Functional Hair Cell Mechanotransducer Channels Are Required for Aminoglycoside Ototoxicity

Abstract

Aminoglycosides (AG) are commonly prescribed antibiotics with potent bactericidal activities. One main side effect is permanent sensorineural hearing loss, induced by selective inner ear sensory hair cell death. Much work has focused on AG's initiating cell death processes, however, fewer studies exist defining mechanisms of AG uptake by hair cells. The current study investigated two proposed mechanisms of AG transport in mammalian hair cells: mechanotransducer (MET) channels and endocytosis. To study these two mechanisms, rat cochlear explants were cultured as whole organs in gentamicin-containing media. Two-photon imaging of Texas Red conjugated gentamicin (GTTR) uptake into live hair cells was rapid and selective. Hypocalcemia, which increases the open probability of MET channels, increased AG entry into hair cells. Three blockers of MET channels (curare, quinine, and amiloride) significantly reduced GTTR uptake, whereas the endocytosis inhibitor concanavalin A did not. Dynosore quenched the fluorescence of GTTR and could not be tested. Pharmacologic blockade of MET channels with curare or quinine, but not concanavalin A or dynosore, prevented hair cell loss when challenged with gentamicin for up to 96 hours. Taken together, data indicate that the patency of MET channels mediated AG entry into hair cells and its toxicity. Results suggest that limiting permeation of AGs through MET channel or preventing their entry into endolymph are potential therapeutic targets for preventing hair cell death and hearing loss.

Validation of a Strategy for Cancer Therapy: Delivering Aminoglycoside Drugs to Mitochondria in HeLa Cells

“…In previous study, it has been reported that AGs have a high affinity for ribosomal RNA and permeability transition in mitochondria, resulting in apoptosis via the inhibition of mitochondrial transcription or exaggerated oxidative stress, which are consistent with the molecular mechanisms for the ototoxicity and nephrotoxicity induced by AGs [24][25][26] . AGs have been clinically available as antibiotics for many years, and, unfortunately, can exert some irreversible adverse-effects, especially in the patients with a mitochondrial disease.…”

Designer aminoglycosides prevent cochlear hair cell loss and hearing loss

“…Although the molecular nature of this channel remains controversial, a significant body of literature exists on the biophysical properties (24,(26)(27)(28)(29)(30). Given that aminoglycosides are permeable blockers of the MET channel, we hypothesized that targeted modification to reduce positively charged sites on the aminoglycoside molecule would hinder drug entry into hair cells and thus prevent ototoxicity (9). Using established synthetic methods (31), we first chemically protected 3 of the amine groups on the aminoglycoside sisomicin, a biosynthetic precursor of gentamicin, thus allowing subsequent modification of the N1 and/or N3′′ amine site (Supplemental Figure 1, A and B; supplemental material available online with this article; doi:10.1172/JCI77424DS1).…”

“…Using established synthetic methods (31), we first chemically protected 3 of the amine groups on the aminoglycoside sisomicin, a biosynthetic precursor of gentamicin, thus allowing subsequent modification of the N1 and/or N3′′ amine site (Supplemental Figure 1, A and B; supplemental material available online with this article; doi:10.1172/JCI77424DS1). Sisomicin was selected because it is hair cell MET channel is required for drug toxicity (9,(20)(21)(22)(23), we have redesigned and modified the aminoglycoside sisomicin based on the biophysical properties of the hair cell MET channels (24) and aminoglycoside-bacterial ribosome interactions (25). Here, we have characterized 9 sisomicin derivatives with diminished ototoxicity in vitro, with 3 of the 9 compounds exhibiting comparable anti-E. coli activities to the parent compound sisomicin.…”

“…The drug is first transported across the blood-labyrinth barrier to enter the endolymph-filled inner ear compartments (8). It then traverses the mechanotransducer channels or other cation channels to enter hair cells (9)(10)(11), resulting in cell demise most likely as a result of the generation of reactive oxygen species (3,12). Past efforts to neutralize aminoglycoside ototoxicity have emphasized the administration of antioxidants, yet the efficacy of this approach is limited (4,(13)(14)(15), in part because of the long half-life of aminoglycosides once they enter hair cells (6).…”

Therapeutic and protective effects of autologous serum in amikacin-induced ototoxicity

“…When the formation of reactive oxygen species overwhelms the capacity of the intrinsic protective and repair systems, the cell undergoes apoptotic cell death, resulting in ototoxicity and deafness. 3,33,34 The present study evaluated the possible therapeutic and protective effects of autologous serum on amikacin-induced apoptosis in an animal model, by both Data represent mean apoptotic cell count, unless indicated otherwise. * P-values concern the differences between the study group and the positive control group.…”