Biology subjects

Marcotti, W.

Publications and source records attributed to Marcotti, W..

8 recordsLinked to original sources

bioRxiv2026

Estrogen-related receptor gamma is required for normal auditory innervation and is essential for hearing.

Estrogen-related receptor gamma (ESRRG), an orphan nuclear receptor with structural homology to the classical estrogen receptors, is widely recognised as a key metabolic regulator involved in mitochondrial, synaptic, and ion-homeostatic pathways. Previous clinical studies suggest a link between ESRRG and auditory function; for example, ESRRG has been associated with susceptibility to age-related hearing loss in women and implicated in congenital hearing loss. However, the biological mechanisms by which ESRRG may mediate hearing function remain largely unknown. Here, using a combination of in vivo auditory physiological recordings, immunofluorescence analyses, single hair-cell electrophysiology, and transcriptomic approaches, we characterise the phenotype of a new inner-ear conditional Esrrg knockout (Esrrg-cKO) to investigate the role of Esrrg in the auditory system. We found that Esrrg-cKO mice of both sexes develop early-onset hearing loss, as evidenced by elevated auditory brainstem response thresholds and reduced wave 1 amplitudes from two weeks of age. These auditory deficits arise from a combination of early-onset cochlear neuronal and innervation malformations, together with inner hair cell synaptic defects and delayed myelination that persist into adulthood. Furthermore, distortion product otoacoustic emissions and endocochlear potential recordings are normal in Esrrg-cKO mice, and although sensory hair cells are preserved, IHCs retain immature biophysical properties. These findings are consistent with auditory neuropathy, and together with our comparative transcriptome analyses, indicate that Esrrg is an essential molecular driver of normal cochlear innervation and maturation.

neuroscience↗

bioRxiv2024

An Open-Source Deep Learning-Based GUI Toolbox For Automated Auditory Brainstem Response Analyses (ABRA)

Hearing loss is a pervasive global health challenge with profound impacts on communication, cognitive function, and quality of life. Recent studies have established age-related hearing loss as a significant risk factor for dementia, highlighting the importance of hearing loss research. Auditory brainstem responses (ABRs), which are electrophysiological recordings of acoustically evoked synchronized neural activity from the auditory nerve and brainstem, serve as in vivo correlates for sensory hair cell and synaptic function, hearing sensitivity, and other critical readouts of auditory pathway physiology, making them highly valuable for both basic neuroscience and clinical research. Despite their utility, traditional ABR analyses rely heavily on subjective manual interpretation, which may introduce variability and pose challenges for reproducibility across studies. Here, we introduce Auditory Brainstem Response Analyzer (ABRA), a novel suite of open-source ABR analysis tools powered by deep learning, which automates and standardizes ABR waveform analysis. ABRA employs convolutional neural networks trained on diverse datasets collected from multiple experimental settings, achieving rapid and unbiased extraction of key ABR metrics, including peak amplitude, latency, and auditory threshold estimates. We demonstrate that ABRAs deep learning models provide performance comparable to expert human annotators while dramatically reducing analysis time and enhancing reproducibility across datasets from different laboratories. By bridging hearing research, sensory neuroscience, and advanced computational techniques, ABRA facilitates broader interdisciplinary insights into auditory function. An online version of the tool is available for use at no cost at https://abra.ucsd.edu.

neuroscience↗

bioRxiv2024

Lack of Oncomodulin Increases ATP-Dependent Calcium Signaling and Susceptibility to Noise in Adult Mice

Tight regulation of Ca2+ is crucial for the function of cochlear outer hair cells (OHCs). Dysregulation of Ca2+ homeostasis in OHCs is associated with impaired hearing and contributes to increased vulnerability to insults such as noise exposure. Ca2+ signaling in developing OHCs is modulated by oncomodulin (OCM), an EF-hand calcium-binding protein. Here, we investigated whether the lack of OCM disrupts the control of intracellular Ca2+ in mature OHCs, and influences vulnerability to acoustic injury. Using young adult CBA/CaJ mice, we found that OHCs from Ocm-knockout (Ocm-/-) mice showed normal biophysical profiles, electromotile responses, and synaptic innervation compared to littermate controls. Moderate noise exposure (95 dB SPL, 2 hrs) caused temporary threshold shifts in Ocm+/+ and Ocm-/- mice. However, while Ocm+/+ fully recovered thresholds 2 weeks after noise exposure, Ocm-/- mice showed permanent threshold shifts. Additionally, Ocm-/- mice had auditory brainstem responses with highly variable latencies and amplitudes both before and after noise exposure compared to Ocm+/+ mice. Using a genetically encoded Ca2+ sensor (GCaMP6s) expressed in Ocm+/+ and Ocm-/- OHCs, we found that prolonged noise exposure (95 dB SPL, 9 hrs) significantly increased GCaMP6s fluorescence, ATP-induced Ca2+ signaling and also caused greater threshold shifts in Ocm-/- compared to Ocm+/+ OHCs. However, prolonged noise exposure had no significant change in the number of presynaptic OHC ribbons in either Ocm+/+ or Ocm-/- mice. We assessed whether the ATP-induced responses were due to changes in P2X2 receptor expression. Prior to noise exposure, P2X2 expression was higher in the cochlea of Ocm-/- mice compared to Ocm+/+ mice. Following prolonged noise, P2X2 receptors were upregulated in the cochlea of Ocm+/+ but not in the Ocm-/- mice, which retained their pre-noise expression level. We propose that the lack of OCM increases susceptibility to cochlear pathology and that purinergic signaling and dysregulation of cytosolic Ca2+ homeostasis likely contribute to early onset hearing loss in the Ocm-/- mice.

