Auditory Sensory Organ Research Articles

Regulation of polarized extension and planar cell polarity in the cochlea by the vertebrate PCP pathway

The mammalian auditory sensory organ, the organ of Corti, consists of sensory hair cells with uniformly oriented stereocilia on the apical surfaces and has a distinct planar cell polarity (PCP) parallel to the sensory epithelium. It is not certain how this polarity is achieved during differentiation. Here we show that the organ of Corti is formed from a thicker and shorter postmitotic primordium through unidirectional extension, characteristic of cellular intercalation known as convergent extension. Mutations in the PCP pathway interfere with this extension, resulting a shorter and wider cochlea as well as misorientation of stereocilia. Furthermore, parallel to the homologous pathway in Drosophila melanogaster, a mammalian PCP component Dishevelled2 shows PCP-dependent polarized subcellular localization across the organ of Corti. Taken together, these data suggest that there is a conserved molecular mechanism for PCP pathways in invertebrates and vertebrates and indicate that the mammalian PCP pathway might directly couple cellular intercalations to PCP establishment in the cochlea.

Promyelocytic leukemia zinc finger protein localizes to the cochlear outer hair cells and interacts with prestin, the outer hair cell motor protein

Hair cells in the auditory sensory organ are specialized mechanoreceptors common to mammalian and non-mammalian species. The mammalian cochlear outer hair cells (OHC) possess a distinct motile property, dubbed membrane-based electromotility, that enhances the receptor function. This electromotility is believed to be the basis of cochlear amplification that increases sensitivity of the mammalian ear to sound. Prestin, a unique voltage-sensitive motor molecule localized in the lateral membrane of OHC, is presumably responsible for OHC electromotility. It has been documented that prestin null-animals lack electromotility and suffer from ∼50 dB loss of hearing sensitivity. To identify proteins that interact with prestin we carried out a yeast two-hybrid library screen using the C-terminal intracellular domain of prestin as bait. Seven bait-dependent prey clones were identified independently. Further analysis revealed that they encode partially over-lapping regions of a single protein: a transcriptional repressor, promyleocytic leukemia zinc finger protein (PLZF). PLZF encodes a POZ/domain Kruppel-type zinc finger transcription factor reported to have pro-apoptotic and anti-proliferative activity. The interaction between endogenous prestin and PLZF proteins in the cochlea was confirmed by co-immunoprecipitation using organ of Corti lysates. Furthermore, immunohistochemical studies strongly suggest that PLZF co-localizes with prestin near the lateral membrane of cochlear OHC. The physiological significance of this interaction remains to be explored.

Noradrenergic modulation of brainstem nuclei alters cochlear neural output

The peripheral auditory sense organ, the cochlea, receives innervation from lateral and medial olivocochlear neurons in the brainstem. These neurons are able to modulate cochlear neural output. Anatomical studies have shown that one of the neurotransmitters which is present in varicosities surrounding the olivocochlear neurons in the brainstem is noradrenaline and previous work on brainstem slices has demonstrated a generally excitatory effect of noradrenaline on medial olivocochlear neurons. In order to assess in vivo the function of the noradrenergic inputs to olivocochlear neurons, we injected noradrenaline in the brainstem of anaesthetised guinea pigs and recorded ipsilateral cochlear electrical activity. Injections of noradrenaline close to the lateral olivocochlear neurons evoked increases in the sound-driven neural activity from the cochlea, measured as compound action potential (CAP) amplitude, as well as in the spontaneous activity, measured as amplitude of the 900 Hz peak of the spectrum of the neural noise in the cochlear fluids. In contrast, noradrenaline in the vicinity of the medial olivocochlear neurons evoked inhibitory effects on both the CAP amplitude and 900 Hz peak. These results indicate most likely an excitatory action of noradrenaline on both the lateral and medial olivocochlear neurons in the brainstem, and show that such noradrenergic inputs can modulate cochlear function.

Expression of Islet1 marks the sensory and neuronal lineages in the mammalian inner ear.

