New Automatic and Robust Measures to Evaluate Hearing Loss and Tinnitus in Preclinical Models

  • A. Laboulais
  • S. Malmström
  • C. Dejean
  • M. Cardoso
  • T. Le Meur
  • L. Almeida
  • C. Goze-Bac
  • S. PucheuEmail author


During this collaboration between CILcare and KeenEye Technologies, a full pipeline has been designed to automatically classify and quantify the number of hair cells in 3D cochlea images. This project introduced many challenges with regard to specific pre- and post-data processing and an adaptive model for 3D object detection. The model has been trained using transfer learning with mini batch images keeping the context information around the different types of cells. This new automatic counting method performed 10 times faster than humans, with on average 3.5 min to analyze one fragment image. The algorithm gave performance metrics of 90% for precision and 70% for sensitivity. While the precision value is good, additional work is needed to increase the overall sensitivity and reduce its variance. In addition, an objective quantification method to detect tinnitus on rats was developed in collaboration between CILcare and Charles Coulomb Laboratory (L2C-BioNanoNMRI team). Tinnitus, a phantom auditory sensation, which occurs in the absence of an external sound stimulus, is generated presumably within the auditory brain. Here we focus on the inferior colliculus (IC), a midbrain structure that integrates auditory information from both ears as well as information from other sensory systems. Some studies reveal neural hyperactivity in the IC after salicylate drug administration. In this study, we present an innovative manganese-enhanced magnetic resonance imaging (MEMRI) analysis method called ∆R2/R2. This quantitative method detects 1H NMR relaxation rate changes in the absence or presence of tinnitus. The ∆R2/R2 method generates relevant data comparable to those obtained with the signal-to-noise ratio (SNR) and signal intensity ratio (SIR) methods when manganese is administered by the transtympanic or intraperitoneal route. A major advantage of the ∆R2/R2 method is that it is automatic, robust, and reveals quantitative markers compared to qualitative methods like SNR and SIR.


