Neurosensory development and cell fate determination in the human cochlea.

BACKGROUND: Hearing depends on correct functioning of the cochlear hair cells, and their innervation by spiral ganglion neurons. Most of the insight into the embryological and molecular development of this sensory system has been derived from animal studies. In contrast, little is known about the molecular expression patterns and dynamics of signaling molecules during normal fetal development of the human cochlea. In this study, we investigated the onset of hair cell differentiation and innervation in the human fetal cochlea at various stages of development. RESULTS: At 10 weeks of gestation, we observed a prosensory domain expressing SOX2 and SOX9/SOX10 within the cochlear duct epithelium. In this domain, hair cell differentiation was consistently present from 12 weeks, coinciding with downregulation of SOX9/SOX10, to be followed several weeks later by downregulation of SOX2. Outgrowing neurites from spiral ganglion neurons were found penetrating into the cochlear duct epithelium prior to hair cell differentiation, and directly targeted the hair cells as they developed. Ubiquitous Peripherin expression by spiral ganglion neurons gradually diminished and became restricted to the type II spiral ganglion neurons by 18 weeks. At 20 weeks, when the onset of human hearing is thought to take place, the expression profiles in hair cells and spiral ganglion neurons matched the expression patterns of the adult mammalian cochleae. CONCLUSIONS: Our study provides new insights into the fetal development of the human cochlea, contributing to our understanding of deafness and to the development of new therapeutic strategies to restore hearing.

GATA3 controls the specification of prosensory domain and neuronal survival in the mouse cochlea.

HDR syndrome (also known as Barakat syndrome) is a developmental disorder characterized by hypoparathyroidism, sensorineural deafness and renal disease. Although genetic mapping and subsequent functional studies indicate that GATA3 haplo-insufficiency causes human HDR syndrome, the role of Gata3 in sensorineural deafness and auditory system development is largely unknown. In this study, we show that Gata3 is continuously expressed in the developing mouse inner ear. Conditional knockout of Gata3 in the developing inner ear disrupts the morphogenesis of mouse inner ear, resulting in a disorganized and shortened cochlear duct with significant fewer hair cells and supporting cells. Loss of Gata3 function leads to the failure in the specification of prosensory domain and subsequently, to increased cell death in the cochlear duct. Moreover, though the initial generation of cochleovestibular ganglion (CVG) cells is not affected in Gata3-null mice, spiral ganglion neurons (SGNs) are nearly depleted due to apoptosis. Our results demonstrate the essential role of Gata3 in specifying the prosensory domain in the cochlea and in regulating the survival of SGNs, thus identifying a molecular mechanism underlying human HDR syndrome.

Making connections in the inner ear: recent insights into the development of spiral ganglion neurons and their connectivity with sensory hair cells.

In mammals, auditory information is processed by the hair cells (HCs) located in the cochlea and then rapidly transmitted to the CNS via a specialized cluster of bipolar afferent connections known as the spiral ganglion neurons (SGNs). Although many anatomical aspects of SGNs are well described, the molecular and cellular mechanisms underlying their genesis, how they are precisely arranged along the cochlear duct, and the guidance mechanisms that promote the innervation of their hair cell targets are only now being understood. Building upon foundational studies of neurogenesis and neurotrophins, we review here new concepts and technologies that are helping to enrich our understanding of the development of the nervous system within the inner ear.

Lmx1a maintains proper neurogenic, sensory, and non-sensory domains in the mammalian inner ear.

Lmx1a is a LIM homeodomain-containing transcription factor, which is required for the formation of multiple organs. Lmx1a is broadly expressed in early stages of the developing inner ear, but its expression is soon restricted to the non-sensory regions of the developing ear. In an Lmx1a functional null mutant, dreher (dr(J)/dr(J)), the inner ears lack a non-sensory structure, the endolymphatic duct, and the membranous labyrinth is poorly developed. These phenotypes are consistent with Lmx1a's role as a selector gene. More importantly, while all three primary fates of the inner ear - neural, sensory, and non-sensory - are specified in dr(J)/dr(J), normal boundaries among these tissues are often violated. For example, the neurogenic domain of the ear epithelium, from which cells delaminate to form the cochleovestibular ganglion, is expanded. Within the neurogenic domain, the demarcation between the vestibular and auditory neurogenic domains is most likely disrupted as well, based on the increased numbers of vestibular neuroblasts and ectopic expression of Fgf3, which normally is associated specifically with the vestibular neurogenic region. Furthermore, aberrant and ectopic sensory organs are observed; most striking among these is vestibular-like hair cells located in the cochlear duct.

Spiral ligament pathology: a major aspect of age-related cochlear degeneration in C57BL/6 mice.

Data from systematic, light microscopic examination of cochlear histopathology in an age-graded series of C57BL/6 mice (1.5-15 months) were compared with threshold elevations (measured by auditory brain stem response) to elucidate the functionally important structural changes underlying age-related hearing loss in this inbred strain. In addition to quantifying the degree and extent of hair cell and neuronal loss, all structures of the cochlear duct were qualitatively evaluated and any degenerative changes were quantified. Hair cell and neuronal loss patterns suggested two degenerative processes. In the basal half of the cochlea, inner and outer hair cell loss proceeded from base to apex with increasing age, and loss of cochlear neurons was consistent with degeneration occurring secondary to inner hair cell loss. In the apical half of the cochlea with advancing age, there was selective loss of outer hair cells which increased from the middle to the extreme apex. A similar gradient of ganglion cell loss was noted, characterized by widespread somatic aggregation and demyelination. In addition to these changes in hair cells and their innervation, there was widespread degeneration of fibrocytes in the spiral ligament, especially among the type IV cell class. The cell loss in the ligament preceded the loss of hair cells and/or neurons in both space and time suggesting that fibrocyte pathology may be a primary cause of the hearing loss and ultimate sensory cell degeneration in this mouse strain.

