Acoustic Trauma Increases Cochlear and Hair Cell Uptake of Gentamicin

Exposure to intense sound or high doses of aminoglycoside antibiotics can increase hearing thresholds, induce cochlear dysfunction, disrupt hair cell morphology and promote hair cell death, leading to permanent hearing loss. When the two insults are combined, synergistic ototoxicity occurs, exacerbating cochlear vulnerability to sound exposure. The underlying mechanism of this synergism remains unknown. In this study, we tested the hypothesis that sound exposure enhances the intra-cochlear trafficking of aminoglycosides, such as gentamicin, leading to increased hair cell uptake of aminoglycosides and subsequent ototoxicity.Juvenile C57Bl/6 mice were exposed to moderate or intense sound levels, while fluorescently-conjugated or native gentamicin was administered concurrently or following sound exposure. Drug uptake was then examined in cochlear tissues by confocal microscopy.Prolonged sound exposure that induced temporary threshold shifts increased gentamicin uptake by cochlear hair cells, and increased gentamicin permeation across the strial blood-labyrinth barrier. Enhanced intra-cochlear trafficking and hair cell uptake of gentamicin also occurred when prolonged sound, and subsequent aminoglycoside exposure were temporally separated, confirming previous observations. Acute, concurrent sound exposure did not increase cochlear uptake of aminoglycosides.Prolonged, moderate sound exposures enhanced intra-cochlear aminoglycoside trafficking into the stria vascularis and hair cells. Changes in strial and/or hair cell physiology and integrity due to acoustic overstimulation could increase hair cell uptake of gentamicin, and may represent one mechanism of synergistic ototoxicity

Cell Type-Specific Reduction of Beta Tubulin Isotypes Synthesized in the Developing Gerbil Organ of Corti

There are seven isotypic forms of the microtubule protein beta tubulin in mammals, but not all isotypes are synthesized in every cell type. In the adult organ of Corti, each of the five major cell types synthesizes a different subset of isotypes. Inner hair cells synthesize only betaI and betaII tubulin, while outer hair cells make betaI and betaIV tubulin. Only betaII and betaIV tubulin are found in inner and outer pillar cells, while betaI, betaII, and betaIV tubulin are present in Deiters cells, and betaI, betaII and betaIII tubulin are found in organ of Corti dendrites. During post-natal organ of Corti development in the gerbil, microtubules are elaborated in an orderly temporal sequence beginning with hair cells, followed by pillar cells and Deiters cells. Using beta tubulin isotype-specific antibodies, we show that, in the gerbil cochlea, the same three isotypes are present in each cell type at birth, and that a cell type-specific reduction in the isotypes synthesized occurs in hair cells and pillar cells at an unusually late stage in development. No beta tubulin isotypes were detected in mature afferent dendrites, but we show that this is because few microtubules are present in mature dendrites. In addition, we show that primary cilia in inner hair cells, a feature of early development, persist much later than previously reported. The findings represent the first description of developmental cell type-specific reductions in tubulin isotypes in any system

Local mechanisms for loud sound-enhanced aminoglycoside entry into outer hair cells.

Loud sound exposure exacerbates aminoglycoside ototoxicity, increasing the risk of permanent hearing loss and degrading the quality of life in affected individuals. We previously reported that loud sound exposure induces temporary threshold shifts (TTS) and enhances uptake of aminoglycosides, like gentamicin, by cochlear outer hair cells (OHCs). Here, we explore mechanisms by which loud sound exposure and TTS could increase aminoglycoside uptake by OHCs that may underlie this form of ototoxic synergy. Mice were exposed to loud sound levels to induce TTS, and received fluorescently-tagged gentamicin (GTTR) for 30 min prior to fixation. The degree of TTS was assessed by comparing auditory brainstem responses (ABRs) before and after loud sound exposure. The number of tip links, which gate the GTTR-permeant mechanoelectrical transducer (MET) channels, was determined in OHC bundles, with or without exposure to loud sound, using scanning electron microscopy. We found wide-band noise (WBN) levels that induce TTS also enhance OHC uptake of GTTR compared to OHCs in control cochleae. In cochlear regions with TTS, the increase in OHC uptake of GTTR was significantly greater than in adjacent pillar cells. In control mice, we identified stereociliary tip links at ~50% of potential positions in OHC bundles. However, the number of OHC tip links was significantly reduced in mice that received WBN at levels capable of inducing TTS. These data suggest that GTTR uptake by OHCs during TTS occurs by increased permeation of surviving, mechanically-gated MET channels, and/or non-MET aminoglycoside-permeant channels activated following loud sound exposure. Loss of tip links would hyperpolarize hair cells and potentially increase drug uptake via aminoglycoside-permeant channels expressed by hair cells. The effect of TTS on aminoglycoside-permeant channel kinetics will shed new light on the mechanisms of loud sound-enhanced aminoglycoside uptake, and consequently on ototoxic synergy