Two-photon microscopy of the mouse cochleain situfor cellular diagnosis

Abstract

Abstract. Sensorineural hearing loss is the most common type of hearing loss worldwide, yet the underlying cause is typically unknown because the inner ear cannot be biopsied today without destroying hearing, and intracochlear cells have not been imaged with resolution sufficient to establish diagnosis. Intracochlear imaging has been technologically challenging because of the cochlea's small size and encasement in bone. We report, for the first time, imaging of the mouse cochlea in situ without exogenous dyes, through a membranous round window, using a near-infrared femtosecond laser as the excitation and endogenous two-photon excitation fluorescence (TPEF) and second harmonic generation as the contrast mechanisms. We find that TPEF exhibits strong contrast allowing cellular, and even subcellular resolution, and detection of specific, noise-induced pathologic changes. Our results demonstrate that the round window provides a useful access to the cochlea through the middle ear, and they motivate future development of a new and efficient diagnostic tool based on two-photon micro-endoscopy. © The Authors.Published by SPIE under a Creative Commons Attribution 3.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.

Human audiometric thresholds do not predict specific cellular damage in the inner ear

“…Although the cochlea’s small size and bony capsule make intracochlear interrogation challenging, better non- or minimally-invasive tools to diagnose cellular damage in the inner ear are needed. Promising new technologies soon expected to enter clinical applications include confocal fluorescence microscopy, optical coherence tomography, and in particular two-photon micro-endoscopy, which has been shown in animals to enable the detection of cell-specific damage in situ (Chen et al, 2007; MacDonald et al, 2008; Wong et al, 2000; Yang et al, 2013). Our results suggest that exclusive reliance on audiometric thresholds or word recognition scores in clinical trials of novel therapies for deafness, such as gene therapy, may prematurely disqualify a promising therapy when the current methods for detecting a potentially significant effect are not adequately sensitive.…”

Quantitative polarized light microscopy of unstained mammalian cochlear sections

“…Our results motivate future application of qPLM to characterize various murine models of human hearing loss, based on specific changes in the birefringent properties of cochlear structures. In light of our recent study demonstrating that the optical properties of fixed cochlear tissues are similar to those of unfixed specimens, 38 qPLM data from fixed specimens may ultimately be applied in humans in vivo to guide diagnosis and management through the use of new noninvasive imaging modalities.…”

Quantitative polarized light microscopy of human cochlear sections

“…Establishing the polarization dependent optical properties of normal healthy cochlear tissue is the first step in considering qPLM as a diagnostic tool in vivo. Although the histological sections used in this study were fixed, they are likely representative of unfixed tissue, based on our comparison of polarization dependent optical properties of fixed and unfixed cochlear tissues using two photon microscopy [3]. However, we recognize that fixation can change retardance, scattering and depolarization of polarized light [19], which strongly motivates future studies of unfixed cochlear tissues.…”

“…Today, the only source of information about the cellular basis of human deafness is cadaveric human temporal bones that house the inner ear. To enable future cellular-level intracochlear imaging in alive humans, we have been exploring optical imaging tools [3,4] because optics provides higher spatial resolution at a lower cost than techniques based on ionizing radiation or magnetic resonance.…”