Phytosterols reverse antiretroviral-induced hearing loss, with potential implications for cochlear aging.

Cholesterol contributes to neuronal membrane integrity, supports membrane protein clustering and function, and facilitates proper signal transduction. Extensive evidence has shown that cholesterol imbalances in the central nervous system occur in aging and in the development of neurodegenerative diseases. In this work, we characterize cholesterol homeostasis in the inner ear of young and aged mice as a new unexplored possibility for the prevention and treatment of hearing loss. Our results show that cholesterol levels in the inner ear are reduced during aging, an effect that is associated with an increased expression of the cholesterol 24-hydroxylase (CYP46A1), the main enzyme responsible for cholesterol turnover in the brain. In addition, we show that pharmacological activation of CYP46A1 with the antiretroviral drug efavirenz reduces the cholesterol content in outer hair cells (OHCs), leading to a decrease in prestin immunolabeling and resulting in an increase in the distortion product otoacoustic emissions (DPOAEs) thresholds. Moreover, dietary supplementation with phytosterols, plant sterols with structure and function similar to cholesterol, was able to rescue the effect of efavirenz administration on the auditory function. Altogether, our findings point towards the importance of cholesterol homeostasis in the inner ear as an innovative therapeutic strategy in preventing and/or delaying hearing loss.

Sensitivity to Envelope Interaural Time Differences: Modeling Auditory Modulation Filtering.

For amplitude-modulated sound, the envelope interaural time difference (ITDENV) is a potential cue for sound-source location. ITDENV is encoded in the lateral superior olive (LSO) of the auditory brainstem, by excitatory-inhibitory (EI) neurons receiving ipsilateral excitation and contralateral inhibition. Between human listeners, sensitivity to ITDENV varies considerably, but ultimately decreases with increasing stimulus carrier frequency, and decreases more strongly with increasing modulation rate. Mechanisms underlying the variation in behavioral sensitivity remain unclear. Here, with increasing carrier frequency (4-10 kHz), as we phenomenologically model the associated decrease in ITDENV sensitivity using arbitrarily fewer neurons consistent across populations, we computationally model the variable sensitivity across human listeners and modulation rates (32-800 Hz) as the decreasing range of membrane frequency responses in LSO neurons. Transposed tones stimulate a bilateral auditory-periphery model, driving model EI neurons where electrical membrane impedance filters the frequency content of inputs driven by amplitude-modulated sound, evoking modulation filtering. Calculated from Fisher information in spike-rate functions of ITDENV, for model EI neuronal populations distinctly reflecting the LSO range in membrane frequency responses, just-noticeable differences in ITDENV collectively reproduce the largest variation in ITDENV sensitivity across human listeners. These slow to fast model populations each generally match the best human ITDENV sensitivity at a progressively higher modulation rate, by membrane-filtering and spike-generation properties producing realistically less than Poisson variance. Non-resonant model EI neurons are also sensitive to interaural intensity differences. With peripheral filters centered between carrier frequency and modulation sideband, fast resonant model EI neurons extend ITDENV sensitivity above 500-Hz modulation.

Subthreshold resonance properties contribute to the efficient coding of auditory spatial cues.

Neurons in the medial superior olive (MSO) and lateral superior olive (LSO) of the auditory brainstem code for sound-source location in the horizontal plane, extracting interaural time differences (ITDs) from the stimulus fine structure and interaural level differences (ILDs) from the stimulus envelope. Here, we demonstrate a postsynaptic gradient in temporal processing properties across the presumed tonotopic axis; neurons in the MSO and the low-frequency limb of the LSO exhibit fast intrinsic electrical resonances and low input impedances, consistent with their processing of ITDs in the temporal fine structure. Neurons in the high-frequency limb of the LSO show low-pass electrical properties, indicating they are better suited to extracting information from the slower, modulated envelopes of sounds. Using a modeling approach, we assess ITD and ILD sensitivity of the neural filters to natural sounds, demonstrating that the transformation in temporal processing along the tonotopic axis contributes to efficient extraction of auditory spatial cues.

Constitutive expression of the alpha10 nicotinic acetylcholine receptor subunit fails to maintain cholinergic responses in inner hair cells after the onset of hearing.