Cochleogram Automated method Rat Hearing loss Tinnitus MEMRI 



Broadband noise


Blood oxygen level dependent


Cerebral cortex


US Food and Drug Administration


Functional magnetic resonance imaging


GAP inhibition of the acoustic startle reflex


Hair cells


Inferior colliculus


Inner hair cells




Manganese enhancement magnetic resonance magnetic


Manganese chloride


Nuclear magnetic resonance


Outer hair cells


Positron emission tomography


NMR relaxation rate


Regional cerebral blood flow


Red green blue




Region of interest


Supporting cells


Standard error of the mean


Spiral ganglion neurons


Signal intensity ratio


Salicylate-induced tinnitus


Signal-to-noise ratio


Transversal relaxation time




  1. 1.
    Cunningham LL, Tucci DL (2017) Hearing loss in adults. N Engl J Med 377:2465–2473CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Wilson BS, Tucci DL, Merson MH, O’Donoghue GM (2017) Global hearing health care: new findings and perspectives. Lancet 390:2503–2515CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Sultana H, Mumtaz N, Dawood T (2018) Type and degree of hearing loss in patients with tinnitus. International Journal of Rehabilitation Sciences (IJRS) 7(01):24–27Google Scholar
  4. 4.
    Basile CE, Fournier P, Hutchins S, Hebert S (2013) Psychoacoustic assessment to improve tinnitus diagnosis. PLoS One 8:e82995CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Stolzberg D, Salvi RJ, Allman BL (2012) Salicylate toxicity model of tinnitus. Front Syst Neurosci 6:28CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Brozoski TJ, Ciobanu L, Bauer CA (2007) Central neural activity in rats with tinnitus evaluated with manganese-enhanced magnetic resonance imaging (MEMRI). Hear Res 228:168–179CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Holt AG, Bissig D, Mirza N, Rajah G, Berkowitz B (2010) Evidence of key tinnitus-related brain regions documented by a unique combination of manganese-enhanced MRI and acoustic startle reflex testing. PLoS One 5:e14260CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Yu X, Wadghiri YZ, Sanes DH, Turnbull DH (2005) In vivo auditory brain mapping in mice with Mn-enhanced MRI. Nat Neurosci 8:961–968CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Chen YC, Li X, Liu L, Wang J, Lu CQ, Yang M, Jiao Y, Zang FC, Radziwon K, Chen GD et al (2015) Tinnitus and hyperacusis involve hyperactivity and enhanced connectivity in auditory-limbic-arousal-cerebellar network. elife 4:e06576CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Harding GW, Bohne BA (2009) Relation of focal hair-cell lesions to noise-exposure parameters from a 4- or a 0.5-kHz octave band of noise. Hear Res 254:54–63CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Harding GW, Bohne BA, Ahmad M (2002) DPOAE level shifts and ABR threshold shifts compared to detailed analysis of histopathological damage from noise. Hear Res 174:158–171CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Gauvin DV, Yoder J, Koch A, Zimmermann ZJ, Tapp RL (2017) Down for the count: the critical endpoint in ototoxicity remains the cytocochleogram. J Pharmacol Toxicol Methods 88:123–129CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Viberg A, Canlon B (2004) The guide to plotting a cochleogram. Hear Res 197:1–10CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Neal C, Kennon-McGill S, Freemyer A, Shum A, Staecker H, Durham D (2015) Hair cell counts in a rat model of sound damage: effects of tissue preparation & identification of regions of hair cell loss. Hear Res 328:120–132CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Bohne BA (1972) Location of small cochlear lesions by phase contrast microscopy prior to thin sectioning. Laryngoscope 82:1–16CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Hardie NA, MacDonald G, Rubel EW (2004) A new method for imaging and 3D reconstruction of mammalian cochlea by fluorescent confocal microscopy. Brain Res 1000:200–210CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Bohne BA, Harding GW (2011) Microscopic anatomy of the mouse inner ear, laboratory manual, 3rd edition. Washington University Press, pp 1–2, St. Louis, MO, USAGoogle Scholar
  18. 18.
    Ehret G, Frankenreiter M (1977) Quantitative analysis of cochlear structures in the house mouse in relation to mechanisms of acoustical information processing. J Comp Physiol 122:65–85Google Scholar
  19. 19.
    Keithley EM, Feldman ML (1982) Hair cell counts in an age-graded series of rat cochleas. Hear Res 8:249–262CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Spoendlin H, Brun JP (1974) The block-surface technique for evaluation of cochlear pathology. Arch Otorhinolaryngol 208:137–145CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Akil O, Seal RP, Burke K, Wang C, Alemi A, During M, Edwards RH, Lustig LR (2012) Restoration of hearing in the VGLUT3 knockout mouse using virally mediated gene therapy. Neuron 75:283–293CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Bohne BA, Kimlinger M, Harding GW (2017) Time course of organ of Corti degeneration after noise exposure. Hear Res 344:158–169CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Chen GD, Decker B, Krishnan Muthaiah VP, Sheppard A, Salvi R (2014) Prolonged noise exposure-induced auditory threshold shifts in rats. Hear Res 317(1–8):1CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Takada Y, Takada T, Lee MY, Swiderski DL, Kabara LL, Dolan DF, Raphael Y (2015) Ototoxicity-induced loss of hearing and inner hair cells is attenuated by HSP70 gene transfer. Mol Ther Methods Clin Dev 2:15019CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Akil O, Lustig LR (2013) Mouse cochlear whole mount immunofluorescence. Bio Protoc 3:e332CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Kaiming H, Xiangyu Z, Shaoqing R, Jian S (2015). Deep Residual Learning for Image Recognition. 2016 IEEE Conference on Computer Vision and Pattern Recognition (CVPR): 770–778Google Scholar
  27. 27.
    Sinno P, Qiang, Y (2010). A Survey on Transfer Learning. Knowledge and Data Engineering, IEEE Transactions on. 22. 1345 - 1359. doi: 10.1109/TKDE.2009.191Google Scholar
  28. 28.
    Monro HRS (1951) A stochastic approximation method. Ann Math Stat 22:400–407Google Scholar
  29. 29.
    Ding D, Jiang H, Chen GD, Longo-Guess C, Muthaiah VP, Tian C, Sheppard A, Salvi R, Johnson KR (2016) N-acetyl-cysteine prevents age-related hearing loss and the progressive loss of inner hair cells in gamma-glutamyl transferase 1 deficient mice. Aging (Albany NY) 8:730–750CrossRefGoogle Scholar