The effects of low-frequency ultrasound on the inner ear: an electrophysiological study using the guinea pig cochlea.

By examining 218 albino guinea pigs, electrophysiological methods were used to investigate the effects of low frequency ultrasound at moderate sound pressure levels after long-term exposure to the inner ear. From 10 kHz to 28 kHz, low frequency ultrasound below 100 dB SPL induced significant changes in cochlear microphonics, elevated thresholds and decreased maximum output voltage of action potentials and decreased absolute values of negative potentials of the endocochlear potentials.

An experimental study on the frog semicircular canal--functions of cupula and vestibular ganglion.

An isolated frog posterior semicircular canal was stimulated either by mechanical or thermal endolymphatic flow. Ampullofugal stimulation induced action potentials from the posterior ampullary nerve. These potentials adapted in six to eight seconds. While mechanical endolymphatic flow was induced, the cupula maintained its shift position without swinging back to the neutral point. This indicates that adaptation is mainly of sensorineural origin. When the canal was stimulated by a piece of frozen Ringer's solution, an increase in spike discharge was observed. The responses from the vestibular nerve trunk were compared to those from the posterior ampullary nerve. The time course of the response was shorter and the maximum spike count was smaller in the recordings from the vestibular nerve trunk.

Spatial organization in the saccule and lagena of a teleost: hair cell pattern and innervation.

The relationship between the hair cell orientation pattern and innervation in the saccule and lagena of the teleost Helostoma temmincki (the kissing gourami) was investigated with scanning electron microscopy and the Winkelmann-Schmitt silver impregnation technique. The hair cell pattern in the saccule consists of four orthogonally oriented groups. The anterior two groups are oriented along the animal's rostrocaudal axis, and the posterior two are oriented along its dorsoventral axis. The pattern of hair cell orientations in the lagena is a typical bidirectional one. Two divisions of the eighth nerve innervate the saccule. The anterior division innervates the horizontally oriented hair cell groups, and the posterior division innervates the dorsoventrally oriented groups. A single nerve innervates the lagena, with the majority of fibers innervating one or the other of the two lagenar hair cell groups. The segregated pattern of innervation according to hair cell orientation groups in the saccule was confirmed in other species. Individual types of axonal terminations appear to innervate hair cells of specific ciliary bundle types.

The origin of efferent fibers to the inner ear in a turtle (Terrapene ornata). A horseradish peroxidase study.

The origin of efferent acoustic and vestibular fibers was determined in the turtle Terrapene ornata. After injection of an aqueous solution of horseradish peroxidase (HRP) into either the cochlear duct or into the ampullae of the horizontal and anterior semicircular canals, neurons in the medullary reticular formation were labeled by the reaction production of retrogradely transported HRP. These neurons were located bilaterally in the medial reticular nucleus. The majority were found ipsilateral to the injection site. There was no demonstrable difference in size, shape, and labeling pattern between efferent acoustic and efferent vestibular neurons. The crossed component component of efferent acoustic fibers, however, was rather sparsely developed.

The cochlear nuclei of snakes.

The cochlear nuclei of three burrowing snakes (Xenopeltis unicolor, Cylindrophis rufus, and Eryx johni) and three non-burrowing snakes (Epicrates cenchris, Natrix sipedon, and Pituophis catenifer) were studied. The posterior branch of the statoacoustic nerve and its posterior ganglion were destroyed and the degenerated nerve fibers and terminals traced to primary cochlear nuclei in 13 specimens of Pituophis catenifer. All these snake species possess three primary and one secondary cochlear nuclei. The primary cochlear nuclei consist of a small nucleus angularis located at the cerebello-medullary junction and a fairly large nucleus magnocellularis forming a dorsal cap over the cephalic end of the alar eminence. Nucleus magnocellularis may be subdivided into a medially placed group of rounder cells, nucleus magnocellularis medialis, and a laterally placed group of more ovate and paler-staining cells, nucleus magnocellularis lateralis. A small but well-defined secondary nucleus which showed no degenerated nerve terminals after nerve root section, nucleus laminaris, underlies the cephalic part of both nucleus magnocellularis medialis and nucleus magnocellularis lateralis. Larger and better-developed cochlear nuclei were found in burrowing species than in non-burrowing species of snakes. Of the three burrowing species studied, Xenopeltis showed the greatest development of cochlear nuclei; Eryx cochlear nuclei were not quite as large but were better differentiated than in Xenopeltis; and Cylindrophis cochlear nuclei were fairly large but not as well developed nor as well differentiated as in either Xenopeltis or Eryx. The cochlear nuclei of the three non-burrowing snakes, Epicrates, Natrix, and Pituophis, were not as large nor as well developed as those of the burrowing snakes. There is some, but not complete, correlation between cochlear development and papilla basilaris length and number of hair cells. Thus, Xenopeltis and Eryx, with well-developed cochlear nuclei, have relatively long papillae basilares; but the boid, Epicrates, with less well-developed cochlear nuclei, has a fairly well-developed papilla basilaris. Cylindrophis, a burrowing species, shows only a moderate degree of cochear nuclei and papilla basilaris development. The non-burrowers, Natrix and Pituophis, have both small cochlear nuclei and relatively short papillae basilares.