neuroscience↗

bioRxiv2023

Oncomodulin Regulates Spontaneous Calcium Signaling and Maturation of Afferent Innervation in Cochlear Outer Hair Cells

Cochlear outer hair cells (OHCs) are responsible for the exquisite frequency selectivity and sensitivity of mammalian hearing. During development, the maturation of OHC afferent connectivity is refined by coordinated spontaneous Ca2+ activity in both sensory and non-sensory cells. Calcium signaling in neonatal OHCs can be modulated by Oncomodulin (OCM, {beta}-parvalbumin), an EF-hand calcium-binding protein. Here, we investigated whether OCM regulates OHC spontaneous Ca2+ activity and afferent connectivity during development. Using a genetically encoded Ca2+ sensor (GCaMP6s) expressed in OHCs in wild-type (Ocm+/+) and Ocm knockout (Ocm-/-) littermates, we found increased spontaneous Ca2+ activity and upregulation of purinergic receptors in OHCs from GCaMP6s Ocm-/- cochlea immediately following birth. The afferent synaptic maturation of OHCs was delayed in the absence of OCM, leading to an increased number of ribbon synapses and afferent fibers on GCaMP6s Ocm-/- OHCs before hearing onset. We propose that OCM regulates the spontaneous Ca2+ signaling in the developing cochlea and the maturation of OHC afferent innervation.

developmental biology↗

bioRxiv2022

Oncomodulin (OCM) uniquely regulates calcium signaling in neonatal cochlear outer hair cells.

In cochlear outer hair cells (OHCs), a network of Ca2+ channels, pumps and Ca2+-binding proteins (CaBPs) regulates the localization, spread, and magnitude of free Ca2+ ions. During early postnatal development, OHCs express three prominent mobile EF-hand CaBPs: oncomodulin (OCM), -parvalbumin (APV) and sorcin. We have previously shown that deletion of Ocm (Ocm-/-) gives rise to progressive cochlear dysfunction in young adult mice. Here, we show that changes in Ca2+ signaling begin early in postnatal development of Ocm-/- mice. While mutant OHCs exhibit normal electrophysiological profiles compared to controls, their intracellular Ca2+ signaling is altered. The onset of OCM expression at postnatal day 3 (P3) causes a developmental change in KCl-induced Ca2+ transients in OHCs and leads to slower KCl-induced Ca2+ transients than those elicited in cells from Ocm-/- littermates. We compared OCM buffering kinetics with other CaBPs in animal models and cultured cells. In a double knockout of Ocm and Apv (Ocm-/-;Apv-/-), mutant OHCs show even faster Ca2+ kinetics, suggesting that APV may also contribute to early postnatal Ca2+ signaling. In transfected HEK293T cells, OCM slows Ca2+ kinetics more so than either APV or sorcin. We conclude that OCM controls the intracellular Ca2+ environment by lowering the amount of freely available [Ca2+]i in OHCs and in transfected HEK293T cells. We propose that OCM plays an important role in shaping the development of early OHC Ca2+ signals through its inimitable Ca2+ buffering capacity.

developmental biology↗

bioRxiv2021

Neuroplastin genetically interacts with Cadherin 23 and the encoded isoform Np55 is sufficient for cochlear hair cell function and hearing

