Topological and Developmental Expression Gradients of Kir2.1, an Inward Rectifier K⁺ Channel, in Spiral Ganglion and Cochlear Hair Cells of Mouse Inner Ear

Abstract

Objectives: Inwardly rectifying potassium current plays critical roles in setting resting membrane potential and thus modulating the excitability of many excitable cells including the hair cells in inner ears, which are excitable during early development. Up to 7 subfamilies have been identified as channels for this current. The present study investigated the developmental and spatial expression of one member, Kir2.1, in the sensorineural epithelia and spiral ganglion neurons of mouse cochleae starting from neonatal stage to the maturation of hearing function, in an attempt to verify its role in the postnatal cochlear development. Methods: The distribution of Kir2.1 protein and mRNA was observed using immunofluorescence staining and in situ hybridization on samples of surface preparation and cross sections of cochleae. The protein and mRNA were further measured with semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blotting to show the developmental change of this channel in the cochlea of mice. Results: The expression of Kir2.1 appears to have a clear temporal pattern: increasing from the first postnatal day (PD0) to PD12 and then quickly decreasing after that. The expression cannot be detected on PD20. Throughout this developmental pattern, the expression was stronger at the basal turn on PD0 and shifted upwards (longitudinal gradient), so that stronger signals were generally seen in the apical parts of the cochlea after PD0. A radial gradient of the Kir2.1 protein was also evident with a stronger and consistent signal usually occurring on row 3 outer hair cells. The spatial and developmental changes of Kir2.1 mRNA in in situ hybridization exhibited similar patterns as seen for the Kir2.1 protein on PD8 and PD12. Western blot data showed a slightly higher concentration of the protein in the apical half of the cochlea and a slight increase from PD4 to PD12. This result is consistent with the quantification of mRNA in RT-PCR. No significant expression of Kir2.1 was found in spiral ganglion neurons. Conclusions: Kir2.1 exhibits a clear and temporal expression in the hair cells of mouse cochleae which may be related to the functional maturation of the hair cells and the neurons.

The function and molecular identity of inward rectifier channels in vestibular hair cells of the mouse inner ear

“…Consistent with previous studies (Géléoc et al 2004;Rüsch et al 1998), we observed strong inward rectifier currents throughout early development in mouse utricle type II hair cells. We used the whole-cell, tight-seal technique in voltage-clamp mode to record fast inward rectifier currents, which were evoked by a protocol that included hyperpolarizing voltage steps from a holding potential of Ϫ64 mV to potentials between Ϫ124 and Ϫ54 mV in increments of 10 mV.…”

“…Small changes in the electrical properties of the basolateral membrane can modify the hair cell receptor potential and consequently, neurotransmitter release at the afferent synapse. These changes include the developmental acquisition of various ion channels required for proper modulation and transmission of the sensory signal, which continues until hair cells reach functional maturity, around postnatal day 8 (P8) (Rüsch et al 1998), as well as modulation by various efferent mechanisms. To extend an understanding of the functional development and the molecular components required for hair cell signaling, we focus here on the Kir2 family of potassium inward rectifier channels and their contributions to mammalian vestibular hair cell function.…”

“…The Kir2 ion channel family includes four subunits in the mouse and two additional subunits, Kir2.5 and -2.6, which are present in the carp and human genomes, respectively (Hassinen et al 2008;Ryan et al 2010). Most Kir2 channels form functional homomeric tetramers, whereas some assemble into heteromeric tetramers (Preisig-Müller et al 2002;Schram et al 2002).…”

“…Expression of Kir2 channels has been reported in mouse auditory hair cells (Ruan et al 2008) and avian vestibular hair cells (Correia et al 2004;Navaratnam et al 1995), but their physiological correlates and contributions have not been elucidated, particularly in mammals. In inner ear hair cells, the inward rectifier current, I K1 , contributes to sensory signaling by maintaining resting potential at more negative values, lowering input resistance (Brichta et al 2002;Géléoc et al 2004;Holt and Eatock 1995;Rüsch et al 1998) and enhancing receptor potentials by deactivation of their negative-feedback mechanism at depolarized membrane potentials (Goodman and Art 1996).…”

“…In inner ear hair cells, the inward rectifier current, I K1 , contributes to sensory signaling by maintaining resting potential at more negative values, lowering input resistance (Brichta et al 2002;Géléoc et al 2004;Holt and Eatock 1995;Rüsch et al 1998) and enhancing receptor potentials by deactivation of their negative-feedback mechanism at depolarized membrane potentials (Goodman and Art 1996). Previously, Géléoc et al (2004) showed a correlation between the acquisition of I K1 in mouse utricle in early development and more negative resting potentials.…”