  30. 30.
    Sanz L, Murillo-Cuesta S, Cobo P, Cediel-Algovia R, Contreras J, Rivera T, Varela-Nieto I, Avendano C (2015) Swept-sine noise-induced damage as a hearing loss model for preclinical assays. Front Aging Neurosci 7:7PubMedPubMedCentralGoogle Scholar
  31. 31.
    Lockwood AH, Salvi RJ, Burkard RF (2002) Tinnitus. N Engl J Med 347:904–910CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Jastreboff PJ, Brennan JF, Coleman JK, Sasaki CT (1988) Phantom auditory sensation in rats: an animal model for tinnitus. Behav Neurosci 102:811–822CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Boyen K, Baskent D, van Dijk P (2015) The gap detection test: can it be used to diagnose tinnitus? Ear Hear 36:e138–e145CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Turner JG, Brozoski TJ, Bauer CA, Parrish JL, Myers K, Hughes LF, Caspary DM (2006) Gap detection deficits in rats with tinnitus: a potential novel screening tool. Behav Neurosci 120:188–195CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Muca A, Standafer E, Apawu AK, Ahmad F, Ghoddoussi F, Hali M, Warila J, Berkowitz BA, Holt AG (2018) Tinnitus and temporary hearing loss result in differential noise-induced spatial reorganization of brain activity. Brain Struct Funct 223:2343–2360CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Galazyuk A, Hebert S (2015) Gap-prepulse inhibition of the acoustic startle reflex (GPIAS) for tinnitus assessment: current status and future directions. Front Neurol 6:88CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Lee JW, Park JA, Lee JJ, Bae SJ, Lee SH, Jung JC, Kim MN, Lee J, Woo S, Chang Y (2007) Manganese-enhanced auditory tract-tracing MRI with cochlear injection. Magn Reson Imaging 25:652–656CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Pautler RG (2004) In vivo, trans-synaptic tract-tracing utilizing manganese-enhanced magnetic resonance imaging (MEMRI). NMR Biomed 17:595–601CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Cheung SW, Nagarajan SS, Bedenbaugh PH, Schreiner CE, Wang X, Wong A (2001) Auditory cortical neuron response differences under isoflurane versus pentobarbital anesthesia. Hear Res 156:115–127CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Malheiros JM, Paiva FF, Longo BM, Hamani C, Covolan L (2015) Manganese-enhanced MRI: biological applications in neuroscience. Front Neurol 6:161CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Schumacher JW, Schneider DM, Woolley SM (2011) Anesthetic state modulates excitability but not spectral tuning or neural discrimination in single auditory midbrain neurons. J Neurophysiol 106:500–514CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Ter-Mikaelian M, Sanes DH, Semple MN (2007) Transformation of temporal properties between auditory midbrain and cortex in the awake Mongolian gerbil. J Neurosci 27:6091–6102CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Jin SU, Lee JJ, Hong KS, Han M, Park JW, Lee HJ, Lee S, Lee KY, Shin KM, Cho JH et al (2013) Intratympanic manganese administration revealed sound intensity and frequency dependent functional activity in rat auditory pathway. Magn Reson Imaging 31:1143–1149CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Lee HJ, Yoo SJ, Lee S, Song HJ, Huh MI, Jin SU, Lee KY, Lee J, Cho JH, Chang Y (2012) Functional activity mapping of rat auditory pathway after intratympanic manganese administration. Neuroimage 60:1046–1054CrossRefPubMedPubMedCentralGoogle Scholar
  45. 45.
    Paul AK, Lobarinas E, Simmons R, Wack D, Luisi JC, Spernyak J, Mazurchuk R, Abdel-Nabi H, Salvi R (2009) Metabolic imaging of rat brain during pharmacologically-induced tinnitus. Neuroimage 44:312–318CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Rancz EA, Moya J, Drawitsch F, Brichta AM, Canals S, Margrie TW (2015) Widespread vestibular activation of the rodent cortex. J Neurosci 35:5926–5934CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Silva AC, Bock NA (2008) Manganese-enhanced MRI: an exceptional tool in translational neuroimaging. Schizophr Bull 34:595–604CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Groschel M, Gotze R, Muller S, Ernst A, Basta D (2016) Central nervous activity upon systemic salicylate application in animals with kanamycin-induced hearing loss—a manganese-enhanced MRI (MEMRI) study. PLoS One 11:e0153386CrossRefPubMedPubMedCentralGoogle Scholar
  49. 49.
    Jung DJ, Han M, Jin SU, Lee SH, Park I, Cho HJ, Kwon TJ, Lee HJ, Cho JH, Lee KY et al (2014) Functional mapping of the auditory tract in rodent tinnitus model using manganese-enhanced magnetic resonance imaging. Neuroimage 100:642–649CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Liu Y, Li X, Ma C, Liu J, Lu H (2005) Salicylate blocks L-type calcium channels in rat inferior colliculus neurons. Hear Res 205:271–276CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    Hu SS, Mei L, Chen JY, Huang ZW, Wu H (2014) Expression of immediate-early genes in the inferior colliculus and auditory cortex in salicylate-induced tinnitus in rat. Eur J Histochem 58:2294CrossRefPubMedPubMedCentralGoogle Scholar
  52. 52.
    Spivak M, Weston J, Bottou L, Kall L, Noble WS (2009) Improvements to the percolator algorithm for peptide identification from shotgun proteomics data sets. J Proteome Res 8:3737–3745CrossRefPubMedPubMedCentralGoogle Scholar
  53. 53.
    Bauer CA, Brozoski TJ, Rojas R, Boley J, Wyder M (1999) Behavioral model of chronic tinnitus in rats. Otolaryngol Head Neck Surg 121:457–462CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Caravan P, Farrar CT, Frullano L, Uppal R (2009) Influence of molecular parameters and increasing magnetic field strength on relaxivity of gadolinium- and manganese-based T1 contrast agents. Contrast Media Mol Imaging 4:89–100CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2020

Authors and Affiliations

  • A. Laboulais
    • 1
    • 2
  • S. Malmström
    • 2
  • C. Dejean
    • 2
  • M. Cardoso
    • 1
  • T. Le Meur
    • 3
  • L. Almeida
    • 3
  • C. Goze-Bac
    • 1
  • S. Pucheu
    • 2
    Email author
  1. 1.Charles Coulomb Laboratory (L2C-BioNanoNMRI Team)UMR 5221 Centre National de la Recherche Scientifique—UniversityMontpellierFrance
  2. 2.CILcare, Advanced Solution for Drug Development in Hearing DisorderMontpellierFrance
  3. 3.KeeneyeParisFrance

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