Molecular mechanisms of inner ear development

Abstract

The inner ear is a structurally complex vertebrate organ built to encode sound, motion, and orientation in space. Given its complexity, it is not surprising that inner ear dysfunction is a relatively common consequence of human genetic mutation. Studies in model organisms suggest that many genes currently known to be associated with human hearing impairment are active during embryogenesis. Hence, the study of inner ear development provides a rich context for understanding the functions of genes implicated in hearing loss. This chapter focuses on molecular mechanisms of inner ear development derived from studies of model organisms.

Figure 1.

Morphogenesis of the mouse inner ear. Lateral views of paint-filled, right inner ears from E10.75 to E17 (E, embryonic age). Abbreviations: aa, anterior ampulla; asc, anterior semicircular canals; cc, common crus; co, cochlear duct; csd, cochlear saccular duct; ed, endolymphatic duct; es, endolymphatic sac; hp, horizontal canal pouch; la, lateral ampulla; lsc, lateral semicircular canal; pa, posterior ampulla; psc, posterior semicircular canal; s, saccule; u, utricle; vp, vertical canal pouch. Orientations: A, anterior; D, dorsal. (Adapted from Morsli et al. 1998.)

Figure 2.

Development of the neural–sensory domain. (A) Proposed derivatives of the NSD based on gene expression patterns and fate-mapping studies. The NSD originated in the anterior region of the invaginating otic placodes, where neuroblasts delaminate to form neurons of the CVG (red). This region also gives rise to macula of the utricle and saccule, as well as nonsensory regions (gray) between sensory patches and the GER of the cochlea. Three separate domains (pink with blue stripes) that form at the fringe or within a subdivision of the NSD at the otocyst stage give rise to the three cristae and organ of Corti. (B) Neural-competent cells expressing Ngn1 and Delta within the neural–sensory domain delaminate from the otic epithelium to form neuroblasts (red). Delta1 inhibits neighboring cells from developing into neuroblasts via lateral inhibition. The remaining sensory epithelium, including cells that once express Ngn1, develops into sensory hair cells (green), supporting cells (pink), and some nonsensory cells (gray). Asterisks represent genes expressed in a subpopulation of cells within the NSD. Abbreviations: ac, anterior crista; asc, anterior semicircular canal; CVG, cochleovestibular ganglion; ed, endolymphatic duct; es, endolymphatic sac; GER, greater epithelial ridge; lc, lateral crista; lsc, lateral semicircular canal; ms, macula of the saccule; mu, macula of the utricle; NSD, neural–sensory-competent domain; oc, organ of Corti; pc, posterior crista; psc, posterior semicircular canal; SG, spiral ganglion; VG, vestibular ganglion.

Figure 3.

Anterior–posterior axial specification. (A) X-Gal histochemical staining in RARE-lacZ mouse embryos shows a transient gradient of RA responsiveness in the developing inner ear. At the otic cup stage (E8.75), the anterior border of the RA responsiveness is in the middle of the otic cup, and this border shifts caudally by the time the otocyst is formed at E9.5. (B) Model of RA signaling in AP patterning of the chicken inner ear. Somites expressing high levels of the RA synthetic enzyme Raldh2 act as the main source of RA for patterning the inner ear. The RA degradation enzyme Cyp26C1, expressed in the ectoderm rostral to the ear rudiment, modulates the level of RA signaling perceived by the otic epithelium. Secreted molecules from the hindbrain such as Wnts (purple) and Shh (yellow) provide DV axial information to the inner ear.

Figure 4.

Dorsal–ventral axial specification. The hindbrain is the source of DV axial signaling for the inner ear. In the wild-type embryo, the developing inner ear receives Wnt signaling from the dorsal hindbrain (purple triangle) and Sonic hedgehog (Shh) from the ventral midline structures of the floor plate and notochord. The graded Shh signaling (yellow triangle) results in various levels of Gli activator (blue triangle) and repressor (red triangle) activities in the developing inner ear. The distal region of the cochlear duct (blue region) requires a high level of Gli activators. The proximal region of the cochlear duct and the saccule (pale red and blue region) require relatively lower levels of Shh signaling for counteracting the Gli3 repressor function. The dorsal vestibular region of the inner ear (red region) requires Gli3 repressor function. In Shh^−/− ears only the dorsal vestibular region develops. In Wnt1/Wnt3a double mutants, the inner ear is rudimentary and cystic. However, Shh signaling appears normal based on the expression of putative Shh target genes such as Pax2 and Otx2. Whether Gli3 repressor activities are still present in the inner ear of the compound mutant is not clear.