Mammalian hearing involves the mechanoelectrical transduction (MET) of sound-induced fluid waves in the cochlea. Essential to this process are the specialised sensory cochlear cells, the inner (IHCs) and outer hair cells (OHCs). While genetic hearing loss is highly heterogeneous, understanding the requirement of each gene will lead to a better understanding of the molecular basis of hearing and also to therapeutic opportunities for deafness. The Neuroplastin (Nptn) gene, which encodes two protein isoforms Np55 and Np65, is required for hearing, and homozygous loss-of-function mutations that affect both isoforms lead to profound deafness in mice. Here we have utilised several distinct mouse models to elaborate upon the spatial, temporal, and functional requirement of Nptn for hearing. While we demonstrate that both Np55 and Np65 are present in cochlear cells, characterisation of a Np65-specific mouse knockout shows normal hearing thresholds indicating that Np65 is functionally redundant for hearing. In contrast, we find that Nptn-knockout mice have significantly reduced maximal MET currents and MET channel open probabilities in mature OHCs, with both OHCs and IHCs also failing to develop fully mature basolateral currents. Furthermore, comparing the hearing thresholds and IHC synapse structure of Nptn-knockout mice with those of mice that lack Nptn only in IHCs and OHCs shows that the majority of the auditory deficit is explained by hair cell dysfunction, with abnormal afferent synapses contributing only a small proportion of the hearing loss. Finally, we show that continued expression of NEUROPLASTIN in OHCs of adult mice is required for membrane localisation of Plasma Membrane Ca2+ ATPase 2 (PMCA2), which is essential for hearing function. Moreover, Nptn haploinsufficiency phenocopies Atp2b2 (encodes PMCA2) mutations, with heterozygous Nptn-knockout mice exhibiting hearing loss through genetic interaction with the Cdh23ahl allele. Together, our findings provide further insight to the functional requirement of Neuroplastin for mammalian hearing. Author SummarySensorineural hearing loss, caused by problems with sensory cells in the cochlea or the auditory nerve, is the most common type of hearing loss. Mutations in Neuroplastin have already been implicated in deafness in mice. We have used mutant mouse models to investigate where Neuroplastin is expressed in the cochlea and its function. When mice do not express a functioning copy of Neuroplastin they have disruptions to the primary sensory synapse. We show that although synaptic disruption contributes to the loss of hearing function it is not the primary cause. Instead, continued expression of Neuroplastin is needed to maintain the localisation of Plasma Membrane Ca2+ ATPase 2 channels which help regulate calcium flow. We have also shown that two types of NEUROPLASTIN protein (isoforms) are both expressed within the cochlea, although only one of these isoforms needs to be expressed for normal hearing. Finally, we also demonstrate that the hearing loss caused by the absence of Neuroplastin is made worse when combined with a common mutation within a gene called Cadherin 23 (Cdh23ahl). This is an important finding as although there are currently no human patients with an identified NEUROPLASTIN mutation, it may be involved in human deafness in combination with other mutations.

neuroscience↗

bioRxiv2019

Dynamic coupling of cochlear inner hair cell intrinsic Ca2+ action potentials to Ca2+ signaling of non-sensory cells

The relationship between Ca2+ action potential (AP) activity in immature inner hair cells (IHCs) and the spontaneous ATP-dependent intercellular Ca2+ signaling in cochlear non-sensory cells (NSCs) of the greater epithelial ridge (GER) is unclear. Here, we determined that IHCs fired asynchronous Ca2+ APs also in the absence of Ca2+ activity in the GER. Patch clamp recordings from IHCs isolated from the rest of the sensory epithelium confirmed that this firing activity is an intrinsic property of immature IHCs. However, frequency, correlation index and burst duration of IHC APs increased significantly during Ca2+ wave propagation in NSCs, and depended on wave extension in the GER. Furthermore, IHC depolarization under whole cell patch clamp conditions triggered Ca2+ signals in nearby NSCs with a delay that was proportional to the distance from the stimulated IHC. Thus the immature mammalian cochlea supports bidirectional exchange of Ca2+ signals between IHCs and NSCs.\n\nIMPACT STATEMENTIn inner hair cells of the developing mammalian cochlea, Ca2+ action potentials are both intrinsic and bidirectionally coupled to the ATP-dependent Ca2+ signaling of the surrounding non-sensory cells.

developmental biology↗

bioRxiv2018

Spontaneous and coordinated Ca2+ activity of cochlear sensory and non-sensory cells drives the maturation of OHC afferent innervation

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 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+ spikes during a narrow period of immature development. Simultaneously, waves of Ca2+ activity in the non-sensory greater epithelial ridge act, via ATP-induced activation of P2X receptors, to synchronize OHC firing, resulting in the refinement of their 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 in OHCs requires experience-independent Ca2+ signals from sensory and non-sensory cells.

neuroscience↗