Daniel J. Jagger

Objective . Cochlear implants (CI) partially restore hearing in profoundly hearing-impaired or deaf individuals by electrically stimulating the auditory nerve. A primary bottleneck in electrical CIs is the broad spread of electrical current from each electrode that limits the transfer of spectral information, which might be overcome by future spatially confined optogenetic stimulation. Here we established an in silico framework, FraSCO, to model sound encoding in the human cochlea by an optical CI (oCI) for testing the potential of optogenetic hearing restoration. Approach . The biophysical modelling framework combined an optical ray tracing model implementing a human cochlea implanted with a waveguide-based oCI with a single compartment model of optogenetically modified spiral ganglion neurons (SGNs). The input was an optogenetic sound coding strategy and the quality of the neural representation was evaluated based on comparison of neurograms evoked by optogenetic and electrical stimulation to the spectrogram of the sound applied. The model incorporated technologically feasible properties of the oCIs with 64 stimulation channels. Main results . The biophysical modelling framework successfully captured essential physiological features of optogenetic SGN stimulation with a minimal set of ion channel types expressed in the SGN soma. Working with a sample of 1000 SGNs distributed along the tonotopic axis to represent sound encoding, we found that improved spectral selectivity achieves comparable information encoding to electrical implants despite lower stimulation rates in current implementations of optogenetic stimulation. We also found that potential oCI users are predicted to be less impacted by noisy backgrounds. Significance . The established computational framework enables an in silico investigation and benchmarking of sound encoding in the cochlea by future oCI and state-of-the-art electrical CI. The results of the exemplary application indicate that optogenetic sound encoding has potential to improve speech understanding in noisy environments for CI users.

Cochlear implants (CI) partially restore hearing in profoundly hearing impaired or deaf people by electrically stimulating the auditory nerve. A bottleneck of electrical CIs is the broad spread of electrical current from each electrode that limits the transfer of spectral information, which might be overcome by future spatially confined optogenetic stimulation. Here we established an in silico framework, FraSCO, to model sound encoding in the human cochlea by an optogenetic CI (oCI) for testing the potential of optogenetic hearing restoration. The biophysical modeling framework combined an optical raytracing model implementing a human cochlea implanted with a waveguide-based oCI with a single compartment model of optogenetically modified spiral ganglion neurons (SGNs). The input was an optogenetic sound coding strategy and the quality of the neural representation was evaluated based on comparison of neurograms evoked by optogenetic and electrical stimulation to the spectrogram of the sound applied. The model aimed for technologically feasible properties of the oCIs with 64 stimulation channels. The biophysical modeling framework successfully captured essential physiological features of optogenetic SGN stimulation with a minimal set of ion channel types expressed in the SGN soma. Working with a sample of 1000 SGNs distributed along the tonotopic axis to represent sound encoding, we found that improved spectral selectivity more than compensates for lower temporal fidelity of current implementations of optogenetic stimulation. The established computational framework enables in silico investigation and benchmarking of sound encoding in the cochlea by future oCI and state-of-the-art eCI. The results indicate that optogenetic sound encoding has potential to improve speech understanding in noisy environments for CI users.

In multicellular tissues, the size and shape of cells are intricately linked with their physiological functions. In the vertebrate auditory organ, the neurosensory epithelium develops as a mosaic of sensory hair cells (HCs), and their glial-like supporting cells, which have distinct morphologies and functional properties at different frequency positions along its tonotopic long axis. In the chick cochlea, the basilar papilla (BP), proximal (high-frequency) HCs, are larger than their distal (low-frequency) counterparts, a morphological feature essential for sound perception. Mitochondrial dynamics, which constitute the equilibrium between fusion and fission, regulate differentiation and functional refinement across a variety of cell types. We investigate this as a potential mechanism for regulating the shape of developing HCs. Using live imaging in intact BP explants, we identify distinct remodelling of mitochondrial networks in proximal compared with distal HCs. Manipulating mitochondrial dynamics in developing HCs alters their normal morphology along the proximal-distal (tonotopic) axis. Inhibition of the mitochondrial fusion machinery decreased proximal HC surface area, whereas promotion of fusion increased the distal HC surface area. We identify mitochondrial dynamics as a key regulator of HC morphology in developing inner ear epithelia.