Figure 5.

Fate-mapping studies of the mouse and chicken inner ear. (A) Genetic fate mapping of the Wnt-responsive cells in the mouse otic cup. X-Gal histochemical staining of a Wnt reporter strain, Topgal, shows that Wnt-responsive cells are restricted to the dorsal otic placode (a) and otocyst (b). However, tracing the progeny of the Wnt-responsive cells in TopCreERT2/Rosa-lacZ embryos after administration of tamoxifen at E8.75 shows these cells expanded ventrally (d, arrows) beyond the initial TopCreERT2 mRNA domain (c). (B) Fate mapping of the rim of the chicken otic cup. Injection of lipophilic dyes to the indicated numerical positions at the rim of the otic cup shows that a majority of the cells that constitute the lateral wall of the otocyst (light blue) originated from the ventral posterior region of the otic cup at positions 6, 7, and 8. The most dorsal region of the otocyst, constituted by cells in positions 1, 2, 10, 11, and 12, displaces medially to form the endolymphatic duct (tan). The lateral region of the otocyst gives rise to the vertical and horizontal canal pouches. (A adapted from Riccomagno et al. 2005; B adapted from Brigande et al. 2000; Fig. 5 reprinted, with permission, © Elsevier.)

Figure 6.

Model of semicircular canal formation. The growth of the canal pouch is promoted by a canal genesis zone (dark blue area), which is induced by the presumptive cristae (black circles). The canal genesis zone gives rise to most of the cells in the canals (blue) and some cells within the common crus (blue dots). The center region of each prospective canal (tan) forms a fusion plate that resorbs, leaving behind two canals joined by the common crus. The bottom panel shows examples of a number of mutant phenotypes. Failure to induce proper growth of the canal pouch or in specification of the resorption domain can result in only a canal pouch, such as in Netrin1 and Fgf9 mutants. Expansion of the resorption domain, as in the case of Lrig3 nulls, or failure to specify the rim of the canal pouch can lead to absence or truncation of the canal, or possibly a canal with smaller caliber.

Figure 7.

Development of the cochlear duct. Schematic cross sections through the base of the cochlear duct at the indicated ages. At E12 the prosensory domain, as defined by expression of Jag1/Sox2/Lfng, is already present, as is a domain of expression of Bmp4 located at the lateral edge of the duct. There is limited overlap between the two domains. In addition, developing spiral ganglion neurons are located near the medial edge of the duct. At E13 developing spiral ganglion neurons begin to express Sonic hedgehog (Shh), which is believed to diffuse into the medial side of the duct, and p27^kip1 is expressed in a subset of the Jag1/Sox2/Lfng-positive cells. BMP4 and Shh are believed to restrict sensory formation to the middle of the duct. At E14 Atoh1 expression begins in a subset of the p27^kip1-positive cells. By E16 Atoh1 expression is resolved to hair cells, and Fgfr3 is expressed in developing pillar and Deiters’ cells. At postnatal day 0 (P0), cellular patterning is essentially complete. Atoh1-positive hair cells are surrounded by supporting cells and spiral ganglion neurons no longer express Shh. Abbreviations: IHC, inner hair cells; IP, inner pillar cell; KO, Kolliker’s organ; OC, organ of Corti; OP, outer pillar cell.

Figure 8.

Planar cell polarity in the inner ear. (A) Schematic surface view of the organ of Corti, illustrating the uniform orientation of stereociliary bundles (green) in hair cells (blue). (B) Schematics of the saccule (upper) and utricle (lower) from a mouse. Arrows, orientation of the stereociliary bundles. The striolar reversal zones are marked in green. (C) Development of a stereociliary bundle includes movement of the developing kinocilium (red) from the center of the lumenal surface of the cell toward the outer edge. At the same time a subset of microvilli (green) grows and extends to develop as stereocilia, while other microvilli recede. (D) The PCP molecules Dvl1, -2, and -3, Vangl2, and Fz3 and -6 become asymmetrically localized to different sides of developing hair cells before the movement of the developing kinocilium to the lateral edge of the lumenal surface. The direction of the kinocilium movement is toward the side of the cell with Dvl1, -2, and -3, and away from the side of the cell that contains Vangl2 and Fz3 and -6.