Effects of extracellular Ca2+ 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.sup.2+] is thought to be required in mechanoelectrical transduction, for exposure of hair bundles to [Ca.sup.2+] chelators eliminates responsiveness by disrupting tip links, filamentous interstereociliary connections that probably are the gating springs. [Ca.sup.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.sup.2+] concentration on stiffness before and after the disruption of gating springs. The stiffness of intact hair bundles depended nonmonotonically on the extracellular [Ca.sup.2+] concentration; the maximal stiffness of [approximately equal to] 1200 [Mu]N[center dot][m.sup.-1] occurred when bundles were bathed in solutions containing 250 [[micro]molar] [Ca.sup.2+], approximately the concentration found in frog endolymph. For cells exposed to solutions with sufficient chelator capacity to reduce the [Ca.sup.2+] concentration below [approximately equal to] 100 nM, hair-bundle stiffness fell to [approximately equal to] 200 [Mu]N[center dot][m.sup.-1] and no longer exhibited [Ca.sup.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.sup.2+] concentration.