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Myosin-VIIA (MYO7A) is an unconventional myosin responsible for syndromic (Usher 1B) or nonsyndromic forms of deafness in humans when mutated. In the cochlea, MYO7A is expressed in hair cells, where it is believed to act as the motor protein tensioning the mechanoelectrical transducer (MET) channels, thus setting their resting open probability (Po). However, direct evidence for this unique role for an unconventional myosin in mature hair cells is lacking. Here, we show that MYO7A has a distinct role in hair cells, being crucial for the structural integrity of hair bundles. Postnatal deletion of Myo7a leads to 87 to 96% reduction in MYO7A from hair cells by postnatal day 20 (P20), without affecting hearing function. During the following week, mice showed progressive decline in both hearing function and MET current amplitude in hair cells without affecting the resting Po and calcium sensitivity of the MET channel. Hair-bundle stiffness was normal at P20 but halved at P30, despite it having a normal staircase morphology and tip links. The reduction of MYO7A in the stereocilia (>87%) increased their vulnerability to sound-induced damage, with significantly more hearing loss and hair bundle deterioration than in control mice. RNA-sequencing identified a downregulation of several stereociliary genes in the Myo7a-deficient cochlea, indicating the presence of indirect compensatory mechanisms. This study reveals that mature hair cells seem to use a MYO7A-independent mechanism to maintain the resting Po of the MET channels. Instead, MYO7A is essential for maintaining the structural and functional integrity of the hair bundles. [ABSTRACT FROM AUTHOR]

Spiral ganglion neurons (SGNs) are primary sensory afferent neurons that relay acoustic information from the cochlear inner hair cells (IHCs) to the brainstem. The response properties of different SGNs diverge to represent a wide range of sound intensities in an action‐potential code. This biophysical heterogeneity is established during pre‐hearing stages of development, a time when IHCs fire spontaneous Ca2+ action potentials that drive glutamate release from their ribbon synapses onto the SGN terminals. The role of spontaneous IHC activity in the refinement of SGN characteristics is still largely unknown. Using pre‐hearing otoferlin knockout mice (Otof−/−), in which Ca2+‐dependent exocytosis in IHCs is abolished, we found that developing SGNs fail to upregulate low‐voltage‐activated K+‐channels and hyperpolarisation‐activated cyclic‐nucleotide‐gated channels. This delayed maturation resulted in hyperexcitable SGNs with immature firing characteristics. We have also shown that SGNs that synapse with the pillar side of the IHCs selectively express a resurgent K+ current, highlighting a novel biophysical marker for these neurons. RNA‐sequencing showed that several K+ channels are downregulated in Otof−/− mice, further supporting the electrophysiological recordings. Our data demonstrate that spontaneous Ca2+‐dependent activity in pre‐hearing IHCs regulates some of the key biophysical and molecular features of the developing SGNs. Key points: Ca2+‐dependent exocytosis in inner hair cells (IHCs) is otoferlin‐dependent as early as postnatal day 1.A lack of otoferlin in IHCs affects potassium channel expression in SGNs.The absence of otoferlin is associated with SGN hyperexcitability.We propose that type I spiral ganglion neuron functional maturation depends on IHC exocytosis. [ABSTRACT FROM AUTHOR]

Sensory‐independent Ca2+ spiking regulates the development of mammalian sensory systems. In the immature cochlea, inner hair cells (IHCs) fire spontaneous Ca2+ action potentials (APs) that are generated either intrinsically or by intercellular Ca2+ waves in the nonsensory cells. The extent to which either or both of these Ca2+ signalling mechansims are required for IHC maturation is unknown. We find that intrinsic Ca2+ APs in IHCs, but not those elicited by Ca2+ waves, regulate the maturation and maintenance of the stereociliary hair bundles. Using a mouse model in which the potassium channel Kir2.1 is reversibly overexpressed in IHCs (Kir2.1‐OE), we find that IHC membrane hyperpolarization prevents IHCs from generating intrinsic Ca2+ APs but not APs induced by Ca2+ waves. Absence of intrinsic Ca2+ APs leads to the loss of mechanoelectrical transduction in IHCs prior to hearing onset due to progressive loss or fusion of stereocilia. RNA‐sequencing data show that pathways involved in morphogenesis, actin filament‐based processes, and Rho‐GTPase signaling are upregulated in Kir2.1‐OE mice. By manipulating in vivo expression of Kir2.1 channels, we identify a "critical time period" during which intrinsic Ca2+ APs in IHCs regulate hair‐bundle function. Synopsis: Immature inner hair cells (IHCs) generate spontaneous Ca2+ action potentials (APs) that are triggered either intrinsically or by intercellular Ca2+ waves in the nonsensory cells. This study shows that the absence of intrinsic APs blocks the maturation of the mechanoelectrical transducer apparatus. Overexpression of Kir2.1 potassium channels in IHCs prevents intrinsic APs but not APs induced by Ca2+ waves.Absence of intrinsic APs in IHCs causes them to lose mechanoelectrical transduction before the age of hearing onset.Intrinsic APs are required over a defined time window during prehearing development to regulate the maturation and maintenance of stereociliary hair bundles in IHCs. [ABSTRACT FROM AUTHOR]

Deafness is a condition with a high prevalence worldwide, produced primarily by the loss of the sensory hair cells and their associated spiral ganglion neurons (SGNs). Of all the forms [...]

