Perilymphatic Gentamicin Trafficking through Basolateral Channels May Underlie Enhanced Ototoxicity by Noise‐Induced Threshold Shifts. (2nd September 2014) - Luk, Lauren J.
Abstract : Objectives: (1) Demonstrate gentamicin uptake from perilymph into hair cells via basolateral channels such as the postsynaptic acetylcholine receptor (nAChR) and the TRPA1 in neonatal mice in vivo and in vitro. (2) Recognize alternative gentamicin trafficking pathways to the well established endolymphatic mechanoelectrotransducer channel underlying aminoglycoside ototoxicity. Methods: Neonatal mice were intraperitoneally injected with fluorescently‐conjugated Gentamicin‐Texas Red (GTTR), prior to fixation 30 minutes later. Cochlear explants were treated with GTTR 1 µg/mL, with or without 100 µM acetylcholine or cinnamaldehyde then fixed, and analyzed. Fluorescence intensities were measured by calculating the mean gray value of each cell with Image J. Student t test was used to determine any significant differences in fluorescence values between treatment groups. Results: Application of acetylcholine increased GTTR uptake by wildtype outer hair cells (OHC) in vivo and in vitro. These results were also replicated in Myo7a8J/8J mice, which lack functional mechanoelectrotransducer channels. Application of cinnamaldehyde (TRPA1 agonist) also increased GTTR uptake in OHCs in wildtype, myo7a8J/8J mice and guinea pigs. Conclusions: Endolymphatic gentamicin trafficking via the mechanoelectrotransducer (MET) channel is a well established mechanism of ototoxicity. Noise‐induced threshold shifts (NITS) have been shown to damage the mechanically gated tip link, thus closing MET channel. However, aminoglycoside hair cell uptake is enhanced by NITS. This suggests one or more aminoglycoside entry route(s) into hair cells in addition to, and independent of, the MET channel. The results of this study support perilymphatic trafficking of gentamicin via basolateral channels including the nAChR Ca2+ channel, and TRPA1.