Several basic helix-loop-helix (bHLH) genes have been shown to be essential for the generation of the auditory sensory hair cells or the spiral ganglion (SG) neurons that innervate the hair cells in the cochlea, as well as a variety of cell types in the other nervous systems. However, it remains elusive what cellular context-dependent mechanisms confer the inner ear-specific neuronal or sensory competency/identities. We explored the possibility that one of the mechanisms responsible for generating cellular diversity in the nervous system through cooperative action of bHLH and LIM-homeodomain (LIM-HD) transcriptional factors might also contribute to the inner ear-specific sensory and/or neuronal competency. Here, we show that Islet1 (Isl1), a LIM-HD protein, is expressed early in the otocyst in the region that gives rise to both the auditory sensory organ, the organ of Corti, and SG neurons. Subsequently, the expression of Isl1 is maintained in SG neurons but is transitory in the sensory lineage. At embryonic day 12 (E12) in mice, the expression of Isl1 marks distinctively the ventral portion of the nascent cochlear epithelium encompassing the primordial organ of Corti. At E13, Isl1 is maintained at relatively high levels in the sensory primordium while down-regulated in the other regions of the cochlear duct. As the sensory epithelium starts to differentiate, it is down-regulated in the entire cochlear epithelium. The expression of Isl1 in the developing inner ear reveals an early and likely a common step in the development of both sensory and neuronal lineages of the inner ear, and suggests its potential role in the inner ear-specific sensory and neuronal cell development.

Otx2, Gbx2, and Fgf8 expression patterns in the chick developing inner ear and their possible roles in otic specification and early innervation

The chick inner ear is a complex structure containing auditory and vestibular sensory organs innervated by neurons of the acoustic-vestibular ganglion. The molecular signals involved in the specification and initial innervation of the otic epithelium are poorly understood. Here, we present a detailed description of the Otx2, Gbx2, and Fgf8 gene expression patterns in the chick developing inner ear, comparing them with the Bmp4 expression, a putative sensory-organ marker. The Otx2 expression was detected in the ventro-lateral wall of the otic anlage and could play a role in the segregation of the saccule and utricle maculae. The relationship between Gbx2 and Fgf8 expression changed during inner ear development but was always related to the macula sacculi innervation and endolymphatic duct formation. Our results also suggest that the maculae of the saccule and lagena, and the medial portion of the macula utriculi could arise within a broad Fgf8-positive domain previously observed at the otocyst stage. The spatial and temporal relationships between these gene expression domains and the initial innervation of the epithelium by some subpopulations of otic axons suggest that expression domain boundaries could be involved in the specification and early innervation of presumptive sensory patches.

The structure and function of auditory chordotonal organs in insects

Insects are capable of detecting a broad range of acoustic signals transmitted through air, water, or solids. Auditory sensory organs are morphologically diverse with respect to their body location, accessory structures, and number of sensilla, but remarkably uniform in that most are innervated by chordotonal organs. Chordotonal organs are structurally complex Type I mechanoreceptors that are distributed throughout the insect body and function to detect a wide range of mechanical stimuli, from gross motor movements to air-borne sounds. At present, little is known about how chordotonal organs in general function to convert mechanical stimuli to nerve impulses, and our limited understanding of this process represents one of the major challenges to the study of insect auditory systems today. This report reviews the literature on chordotonal organs innervating insect ears, with the broad intention of uncovering some common structural specializations of peripheral auditory systems, and identifying new avenues for research. A general overview of chordotonal organ ultrastructure is presented, followed by a summary of the current theories on mechanical coupling and transduction in monodynal, mononematic, Type 1 scolopidia, which characteristically innervate insect ears. Auditory organs of different insect taxa are reviewed, focusing primarily on tympanal organs, and with some consideration to Johnston's and subgenual organs. It is widely accepted that insect hearing organs evolved from pre-existing proprioceptive chordotonal organs. In addition to certain non-neural adaptations for hearing, such as tracheal expansion and cuticular thinning, the chordotonal organs themselves may have intrinsic specializations for sound reception and transduction, and these are discussed. In the future, an integrated approach, using traditional anatomical and physiological techniques in combination with new methodologies in immunohistochemistry, genetics, and biophysics, will assist in refining hypotheses on how chordotonal organs function, and, ultimately, lead to new insights into the peripheral mechanisms underlying hearing in insects.