Signal transmission by sensory auditory and vestibular hair cells relies upon Ca2+-dependent exocytosis of glutamate. The Ca2+ current in mammalian inner ear hair cells is predominantly carried through Ca V 1.3 voltage-gated Ca2+ channels. Despite this, Ca V 1.3 deficient mice (Ca V 1.3–/–) are deaf but do not show any obvious vestibular phenotype. Here, we compared the Ca2+ current (I Ca ) in auditory and vestibular hair cells from wild-type and Ca V 1.3–/– mice, to assess whether differences in the size of the residual I Ca could explain, at least in part, the two phenotypes. Using 5 mM extracellular Ca2+ and near-body temperature conditions, we investigated the cochlear primary sensory receptors inner hair cells (IHCs) and both type I and type II hair cells of the semicircular canals. We found that the residual I Ca in both auditory and vestibular hair cells from Ca V 1.3–/– mice was less than 20% (12–19%, depending on the hair cell type and age investigated) compared to controls, indicating a comparable expression of Ca V 1.3 Ca2+ channels in both sensory organs. We also showed that, different from IHCs, type I and type II hair cells from Ca V 1.3–/– mice were able to acquire the adult-like K+ current profile in their basolateral membrane. Intercellular K+ accumulation was still present in Ca V 1.3–/– mice during I K,L activation, suggesting that the K+-based, non-exocytotic, afferent transmission is still functional in these mice. This non-vesicular mechanism might contribute to the apparent normal vestibular functions in Ca V 1.3–/– mice. [ABSTRACT FROM AUTHOR]

Key points: The aim was to determine whether detachment of the tectorial membrane (TM) from the organ of Corti in Tecta/Tectb−/− mice affects the biophysical properties of cochlear outer hair cells (OHCs).Tecta/Tectb−/− mice have highly elevated hearing thresholds, but OHCs mature normally.Mechanoelectrical transducer (MET) channel resting open probability (Po) in mature OHC is ∼50% in endolymphatic [Ca2+], resulting in a large standing depolarizing MET current that would allow OHCs to act optimally as electromotile cochlear amplifiers.MET channel resting Poin vivo is also high in Tecta/Tectb−/− mice, indicating that the TM is unlikely to statically bias the hair bundles of OHCs.Distortion product otoacoustic emissions (DPOAEs), a readout of active, MET‐dependent, non‐linear cochlear amplification in OHCs, fail to exhibit long‐lasting adaptation to repetitive stimulation in Tecta/Tectb−/− mice.We conclude that during prolonged, sound‐induced stimulation of the cochlea the TM may determine the extracellular Ca2+ concentration near the OHC's MET channels. The tectorial membrane (TM) is an acellular structure of the cochlea that is attached to the stereociliary bundles of the outer hair cells (OHCs), electromotile cells that amplify motion of the cochlear partition and sharpen its frequency selectivity. Although the TM is essential for hearing, its role is still not fully understood. In Tecta/Tectb−/− double knockout mice, in which the TM is not coupled to the OHC stereocilia, hearing sensitivity is considerably reduced compared with that of wild‐type animals. In vivo, the OHC receptor potentials, assessed using cochlear microphonics, are symmetrical in both wild‐type and Tecta/Tectb−/− mice, indicating that the TM does not bias the hair bundle resting position. The functional maturation of hair cells is also unaffected in Tecta/Tectb−/− mice, and the resting open probability of the mechanoelectrical transducer (MET) channel reaches values of ∼50% when the hair bundles of mature OHCs are bathed in an endolymphatic‐like Ca2+ concentration (40 μM) in vitro. The resultant large MET current depolarizes OHCs to near –40 mV, a value that would allow optimal activation of the motor protein prestin and normal cochlear amplification. Although the set point of the OHC receptor potential transfer function in vivo may therefore be determined primarily by endolymphatic Ca2+ concentration, repetitive acoustic stimulation fails to produce adaptation of MET‐dependent otoacoustic emissions in vivo in the Tecta/Tectb−/− mice. Therefore, the TM is likely to contribute to the regulation of Ca2+ levels around the stereocilia, and thus adaptation of the OHC MET channel during prolonged sound stimulation. Key points: The aim was to determine whether detachment of the tectorial membrane (TM) from the organ of Corti in Tecta/Tectb−/− mice affects the biophysical properties of cochlear outer hair cells (OHCs).Tecta/Tectb−/− mice have highly elevated hearing thresholds, but OHCs mature normally.Mechanoelectrical transducer (MET) channel resting open probability (Po) in mature OHC is ∼50% in endolymphatic [Ca2+], resulting in a large standing depolarizing MET current that would allow OHCs to act optimally as electromotile cochlear amplifiers.MET channel resting Poin vivo is also high in Tecta/Tectb−/− mice, indicating that the TM is unlikely to statically bias the hair bundles of OHCs.Distortion product otoacoustic emissions (DPOAEs), a readout of active, MET‐dependent, non‐linear cochlear amplification in OHCs, fail to exhibit long‐lasting adaptation to repetitive stimulation in Tecta/Tectb−/− mice.We conclude that during prolonged, sound‐induced stimulation of the cochlea the TM may determine the extracellular Ca2+ concentration near the OHC's MET channels. [ABSTRACT FROM AUTHOR]