In multicellular tissues, cell size and shape are intricately linked with physiological function. In the vertebrate auditory organ, the neurosensory epithelium develops as a mosaic of sensory hair cells (HCs), and their glial-like supporting cells (SCs), which have distinct morphologies at different frequency positions along its tonotopic long axis. In the chick cochlea, the basilar papilla (BP), proximal (high-frequency) HCs are larger than their distal (low-frequency) counterparts, a morphological feature essential for frequency tuning. Mitochondrial dynamics, which constitute the equilibrium between fusion and fission, regulate differentiation and functional refinement across a variety of cell types. We investigate this a potential mechanism for cell size regulation in developing HCs. Using live imaging in intact BP explants, we identify distinct remodelling of mitochondrial networks in proximal compared to distal HCs. Manipulating mitochondrial dynamics in developing HCs alters their normal morphology along the proximal-distal (tonotopic) axis. Inhibition of the mitochondrial fusion machinery decreased proximal HC size, whilst promotion of fusion increased the distal HC size. We identify mitochondrial dynamics as a key regulator of HC size and morphology in developing inner ear epithelia. Summary Statement Mitochondrial remodelling drives developmental changes in cell size in the auditory sensory epithelium. Our data reveal a fundamental mechanism regulating cell size and frequency-place coding in the developing cochlea.

Our sense of hearing depends on the function of a specialised class of sensory cells, the hair cells, which are found in the organ of Corti of the mammalian cochlea. The unique physiological environment in which these cells operate is maintained by a syncitium of non‐sensory supporting cells, which are crucial for regulating cochlear physiology and metabolic homeostasis. Despite their importance for cochlear function, the role of these supporting cells in age‐related hearing loss, the most common sensory deficit in the elderly, is poorly understood. Here, we investigated the age‐related changes in the expression and function of metabotropic purinergic receptors (P2Y₁, P2Y₂ and P2Y₄) in the supporting cells of the cochlear apical coil. Purinergic signalling in supporting cells is crucial during the development of the organ of Corti and purinergic receptors are known to undergo changes in expression during ageing in several tissues. Immunolabelling and Ca²⁺ imaging experiments revealed a downregulation of P2Y receptor expression and a decrease of purinergic‐mediated calcium responses after early postnatal stages in the supporting cells. An upregulation of P2Y receptor expression was observed in the aged cochlea when compared to 1 month‐old adults. The aged mice also had significantly larger calcium responses and displayed calcium oscillations during prolonged agonist applications. We conclude that supporting cells in the aged cochlea upregulate P2Y₂ and P2Y₄ receptors and display purinergic‐induced Ca²⁺ responses that mimic those observed during pre‐hearing stages of development, possibly aimed at limiting or preventing further damage to the sensory epithelium. image Key points: Age‐related hearing loss is associated with lower hearing sensitivity and decreased ability to understand speech. We investigated age‐related changes in the expression and function of metabotropic purinergic (P2Y) receptors in cochlear non‐sensory supporting cells of mice displaying early‐onset (C57BL/6N) and late‐onset (C3H/HeJ) hearing loss. The expression of P2Y₁, P2Y₂ and P2Y₄ receptors in the supporting cells decreased during cochlear maturation, but that of P2Y₂ and P2Y₄ was upregulated in the aged cochlea. P2Y₂ and P2Y₄ receptors were primarily responsible for the ATP‐induced Ca²⁺ responses in the supporting cells. The degree of purinergic expression upregulation in aged supporting cells mirrored hearing loss progression in the different mouse strains. We propose that the upregulation of purinergic‐mediated signalling in the aged cochlea is subsequent to age‐related changes in the hair cells and may act as a protective mechanism to limit or to avoid further damage to the sensory epithelium.

Norrie disease is caused by mutation of the NDP gene, presenting as congenital blindness followed by later onset of hearing loss. Protecting patients from hearing loss is critical for maintaining their quality of life. This study aimed to understand the onset of pathology in cochlear structure and function. By investigating patients and juvenile Ndp -mutant mice, we elucidated the sequence of onset of physiological changes (in auditory brainstem responses, distortion product otoacoustic emissions, endocochlear potential, blood-labyrinth barrier integrity) and determined the cellular, histological, and ultrastructural events leading to hearing loss. We found that cochlear vascular pathology occurs earlier than previously reported and precedes sensorineural hearing loss. The work defines a disease mechanism whereby early malformation of the cochlear microvasculature precedes loss of vessel integrity and decline of endocochlear potential, leading to hearing loss and hair cell death while sparing spiral ganglion cells. This provides essential information on events defining the optimal therapeutic window and indicates that early intervention is needed. In an era of advancing gene therapy and small-molecule technologies, this study establishes Ndp -mutant mice as a platform to test such interventions and has important implications for understanding the progression of hearing loss in Norrie disease.

