Outer hair cells (OHCs) are highly specialized sensory cells conferring the fine‐tuning and high sensitivity of the mammalian cochlea to acoustic stimuli. Here, by genetically manipulating spontaneous Ca2+ signalling in mice in vivo, through a period of early postnatal development, we find that the refinement of OHC afferent innervation is regulated by complementary spontaneous Ca2+ signals originating in OHCs and non‐sensory cells. OHCs fire spontaneous Ca2+ action potentials during a narrow period of neonatal development. Simultaneously, waves of Ca2+ activity in the non‐sensory cells of the greater epithelial ridge cause, via ATP‐induced activation of P2X3 receptors, the increase and synchronization of the Ca2+ activity in nearby OHCs. This synchronization is required for the refinement of their immature afferent innervation. In the absence of connexin channels, Ca2+ waves are impaired, leading to a reduction in the number of ribbon synapses and afferent fibres on OHCs. We propose that the correct maturation of the afferent connectivity of OHCs requires experience‐independent Ca2+ signals from sensory and non‐sensory cells.

In the adult auditory organ, mechanoelectrical transducer (MET) channels are essential for transducing acoustic stimuli into electrical signals. In the absence of incoming sound, a fraction of the MET channels on top of the sensory hair cells are open, resulting in a sustained depolarizing current. By genetically manipulating the in vivo expression of molecular components of the MET apparatus, we show that during pre-hearing stages the MET current is essential for establishing the electrophysiological properties of mature inner hair cells (IHCs). If the MET current is abolished in adult IHCs, they revert into cells showing electrical and morphological features characteristic of pre-hearing IHCs, including the re-establishment of cholinergic efferent innervation. The MET current is thus critical for the maintenance of the functional properties of adult IHCs, implying a degree of plasticity in the mature auditory system in response to the absence of normal transduction of acoustic signals.

Key pointsr Outer hair cells (OHCs) enhance the sensitivity and the frequency tuning of the mammalian cochlea.r Similar to the primary sensory receptor, the inner hair cells (IHCs), the mature functional characteristics of OHCs are acquired before hearing onset.r We found that OHCs, like IHCs, fire spontaneous Ca 2+ -induced action potentials (APs) during immature stages of development, which are driven by Ca V 1.3 Ca 2+ channels.r We also showed that the development of low-and high-frequency hair cells is differentially regulated during pre-hearing stages, with the former cells being more strongly dependent on experience-independent Ca 2+ action potential activity.Abstract Sound amplification within the mammalian cochlea depends upon specialized hair cells, the outer hair cells (OHCs), which possess both sensory and motile capabilities. In various altricial rodents, OHCs become functionally competent from around postnatal day 7 (P7), before the primary sensory inner hair cells (IHCs), which become competent at about the onset of hearing (P12). The mechanisms responsible for the maturation of OHCs and their synaptic specialization remain poorly understood. We report that spontaneous Ca 2+ activity in the immature cochlea, which is generated by Ca V 1.3 Ca 2+ channels, differentially regulates the maturation of hair cells along the cochlea. Under near-physiological recording conditions we found that, similar to IHCs, immature OHCs elicited spontaneous Ca 2+ action potentials (APs), but only duringThis is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. 152 J.-Y. Jeng and others J Physiol 598.1the first few postnatal days. Genetic ablation of these APs in vivo, using Ca V 1.3 −/− mice, prevented the normal developmental acquisition of mature-like basolateral membrane currents in low-frequency (apical) hair cells, such as I K,n (carried by KCNQ4 channels), I SK2 and I ACh (α9α10nAChRs) in OHCs and I K,n and I K,f (BK channels) in IHCs. Electromotility and prestin expression in OHCs were normal in Ca V 1.3 −/− mice. The maturation of high-frequency (basal) hair cells was also affected in Ca V 1.3 −/− mice, but to a much lesser extent than apical cells. However, a characteristic feature in Ca V 1.3 −/− mice was the reduced hair cell size irrespective of their cochlear location. We conclude that the development of low-and high-frequency hair cells is differentially regulated during development, with apical cells being more strongly dependent on experience-independent Ca 2+ APs.

Eye movements depend on correct patterns of connectivity between cranial motor axons and the extraocular muscles. Despite the clinical importance of the ocular motor system, little is known of the molecular mechanisms underlying its development. We have recently shown that mutations in the Chimaerin-1 gene encoding the signaling protein α2-chimaerin (α2-chn) perturb axon guidance in the ocular motor system and lead to the human eye movement disorder, Duane retraction syndrome (DRS). The axon guidance cues that lie upstream of α2-chn are unknown; here we identify candidates to be the Semaphorins (Sema) 3A and 3C, acting via the PlexinA receptors. Sema3A/C are expressed in and around the developing extraocular muscles and cause growth cone collapse of oculomotor neurons in vitro. Furthermore, RNAi knockdown of α2-chn or PlexinAs in oculomotor neurons abrogates Sema3A/C-dependent growth cone collapse. In vivo knockdown of endogenous PlexinAs or α2-chn function results in stereotypical oculomotor axon guidance defects, which are reminiscent of DRS, whereas expression of α2-chn gain-of-function constructs can rescue PlexinA loss of function. These data suggest that α2-chn mediates Sema3-PlexinA repellent signaling. We further show that α2-chn is required for oculomotor neurons to respond to CXCL12 and hepatocyte growth factor (HGF), which are growth promoting and chemoattractant during oculomotor axon guidance. α2-chn is therefore a potential integrator of different types of guidance information to orchestrate ocular motor pathfinding. DRS phenotypes can result from incorrect regulation of this signaling pathway.