Expression of prestin, a membrane motor protein, in the mammalian auditory and vestibular periphery

Hair cells are specialized mechanoreceptors common to auditory and vestibular sensory organs of mammalian and non-mammalian species. Different hair cells are believed to share common features related to their mechanosensory function. It has been shown that hair cells possess various forms of motile properties that enhance their receptor function. Membrane-based electromotility is a form of hair cell motility observed in isolated outer hair cells (OHCs) of the cochlea. A novel membrane protein, prestin, recently cloned from gerbil and rat tissues, is presumably responsible for electromotility. We cloned prestin from mouse organ of Corti and confirmed strong homology of this protein among different rodent species. We explored whether or not prestin is present in hair cells of the vestibular system. Using reverse transcription-polymerase chain reaction, we demonstrated that prestin is expressed in mouse and rat auditory and vestibular organs, but not in chicken auditory periphery. In situ hybridization and immunolocalization studies confirmed the presence of prestin in OHCs as well as in vestibular hair cells (VHCs) of rodent saccule, utricle and crista ampullaris. However, in the VHCs, staining of varying intensity with anti-prestin antibodies was observed in the cytoplasm, but not in the lateral plasma membrane or in the stereociliary membrane. Whole-cell patch-clamp recordings showed that VHCs do not possess the voltage-dependent capacitance associated with membrane-based electromotility. We conclude that although prestin is expressed in VHCs, it is unlikely that it supports the form of somatic motility observed in OHCs.

Myo15 function is distinct from Myo6, Myo7a and pirouette genes in development of cochlear stereocilia.

The unconventional myosin genes Myo15, Myo6 and Myo7a are essential for hearing in both humans and mice. Despite the expression of each gene in multiple organs, mutations result in identifiable phenotypes only in auditory or ocular sensory organs. The pirouette (pi) mouse also exhibits deafness and an inner ear pathology resembling that of Myo15 mutant mice and thus may be functionally related to Myo15. In order to investigate possible interactions between Myo15 and Myo6, Myo7a, and the gene affected in pirouette, we crossed Myo15(sh2/sh2) mice to the three other mutant mouse strains. Hearing in doubly heterozygous mice was similar to age-matched singly heterozygous animals, indicating that partial deficiency for both Myo15 and one of these other deafness genes does not reduce hearing. Viable double mutants were obtained from each cross, indicating that potential overlapping functions between these genes in other organs are not essential for viability. All critical cell types of the cochlear sensory epithelium were present in double mutant mice and cochlear stereocilia exhibited a superimposition of single mutant phenotypes. These data suggest that the function of Myo15 is distinct from that of Myo6, Myo7a or pi in development and/or maintenance of stereocilia.

Growth Factors and Early Development of Otic Neurons: Interactions between Intrinsic and Extrinsic Signals

The interplay between intrinsic and extrinsic factors is essential for the transit into different cell states during development. Secreted factors mediate crucial steps in development such as cell growth and survival, bias between self-renewal and differentiation, or choices between different cell fates. Three families of growth factors emerge as critical for early steps in otic neurogenesis: fibroblast growth factors (FGFs); nerve growth factor (NGF)-related neurotrophins (NTs); and insulin-related factors, particularly insulin-like growth factor-1 (IGF-1). Otic neurons connect the sensory mechanoreceptors of the ear, the hair cells, with their targets in the central nervous system (CNS). The generation of otic neurons is a sequential process that includes first the specification of otic precursors in the otic epithelium, second the delamination of epithelial neuroblasts to form the cochleovestibular ganglion (CVG), third the proliferative expansion of ganglionar neuroblasts, and finally the differentiation of neurons that innervate back the vestibular and cochlear (auditory) sensory organs. Each step is characterized by the expression of a particular set of transcription factors and differentially regulated by growth factor signaling, based on which a simple model for early otic neurogenesis is proposed.

