Hair Bundle Stiffness in the Turtle Utricle: Structural and Regional Variations

Vestibular hair cells are mechanotransducing sensory receptors in the vertebrate inner ear that detect movement and orientation of the head with respect to gravity. The morphologies of their ciliary bundles vary greatly for different species, endorgans, and within the same endorgan. Bundle morphology in the turtle utricle, like other species, demonstrates highly organized regional variations. These structural differences in bundles impact their mechanical behavior and the process of mechanotransduction. To further understanding of the mechanical behavior of hair bundles, this work experimentally measured the stiffness of bundles with differing morphology, the stiffness contribution of interciliary links and the mechanical properties of the kinocilium in the turtle utricle.

The stiffness of hair bundles of varying structure and location along a medial to lateral transect of the utricle was examined. Bundle stiffness was greatest in the striola and demonstrated a systematic decline with location from the line of polarity reversal. The average stiffness of bundles in the striola and extrastriola were 82 ± 46 (n=48) and 9 ± 5 (n=25) µN/m, respectively. The stiff and weak bundles demonstrated characteristic morphologies. The stiffest bundles have short kinocilium, tall stereocilia, and ratios of kinocilium to tallest stereocilia height (KS) close to 1. In contrast, the compliant bundles have tall kinocilium, short stereocilia, and KS ratios ranging from 1.6 – 8. The stiffer bundles also tend to have longer array lengths and steeper slopes. Measurements of bundle stiffness in the turtle utricle are lower than those previously reported which may be attributed to morphological differences between species.

The stiffness contributions of the interciliary links were also examined through their selective removal with exposure to the Ca²⁺ chelator BAPTA and the protease subtilisin. BAPTA treatment reportedly breaks tip, kinocilial and ankle links while subtilisin breaks the shaft and ankle links. Following BAPTA and subtilisin treatments, bundle stiffness reduced by 65 ± 10% and 63 ± 11%, respectively.

The mechanical properties of the kinocilium were measured with novel techniques. Flexural rigidity (EI) was measure while the kinocilium was fixed at the height of the tallest stereocilia using a glass supporting probe. Through both force deflection and a high speed video technique, measured values of EI ranged from 1460 – 6150 pN·µm2. The rotational stiffness of the kinocilium about its apical insertion was also measured. Bundles were treated with BAPTA to break the kinocilial links and separate the kinocilium from neighboring stereocilia. Using a force deflection technique, the rotational stiffness of the kinocilium was measured as 120 ± 17 pN·µm/rad.