P2X₇receptors (P2X₇Rs) are associated with numerous pathophysiological mechanisms, and this promotes them as therapeutic targets for certain neurodegenerative conditions. However, the identity of P2X₇R-expressing cells in the nervous system remains contentious. Here, we examined P2X₇R functionality in auditory nerve cells from rodents of either sex, and determined their functional and anatomic expression pattern. In whole-cell recordings from rat spiral ganglion cultures, the purinergic agonist 2',3'-O-(4-benzoylbenzoyl)-ATP (BzATP) activated desensitizing currents in spiral ganglion neurons (SGNs) but non-desensitizing currents in glia that were blocked by P2X₇R-specific antagonists. In imaging experiments, BzATP gated sustained Ca²⁺entry into glial cells. BzATP-gated uptake of the fluorescent dye YO-PRO-1 was reduced and slowed by P2X₇R-specific antagonists. In rats, P2X₇Rs were immuno-localized predominantly within satellite glial cells (SGCs) and Schwann cells (SCs). P2X₇R expression was not detected in the portion of the auditory nerve within the central nervous system. Mouse models allowed further exploration of the distribution of cochlear P2X₇Rs. In GENSAT reporter mice, EGFP expression driven via theP2rx7promoter was evident in SGCs and SCs but was undetectable in SGNs. A second transgenic model showed a comparable cellular distribution of EGFP-tagged P2X₇Rs. In wild-type mice the discrete glial expression was confirmed using a P2X₇-specific nanobody construct. Our study shows that P2X₇Rs are expressed by peripheral glial cells, rather than by afferent neurons. Description of functional signatures and cellular distributions of these enigmatic proteins in the peripheral nervous system (PNS) will help our understanding of ATP-dependent effects contributing to hearing loss and other sensory neuropathies.SIGNIFICANCE STATEMENTP2X₇receptors (P2X₇Rs) have been the subject of much scrutiny in recent years. They have been promoted as therapeutic targets in a number of diseases of the nervous system, yet the specific cellular location of these receptors remains the subject of intense debate. In the auditory nerve, connecting the inner ear to the brainstem, we show these multimodal ATP-gated channels localize exclusively to peripheral glial cells rather than the sensory neurons, and are not evident in central glia. Physiologic responses in the peripheral glia display classical hallmarks of P2X₇R activation, including the formation of ion-permeable and also macromolecule-permeable pores. These qualities suggest these proteins could contribute to glial-mediated inflammatory processes in the auditory periphery under pathologic disease states.

Spiral ganglion neurons (SGNs) are the primary afferent neurons of the auditory system, and together with their attendant glia, form the auditory nerve. Within the cochlea, satellite glial cells (SGCs) encapsulate the cell body of SGNs, whereas Schwann cells (SCs) wrap their peripherally‐ and centrally‐directed neurites. Despite their likely importance in auditory nerve function and homeostasis, the physiological properties of auditory glial cells have evaded description. Here, we characterized the voltage‐activated membrane currents of glial cells from the mouse cochlea. We identified a prominent weak inwardly rectifying current in SGCs within cochlear slice preparations (postnatal day P5‐P6), which was also present in presumptive SGCs within dissociated cultures prepared from the cochleae of hearing mice (P14‐P15). Pharmacological block by Ba²⁺ and desipramine suggested that channels belonging to the Kir4 family mediated the weak inwardly rectifying current, and post hoc immunofluorescence implicated the involvement of Kir4.1 subunits. Additional electrophysiological profiles were identified for glial cells within dissociated cultures, suggesting that glial subtypes may have specific membrane properties to support distinct physiological roles. Immunofluorescence using fixed cochlear sections revealed that although Kir4.1 is restricted to SGCs after the onset of hearing, these channels are more widely distributed within the glial population earlier in postnatal development (i.e., within both SGCs and SCs). The decrease in Kir4.1 immunofluorescence during SC maturation was coincident with a reduction of Sox2 expression and advancing neurite myelination. The data suggest a diversification of glial properties occurs in preparation for sound‐driven activity in the auditory nerve.