Hearing is an active process in mosquitoes

Auditory sensitivity and frequency selectivity are enhanced by active mechanics. Known from vertebrates, active audition is established for an invertebrate. The auditory mechanics of antennal sound receivers in mosquitoes exhibits key diagnostic features of active mechanics: (a) physiologically vulnerable sensitivity and tuning. The mechanical sensitivity and tuning sharpness of the receiver decrease after death. (b) Vulnerable nonlinear mechanical response. Nonlinearity is expressed as amplitude-dependent damping: lower stimulus intensities result in larger response gain and sharper resonance peaks. This band-limited nonlinearity disappears post-mortem. (c) Hypoxia sensitivity. In females, transient exposure to CO2 reversibly reduces tuning sharpness. In males, the effect is opposite: CO2 sharpens tuning and elicits large vibrations in the absence of acoustic stimulation. Hypoxia-induced vibrations are unrelated to external forces; they are autonomous and self-sustained, and forcibly generated by an internal motor. (d) Autonomous antennal vibration (AV). AV is also induced by the injection of dimethylsufoxide (DMSO). DMSO-induced AV persists (90 min) at large amplitudes (>400 nm). Antennal deflection shapes rule out the involvement of muscles, narrowing down the source of mechanical activity to the auditory sense organ at the antennas base. [Work supported by the Leopoldina Academic Society, the Swiss National Science Foundation, BBSRC and the University of Bristol.]

Aquaporin 4 expression in the mammalian inner ear and its role in hearing

Aquaporin 4 (Aqp4), a member of a family of water transport proteins, is a candidate for playing a critical role in inner ear fluid homeostasis. In this study, we assess cross-species Aqp4 expression in the inner ear, sequence conservation in the 5 ′-UTR, and hearing in Aqp4 knockout mice. A single Aqp4 isoform was expressed in a highly conserved pattern within the supporting epithelia surrounding the sensory cells of the auditory and vestibular sensory organs and the glial cells surrounding the auditory nerve of the mouse and rat. In the 5 ′-UTR of mouse and rat Aqp4 gene, sequence conservation was highest in the region spanning the transcription start site. Aqp4 knockout mice demonstrated impaired hearing, but normal neural conduction time. Similar Aqp4 expression pattern and regulatory sequence conservation across species suggest a highly conserved role for Aqp4 in the inner ear. In the Aqp4 deficient mouse, cochlear dysfunction is suggested as the primary cause of hearing impairment in the absence of neural conduction abnormality.

The history of cochlear implantation: from Volta to multichannel-intracochlear stimulation

Seit der Consensus Conference 1995 ist das Cochlear Implant (CI) als Methode der Wahl bei cochleäer Taubheit etabliert. Mit Hilfe des CI ist es erstmals möglich, den auditiven Sinneskanal nach dem Funktionsverlust des Perzeptionsorganes wieder zu nutzen. Weltweit tragen heute mehr als 30 000 Patienten ein CI. Mittlerweile erscheint es bereits selbstverständlich, die Therapie mit Innenohrimplantaten in das Spektrum der HNO-ärztlichen Behandlung als eine Standardtherapie einzubeziehen.

The mechanical basis of Drosophila audition.

In Drosophila melanogaster, antennal hearing organs mediate the detection of conspecific songs. Combining laser Doppler vibrometry, acoustic near-field measurements and anatomical analysis, we have investigated the first steps in Drosophila audition, i.e. the conversion of acoustic energy into mechanical vibrations and the subsequent transmission of vibrations to the auditory receptors in the base of the antenna. Examination of the mechanical responses of the antennal structures established that the distal antennal parts (the funiculus and the arista) together constitute a mechanical entity, the sound receiver. Unconventionally, this receiver is asymmetric, resulting in an unusual, rotatory pattern of vibration; in the presence of sound, the arista and the funiculus together rotate about the longitudinal axis of the latter. According to the mechanical response characteristics, the antennal receiver represents a moderately damped simple harmonic oscillator. The receiver's resonance frequency increases continuously with the stimulus intensity, demonstrating the presence of a non-linear stiffness that may be introduced by the auditory sense organ. This surprising, non-linear effect is relevant for close-range acoustic communication in Drosophila; by improving antennal sensitivity at low song intensities and reducing sensitivity when intensity is high, it brings about dynamic range compression in the fly's auditory system.

