SUMMARY Sensory transduction in auditory and vestibular hair cells requires expression of transmembrane channel-like (Tmc) 1 and 2 genes, but the function of these genes is unknown. To investigate the hypothesis that TMC1 and TMC2 proteins are components of the mechanosensitive ion channels that convert mechanical information into electrical signals, we recorded whole-cell and single-channel currents from mouse hair cells that expressed Tmc1, Tmc2 or mutant Tmc1. Cells that expressed mutant Tmc1 had reduced calcium permeability and reduced single-channel currents, while Tmc2 cells had high calcium permeability and large single-channel currents. Cells that expressed both Tmc1 and Tmc2 had a broad range of single-channel currents, suggesting multiple heteromeric assemblies of TMC subunits. The data demonstrate TMC1 and TMC2 are components of hair cell transduction channels and contribute to permeation properties. Gradients in TMC channel composition may also contribute to variation in sensory transduction along the tonotopic axis of the mammalian cochlea.

HCN1-4 subunits form Na+/K+ permeable ion channels that are activated by hyperpolarization and carry the current known as Ih. Ih has been characterized in vestibular hair cells of the inner ear, but its molecular correlates and functional contributions have not been elucidated. We examined Hcn mRNA expression and immunolocalization of HCN protein in the mouse utricle, a mechanosensitive organ that contributes to the sense of balance. We found that HCN1 is the most highly expressed subunit, localized to the basolateral membranes of type I and type II hair cells. We characterized Ih using the whole-cell, voltage-clamp technique and found the current expressed in ~90% of the cells with a mean maximum conductance of 4.4 nS. Ih was inhibited by ZD7288, Cilobradine and by adenoviral expression of a dominant-negative form of HCN2. To determine which HCN subunits carried Ih we examined hair cells from mice deficient in Hcn1, 2, or both. Ih was completely abolished in hair cells of Hcn1−/− mice and Hcn1,2−/− mice but was similar to wild-type in Hcn2−/− mice. To examine the functional contributions of Ih, we recorded hair cell membrane responses to small hyperpolarizing current steps and found that activation of Ih evoked a 5-10 mV sag depolarization and a subsequent 15-20 mV rebound upon termination. The sag and rebound were nearly abolished in Hcn1-deficient hair cells. We also found that Hcn1-deficient mice had deficits in vestibular evoked potentials and balance assays. We conclude that HCN1 contributes to vestibular hair cell function and the sense of balance.

Ex vivo analyses indicate that spontaneous activity in vestibular neurons depends on resting hair cell mechanotransduction and HCN channels in calyx terminals.