Direct detection of ototoxicant-induced reactive oxygen species generation in cochlear explants

William J. Clerici, Kenneth Hensley, Debra L. DiMartino, D. Allan Butterfield

Research output: Contribution to journalArticlepeer-review

272 Scopus citations


The proposal that free-radical generation contributes to the ototoxicities of several chemical agents was studied utilizing electron paramagnetic resonance (EPR) spectrometry to detect directly ototoxicant-induced reactive oxygen species formation in cochlear tissue. Guinea pig cochlear explants in chelexed artificial perilymph (AP; 200 μl) were exposed to an ototoxicant or AP for 10 min. Ototoxic agents included gentamicin sulfate (4.0 mM), kanamycin monosulfate (4.0 mM), ethacrynic acid (0.5 mM), furosemide (0.3 mM), cisplatin (0.1 mM), trimethyltin chloride (0.1 mM), and quinine HCl (3.0 mM). Following incubation, 20 μl of AP/ototoxicant mixture was replaced by the filtered spin trap, 5,5-dimethylpyrroline-N-oxide (DMPO). After 10 min, the EPR spectrum of the mixture was obtained. Four line EPR spectra of relative intensities 1:2:2:1, associated with hydroxyl radical (OH.)/DMPO adduct formation, were evidenced by reaction mixtures containing cochlear explants exposed to each ototoxicant. Cisplatin, quinine and the loop diuretics produced weak OH.-associated EPR signals in the absence of a cochlear explant, which were amplified in its presence; Deferoxamine quenched all OH spectral peaks. Peroxide levels, assayed in parallel experiments, were diminished by each ototoxicant relative to those seen following AP exposure, suggesting possible H2O2 conversion to OH: These data support the proposal that various ototoxic agents are capable of reactive oxygen species generation or promotion in cochlear tissues.

Original languageEnglish
Pages (from-to)116-124
Number of pages9
JournalHearing Research
Issue number1-2
StatePublished - Sep 1 1996

Bibliographical note

Funding Information:
The authorst hankDr. Lihua Yang for technicacl ontri-butions. This researchw as funded by the Division of Otolaryngology-Heaadn d Neck Surgery, Departmenot f Surgery,U niversityo f KentuckyC ollegeof Medicinea nd by Grant DC06701 from the NationalI nstituteo f Environmental Health Sciences( NIEHS), NIH. Its contentsa re solely the responsibilitoyf the authorsa nddo not necessarily representth e official views of the NIEHS, NIH.


  • Cochlea
  • Electron paramagnetic resonance spectrometry
  • Free radical
  • Ototoxicity
  • Reactive oxygen species

ASJC Scopus subject areas

  • Sensory Systems


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