Oxidative stress has been established as the key mechanism of the cochlear damage underlying noise-induced hearing loss, however, emerging evidence suggests that cochlear inflammation may also be a major contributor. This study aimed to improve our understanding of the cochlear inflammatory response associated with acute and chronic noise exposure. C57BL/6 mice were exposed to acute traumatic noise (100 dBSPL, 8-16 kHz for 24 h) and their cochleae collected at various intervals thereafter, up to 7 days. Using quantitative RT-PCR and immunohistochemistry, changes in expression levels of proinflammatory cytokines (TNF-α, IL-1β), chemokines (CCL2) and cell adhesion molecules (ICAM-1) were studied. All gene transcripts displayed similar dynamics of expression, with an early upregulation at 6 h post-exposure, followed by a second peak at 7 days. ICAM-1 immunoexpression increased significantly in the inferior region of the spiral ligament, peaking 24 h post-exposure. The early expression of proinflammatory mediators likely mediates the recruitment and extravasation of inflammatory cells into the noise-exposed cochlea. The occurrence of the latter expression peak is not clear, but it may be associated with reparative processes initiated in response to cochlear damage. Chronic exposure to moderate noise (90 dBSPL, 8-16 kHz, 2 h/day, up to 4 weeks) also elicited an inflammatory response, reaching a maximum after 2 weeks, suggesting that cochlear damage and hearing loss associated with chronic environmental noise exposure may be linked to inflammatory processes in the cochlea. This study thus provides further insight into the dynamics of the cochlear inflammatory response induced by exposure to acute and chronic noise.

The voltage-dependent protein SLC26a5 (prestin) underlies outer hair cell electromotility (eM), which is responsible for cochlear amplification in mammals. The electrical signature of eM is a bell-shaped nonlinear capacitance (NLC), deriving from prestin sensor-charge (Q) movements, which peaks at the membrane voltage, V, where charge is distributed equally on either side of the membrane. Voltage dependencies of NLC and eM differ depending on interrogation frequency and intracellular chloride, revealing slow intermediate conformational transitions between anion binding and voltage-driven Q movements. Consequently, NLC exhibits low-pass characteristics, substantially below prevailing estimates of eM frequency response. Here we study in guinea pig and mouse of either sex synchronous prestin electrical (NLC, Q) and mechanical (eM) activity across frequencies under voltage clamp (whole cell and microchamber). We find that eM and Q magnitude and phase correspond, indicating tight piezoelectric coupling. Electromechanical measures (both NLC and eM) show dual-Lorentzian, low-pass behavior, with a limiting (τ) time constant at V of 32.6 and 24.8 μs, respectively. As expected for voltage-dependent kinetics, voltage excitation away from V has a faster, flatter frequency response, with our fastest measured τ for eM of 18.2 μs. Previous observations of ultrafast eM (τ ≈ 2 μs) were obtained at offsets far removed from V We hypothesize that trade-offs in eM gain-bandwith arising from voltage excitation at membrane potentials offset from V influence the effectiveness of cochlear amplification across frequencies. Of two types of hair cells within the organ of Corti, inner hair cells and outer hair cells, the latter evolved to boost sensitivity to sounds. Damage results in hearing loss of 40-60 dB, revealing amplification gains of 100-1000× that arise from voltage-dependent mechanical responses [electromotility (eM)]. eM, driven by the membrane protein prestin, may work beyond 70 kHz. However, this speed exceeds, by over an order of magnitude, kinetics of typical voltage-dependent membrane proteins. We find eM is actually low pass in nature, indicating that prestin bears kinetics typical of other membrane proteins. These observations highlight potential difficulties in providing sufficient amplification beyond a cutoff frequency near 20 kHz. Nevertheless, observed trade-offs in eM gain-bandwith may sustain cochlear amplification across frequency.