Differential expression of Otx2, Gbx2, Pax2, and Fgf8 in the developing vestibular and auditory sensory organs

The vertebrate inner ear is a complex organ with vestibular and auditory sensory functions, which derives from a single ectoderm structure, the otic placode. The development and regional patterning of the otic primordium is determined by the restricted expression of several genes. Here, we show the expression pattern of three transcription factors ( Otx2, Gbx2, Pax2) and of a member of the fibroblast growth factor family ( Fgf8) in the developing chick inner ear, and we correlate these patterns with the developing sensory and nonsensory elements.

Genes, deafness, and balance disorders

The mammalian auditory sense organ is subdivided into three principle compartments, the outer-, middle- and inner ear. The main task of the outer- and middle ear is to channel sound waves towards the cochlea within the inner ear. The inner ear also contains the vestibule, the end organ for the perception of gravity and acceleration. Hair cells within the sensory epithelia of the cochlea and the vestibule contain stereocilia that harbor mechanically gated ion channels. These ion channels open or close upon deflection of the stereocilia leading to changes in cell polarization and the rate of neurotransmitter release from hair cells onto sensory neurons. In this way, mechanical signals evoked by sound waves or head movement are transformed into electrochemical signals. The positional cloning of human disease genes and the analysis of mouse mutants has led to the identification of numerous genes that cause deafness and balance disorders. These findings provide insights into the molecular and cellular requirements for mechanosensory transduction and establish an entry point to understand deafness at the molecular level. We will summarize results that have shed light on the function of extracellular matrix glycoproteins, cell adhesion molecules, and components of the actin cytoskeleton in the inner ear.

Confocal imaging of sensory organ formation during Xenopus inner ear development

During Xenopus larval development, the otic vesicle gradually gives rise to several auditory and vestibular sensory organs that are innervated by the eighth cranial nerve. Confocal imaging methods were used to examine the appearance of stereociliary bundles in the various sensory epithelia during Xenopus development (stages 28–50), and to gather comparative developmental data that show how the inner ear gradually forms distinct auditory and vestibular compartments discernable by larval stage 50. A BioRad MRC-1024 confocal microscope was used to collect digitized images from inner ears labeled with Alexa 488 phalloidin to detect the actin cytoskeleton, and propidium iodide to highlight DNA in cell nuclei. Confocal images from whole inner ears clearly show the developmental emergence of uniquely organized sensory epithelia populated by hair cell bundles of varying morphology, as well as the position of the sensory ganglia relative to the endorgans. Images from sectioned tissue provide additional information about cell morphology and the arrangement of the sensory epithelium in relation to neural innervation. Taken together, the image data illustrate the complex structural changes that underlie inner ear organogenesis and morphogenesis. [Work supported by grants to EES (NIH NIGMS, NASA) and awards to QQ (NASA NMSGC Fellowship, NIGMS RISE).]

SEM search for sound production and sound perception organs in a variety of Drosophila species.

The discovery of new song types in Hawaiian Drosophila species has raised a question of alternative sound production and sound perception mechanisms in Drosophila. For example D. disjuncta males, which produce song with a carrier frequency of 5,000-6,000 Hz, do not vibrate their wings but only tremble them in a very small amplitude while singing. In our SEM search we found at the wing base of the flies of this species a special structure which could play a part in song production. Our search for auditory sense organs in Drosophila species revealed that the aristae on fly antennae are structurally not as simple as has been assumed. In Hawaiian D. planitibia subgroup species, the fly aristae are more regular than in other Drosophila species and they are also covered with small hairs. We did not find any sign of tympanic hearing organs in any of the studied species, but we discovered on fly legs and wings sensilla which could play a part in sound/vibration perception. Our SEM survey provides a good starting point in the search for alternative pathways for sound production and perception via transmission electron microscopy combined with behavioral studies and electrophysiological recordings in Hawaiian Drosophila species.

