiveness by disrupting tip links, filamentous interstereociliary connections that probably are the gating springs. Ca(2+) also participates in adaptation to stimuli by controlling the activity of a molecular motor that sets gating-spring tension. Using a flexible glass fiber to measure hair-bundle stiffness, we investigated the effect of Ca(2+) concentration on stiffness before and after the disruption of gating springs. The stiffness of intact hair bundles depended nonmonotonically on the extracellular Ca(2+) concentration; the maximal stiffness of ≈1200 μN⋅m(−)(1) occurred when bundles were bathed in solutions containing 250 μM Ca(2+), approximately the concentration found in frog endolymph. For cells exposed to solutions with sufficient chelator capacity to reduce the Ca(2+) concentration below ≈100 nM, hair-bundle stiffness fell to ≈200 μN⋅m(−)(1) and no longer exhibited Ca(2+)-dependent changes. Because cells so treated lost mechanoelectrical transduction, we attribute the reduction in bundle stiffness to tip-link disruption. The results indicate that gating springs are not linearly elastic; instead, they stiffen with increased strain, which rises with adaptation-motor activity at the physiological extracellular Ca(2+) concentration.