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Fragile X syndrome (FXS) is characterized by hypersensitivity to sensory stimuli, including environmental sounds. We compared the auditory brainstem response (ABR) recorded in vivo in mice lacking the gene (Fmr1 ؊/y ) for fragile X mental retardation protein (FMRP) with that in wild-type animals. We found that ABR wave I, which represents input from the auditory nerve, is reduced in Fmr1 ؊/y animals, but only at high sound levels. In contrast, wave IV, which represents the activity of auditory brainstem nuclei is enhanced at all sound levels, suggesting that loss of FMRP alters the central processing of auditory signals. Current-clamp recordings of neurons in the medial nucleus of the trapezoid body in the auditory brainstem revealed that, in contrast to neurons from wild-type animals, sustained depolarization triggers repetitive firing rather than a single action potential. In voltage-clamp recordings, K ϩ currents that activate at positive potentials ("high-threshold" K ϩ currents), which are required for high-frequency firing and are carried primarily by Kv3.1 channels, are elevated in Fmr1 ؊/y mice, while K ϩ currents that activate near the resting potential and inhibit repetitive firing are reduced. We therefore tested the effects of AUT2 [((4-({5-[(4R)-4-ethyl-2,5-dioxo-1-imidazolidinyl]-2-pyridinyl}oxy)-2-(1-methylethyl) benzonitrile], a compound that modulates Kv3.1 channels. AUT2 reduced the high-threshold K ϩ current and increased the low-threshold K ϩ currents in neurons from Fmr1 ؊/y animals by shifting the activation of the high-threshold current to more negative potentials. This reduced the firing rate and, in vivo, restored wave IV of the ABR. Our results from animals of both sexes suggest that the modulation of the Kv3.1 channel may have potential for the treatment of sensory hypersensitivity in patients with FXS. Key words: auditory brainstem response; AUT2; fragile X; high-and low-threshold potassium channels; medial nucleus of the trapezoid body; potassium channelsSignificance Statement mRNA encoding the Kv3.1 potassium channel was one of the first described targets of the fragile X mental retardation protein (FMRP). Fragile X syndrome is caused by loss of FMRP and, in humans and mice, causes hypersensitivity to auditory stimuli. We found that components of the auditory brain response (ABR) corresponding to auditory brainstem activity are enhanced in mice lacking FMRP. This is accompanied by hyperexcitability and altered potassium currents in auditory brainstem neurons. Treatment with a drug that alters the voltage dependence of Kv3.1 channels normalizes the imbalance of potassium currents, as well as ABR responses in vivo, suggesting that such compounds may be effective in treating some symptoms of fragile X syndrome.

The mammalian outer hair cell (OHC) protein prestin (Slc26a5) differs from other Slc26 family members due to its unique piezoelectric-like property that drives OHC electromotility, the putative mechanism for cochlear amplification. Here, we use cryo-electron microscopy to determine prestin’s structure at 3.6 Å resolution. Prestin is structurally similar to the anion transporter Slc26a9. It is captured in an inward-open state which may reflect prestin’s contracted state. Two well-separated transmembrane (TM) domains and two cytoplasmic sulfate transporter and anti-sigma factor antagonist (STAS) domains form a swapped dimer. The transmembrane domains consist of 14 transmembrane segments organized in two 7+7 inverted repeats, an architecture first observed in the bacterial symporter UraA. Mutation of prestin’s chloride binding site removes salicylate competition with anions while retaining the prestin characteristic displacement currents (Nonlinear Capacitance), undermining the extrinsic voltage sensor hypothesis for prestin function.

Since chronic mitochondrial dysfunction is common in many patients, it could lead to developing hearing loss that can be alleviated/rescued by AO treatment. Our study creates a framework for clinical trials and introduces the Fus1 KO model as a powerful platform for developing novel therapeutic strategies to prevent/delay hearing loss associated with mitochondrial dysfunction. Antioxid. Redox Signal. 27, 489-509.

Outer hair cell (OHC) nonlinear capacitance (NLC) represents voltage sensor charge movements of prestin (SLC26a5), the protein responsible for OHC electromotility. Previous measures of NLC frequency response have employed methods which did not assess the influence of dielectric loss (sensor charge movements out of phase with voltage) that may occur, and such loss conceivably may influence prestin’s frequency dependent activity. Here we evaluate prestin’s complex capacitance out to 30 kHz and find that prestin’s frequency response determined using this approach coincides with all previous estimates. We also show that membrane tension has no effect on prestin’s frequency response, despite substantial shifts in its voltage operating range, indicating that prestin transition rate alterations do not account for the shifts. The magnitude roll-off of prestin activity across frequency surpasses the reductions of NLC caused by salicylate treatments that are known to abolish cochlear amplification. Such roll-off likely limits the effectiveness of prestin in contributing to cochlear amplification at the very high acoustic frequencies processed by some mammals.

Decreased energy production and increased oxidative stress are considered to be major contributors to aging and aging-associated pathologies. The role of mitochondrial calcium homeostasis has also been highlighted as an important factor affecting different pathological conditions. Here, we present evidence that loss of a small mitochondrial protein Fus1 that maintains mitochondrial homeostasis results in premature aging, aging-associated pathologies, and decreased survival. We showed that Fus1KO mice develop multiple early aging signs including lordokyphosis, lack of vigor, inability to accumulate fat, reduced ability to tolerate stress, and premature death. Other prominent pathological changes included low sperm counts, compromised ability of adult stem cells to repopulate tissues, and chronic inflammation. At the molecular level, we demonstrated that mitochondria of Fus1 KO cells have low reserve respiratory capacity (the ability to produce extra energy during sudden energy demanding situations), and show significantly altered dynamics of cellular calcium response.Our recent studies on early hearing and memory loss in Fus1 KO mice combined with the new data presented here suggest that calcium and energy homeostasis controlled by Fus1 may be at the core of its aging-regulating activities. Thus, Fus1 protein and Fus1-dependent pathways and processes may represent new tools and targets for anti-aging strategies.