Targeted disruption of otog results in deafness and severe imbalance.

Genes specifically expressed in the inner ear are candidates to underlie hereditary nonsyndromic deafness. The gene Otog has been isolated from a mouse subtractive cDNA cochlear library. It encodes otogelin, an N-glycosylated protein that is present in the acellular membranes covering the six sensory epithelial patches of the inner ear: in the cochlea (the auditory sensory organ), the tectorial membrane (TM) over the organ of Corti; and in the vestibule (the balance sensory organ), the otoconial membranes over the utricular and saccular maculae as well as the cupulae over the cristae ampullares of the three semi-circular canals. These membranes are involved in the mechanotransduction process. Their movement, which is induced by sound in the cochlea or acceleration in the vestibule, results in the deflection of the stereocilia bundle at the apex of the sensory hair cells, which in turn opens the mechanotransduction channels located at the tip of the stereo-cilia. We sought to elucidate the role of otogelin in the auditory and vestibular functions by generating mice with a targeted disruption of Otog. In Otog-/- mice, both the vestibular and the auditory functions were impaired. Histological analysis of these mutants demonstrated that in the vestibule, otogelin is required for the anchoring of the otoconial membranes and cupulae to the neuroepithelia. In the cochlea, ultrastructural analysis of the TM indicated that otogelin is involved in the organization of its fibrillar network. Otogelin is likely to have a role in the resistance of this membrane to sound stimulation. These results support OTOG as a possible candidate gene for a human nonsyndromic form of deafness.

SEM search for sound production and sound perception organs in a variety ofDrosophila species

The discovery of new song types in Hawaiian Drosophila species has raised a question of alternative sound production and sound perception mechanisms in Drosophila. For example D. disjuncta males, which produce song with a carrier frequency of 5,000-6,000 Hz, do not vibrate their wings but only tremble them in a very small amplitude while singing. In our SEM search we found at the wing base of the flies of this species a special structure which could play a part in song production. Our search for auditory sense organs in Drosophila species revealed that the aristae on fly antennae are structurally not as simple as has been assumed. In Hawaiian D. planitibia subgroup species, the fly aristae are more regular than in other Drosophila species and they are also covered with small hairs. We did not find any sign of tympanic hearing organs in any of the studied species, but we discovered on fly legs and wings sensilla which could play a part in sound/vibration perception. Our SEM survey provides a good starting point in the search for alternative pathways for sound production and perception via transmission electron microscopy combined with behavioral studies and electrophysiological recordings in Hawaiian Drosophila species.

Mosquito hearing: sound-induced antennal vibrations in male and femaleAedes aegypti

Male mosquitoes are attracted by the flight sounds of conspecific females. In males only, the antennal flagellum bears a large number of long hairs and is therefore said to be plumose. As early as 1855, it was proposed that this remarkable antennal anatomy served as a sound-receiving structure. In the present study, the sound-induced vibrations of the antennal flagellum in male and female Aedes aegypti were compared, and the functional significance of the flagellar hairs for audition was examined. In both males and females, the antennae are resonantly tuned mechanical systems that move as simple forced damped harmonic oscillators when acoustically stimulated. The best frequency of the female antenna is around 230 Hz; that of the male is around 380 Hz, which corresponds approximately to the fundamental frequency of female flight sounds. The antennal hairs of males are resonantly tuned to frequencies between approximately 2600 and 3100 Hz and are therefore stiffly coupled to, and move together with, the flagellar shaft when stimulated at biologically relevant frequencies around 380 Hz. Because of this stiff coupling, forces acting on the hairs can be transmitted to the shaft and thus to the auditory sensory organ at the base of the flagellum, a process that is proposed to improve acoustic sensitivity. Indeed, the mechanical sensitivity of the male antenna not only exceeds the sensitivity of the female antenna but also those of all other arthropod movement receivers studied so far.