Efferent inhibition of cochlear hair cells is mediated by alpha9alpha10 nicotinic cholinergic receptors (nAChRs) functionally coupled to calcium-activated, small conductance (SK2) potassium channels. Before the onset of hearing, efferent fibers transiently make functional cholinergic synapses with inner hair cells (IHCs). The retraction of these fibers after the onset of hearing correlates with the cessation of transcription of the Chrna10 (but not the Chrna9) gene in IHCs. To further analyze this developmental change, we generated a transgenic mice whose IHCs constitutively express alpha10 into adulthood by expressing the alpha10 cDNA under the control of the Pou4f3 gene promoter. In situ hybridization showed that the alpha10 mRNA is expressed in IHCs of 8-week-old transgenic mice, but not in wild-type mice. Moreover, this mRNA is translated into a functional protein, since IHCs from P8-P10 alpha10 transgenic mice backcrossed to a Chrna10(-/-) background (whose IHCs have no cholinergic function) displayed normal synaptic and acetylcholine (ACh)-evoked currents in patch-clamp recordings. Thus, the alpha10 transgene restored nAChR function. However, in the alpha10 transgenic mice, no synaptic or ACh-evoked currents were observed in P16-18 IHCs, indicating developmental down-regulation of functional nAChRs after the onset of hearing, as normally observed in wild-type mice. The lack of functional ACh currents correlated with the lack of SK2 currents. These results indicate that multiple features of the efferent postsynaptic complex to IHCs, in addition to the nAChR subunits, are down-regulated in synchrony after the onset of hearing, leading to lack of responses to ACh.

A point mutation in the hair cell nicotinic cholinergic receptor prolongs cochlear inhibition and enhances noise protection.

The transduction of sound in the auditory periphery, the cochlea, is inhibited by efferent cholinergic neurons projecting from the brainstem and synapsing directly on mechanosensory hair cells. One fundamental question in auditory neuroscience is what role(s) this feedback plays in our ability to hear. In the present study, we have engineered a genetically modified mouse model in which the magnitude and duration of efferent cholinergic effects are increased, and we assess the consequences of this manipulation on cochlear function. We generated the Chrna9L9'T line of knockin mice with a threonine for leucine change (L9'T) at position 9' of the second transmembrane domain of the alpha9 nicotinic cholinergic subunit, rendering alpha9-containing receptors that were hypersensitive to acetylcholine and had slower desensitization kinetics. The Chrna9L9'T allele produced a 3-fold prolongation of efferent synaptic currents in vitro. In vivo, Chrna9L9'T mice had baseline elevation of cochlear thresholds and efferent-mediated inhibition of cochlear responses was dramatically enhanced and lengthened: both effects were reversed by strychnine blockade of the alpha9alpha10 hair cell nicotinic receptor. Importantly, relative to their wild-type littermates, Chrna9(L9'T/L9'T) mice showed less permanent hearing loss following exposure to intense noise. Thus, a point mutation designed to alter alpha9alpha10 receptor gating has provided an animal model in which not only is efferent inhibition more powerful, but also one in which sound-induced hearing loss can be restrained, indicating the ability of efferent feedback to ameliorate sound trauma.

Effects of quinine, quinidine, and chloroquine on alpha9alpha10 nicotinic cholinergic receptors.

In this study, we report the effects of the quinoline derivatives quinine, its optical isomer quinidine, and chloroquine on alpha9alpha10-containing nicotinic acetylcholine receptors (nAChRs). The compounds blocked acetylcholine (ACh)-evoked responses in alpha9alpha10-injected Xenopus laevis oocytes in a concentration-dependent manner, with a rank order of potency of chloroquine (IC50 = 0.39 microM) > quinine (IC50 = 0.97 microM) approximately quinidine (IC50= 1.37 microM). Moreover, chloroquine blocked ACh-evoked responses on rat cochlear inner hair cells with an IC50 value of 0.13 microM, which is within the same range as that observed for recombinant receptors. Block by chloroquine was purely competitive, whereas quinine inhibited ACh currents in a mixed competitive and noncompetitive manner. The competitive nature of the blockage produced by the three compounds was confirmed by equilibrium binding experiments using [3H]methyllycaconitine. Binding affinities (Ki values) were 2.3, 5.5, and 13.0 microM for chloroquine, quinine, and quinidine, respectively. Block by quinine was found to be only slightly voltage-dependent, thus precluding open-channel block as the main mechanism of interaction of quinine with alpha9alpha10 nAChRs. The present results add to the pharmacological characterization of alpha9alpha10-containing nicotinic receptors and indicate that the efferent olivocochlear system that innervates the cochlear hair cells is a target of these ototoxic antimalarial compounds.