NCBI Bookshelf. A service of the National Library of Medicine, National Institutes of Health.

Park BY, Saint-Jeannet JP. Induction and Segregation of the Vertebrate Cranial Placodes. San Rafael (CA): Morgan & Claypool Life Sciences; 2010.

Cover of Induction and Segregation of the Vertebrate Cranial Placodes

Induction and Segregation of the Vertebrate Cranial Placodes.

Show details

Induction and Segregation of the Cranial Placodes

At the end of gastrulation, the ectoderm of the vertebrate embryo can be divided into three major domains: the non-neural ectoderm and the neural plate separated by a third region known as the neural plate border. The non-neural ectoderm and neural plate will develop into epidermis and central nervous system, respectively, while the neural plate border contains at least two cell populations: neural crest and pre-placodal ectoderm. The neural crest is located lateral to the neural plate but excluded from its most anterior region. The pre-placodal ectoderm is restricted to the anterior region of the embryo, lateral to the neural crest and at the most rostral boundary of the neural plate. The pre-placodal ectoderm abuts directly the neural plate anteriorly, and the neural crest in the lateral regions (Figure 1). The restriction of placodal fate to anterior regions of the neural plate border is not due to restrictions in competence at the gastrula stage since all regions of the ectoderm are competent to form placodes, and this competence is maintained until fairly late in development, at least for some placodes (Jacobson, 1966; Noramly and Grainger, 2002). Instead, the cranial restriction of placodal fate must be explained by local induction.

Formation of the pre-placodal region is initiated through a series of events that first define the neural plate border and subsequently subdivide the border into placode and neural crest precursors (Figure 1). This is achieved through interactions with surrounding tissues, neural plate, the future epidermis and underlying head mesoderm, all of which secrete factors controlling placode versus neural crest fate. Therefore, different signaling events converge to position the placode and the neural crest territories next to the neural plate. Because neural crest and placodes originate from the same region of the embryo, the neural plate border, it is likely that the same set of factors will be differentially deployed for the induction of both cell types (reviewed in Knecht and Bronner-Fraser, 2002; Huang and Saint-Jeannet, 2004). How these signals are integrated at the neural plate border to generate distinct fates is an important and unresolved question.

Placodes are often discussed as independent structures, as they arise from non-overlapping ectodermal thickenings. However, most current models of placode induction involve a multi-step process starting with the formation of a pre-placodal region in which individual fates are initially intermingled (Kozlowski et al., 1997; Streit, 2004; Battacharrya and Bronner-Fraser, 2004; Xu et al., 2008). Later, placodes with distinct identities segregate in a stereotypical antero-posterior pattern in response to additional inductive cues (Figure 1). This model originates primarily in the pioneering studies of Antone Jacobson performed in the 60’s using amphibian embryos (Jacobson, 1963a; b; c; 1966), studies that are now largely supported by molecular analyses (reviewed in Baker and Bronner-Fraser, 2001; Streit, 2004; 2007; Battacharrya and Bronner-Fraser, 2004; Schlosser and Ahrens, 2004; Brugmann and Moody, 2005; Schlosser, 2006; Bailey and Streit, 2006; McCabe and Bronner-Fraser, 2009).

Historical perspective

In a series of experiments performed in two different species of amphibians, Taricha torosa and Ambystoma punctatum, Jacobson evaluated the abilities of different tissues to induce lens, otic or olfactory placodes during development (Jacobson 1966;1963a; 1963b). These studies demonstrated that the lens and olfactory placodes can be induced by early signals from the endoderm and mesoderm, while the otic placode is induced by mesoderm- and neuroectoderm-derived signals. These signals are however not sufficient to elicit development of a complete lens, inner ear or olfactory structure. By evaluating the proximity of the presumptive placodal ectoderm to these inducers during development of the lens, Jacobson proposed that the presumptive lens ectoderm is initially induced by the endoderm, followed by inducing signals from the intervening mesoderm, and eventually from signals derived from the optic cup, adjacent to the lens placode. Based on these observations Jacobson proposed that a similar sequence of inductions could be applied to the formation of other placodes.

In another set of experiments Jacobson demonstrated that within the ectoderm adjacent to the anterior neural plate, cells are competent to give rise to any placode, but progressively lose this ability overtime, as placodal domains start to segregate within the pre-placodal ectoderm (Jacobson, 1963c). In these experiments the pre-placodal ectoderm is dissected and rotated along its antero-posterior axis at the early neurula stage, so that prospective ear ectoderm end up next to forebrain, and prospective nose ectoderm next to hindbrain (Figure 10A). Within the transplanted pre-placodal ectoderm, placodes develop according to their novel (ectopic) position along the antero-posterior axis, with occasional formation of anterior ectopic ears (Figure 10B). In the same experiments performed at a later stage in development, placodes developed according to their original position, suggesting that the identity of the pre-placodal ectoderm was already determined at the stage of transplantation. However, some plasticity persists in the transplanted ectoderm since olfactory tissues are induced adjacent to the ectopic anterior ears, and ears are induced next to ectopic posterior noses (Figure 10B). Therefore, at early neurula stage cells within the pre-placodal ectoderm are competent to generate a placode distinct from their normal fate, while at late neurula stage their fate appears to be determined (Jacobson, 1963c). These experiments highlight three key points regarding the specification of cranial placodes: (i) all placodes originate from a common domain; (ii) there is a shared inductive mechanism for all placodes; and (iii) placode induction is a multi-step process.

Figure 10. Diagrams illustrating Antone Jacobson’s pre-placodal ectoderm rotation experiments.

Figure 10

Diagrams illustrating Antone Jacobson’s pre-placodal ectoderm rotation experiments. (A) The pre-placodal ectoderm of an amphibian neurula stage donor embryo was dissected and transplanted on a host embryo so that its antero-posterior polarity (more...)

A common domain for all cranial placodes

The pre-placodal ectoderm surrounding the anterior neural plate at the late gastrula/early neurula stages has been described in mouse, chick, amphibian and fish embryos. A large body of work, including that of Antone Jacobson described above, has shown that induction of different placodes involves similar sequential interactions between the presumptive pre-placodal region and surrounding tissues (reviewed in Baker and Bronner-Fraser, 2001).

Lineage and fate map analyses in fish and chick embryos at the gastrula and neurula stage have demonstrated that this region of the embryonic ectoderm, adjacent to the prospective brain contains precursors for most cranial placodes (Kozlowski et al., 1997; Bhattacharyya et al., 2004; Streit, 2002). In this territory, precursors for different placodes are initially intermingled and eventually recruited into distinct areas along the antero-posterior axis. Using focal dye labeling, recent fate maps of the olfactory and lens (Bhattacharyya et al., 2004), and the trigeminal, epibranchial, and otic placodes (Streit, 2002; Xu et al., 2008) demonstrated that adjacent ectodermal cell populations can contribute to distinct placodes.

The fact that the pre-placodal ectoderm is formed by an heretogenous population placodal precursors is consistent with the notion that all placodal precursors are generated by a common inductive mechanism and share a similar developmental program. This is a view supported by molecular analyses showing the co-expression of specific genes in the domain surrounding the anterior neural plate (see Tables 17). Recent work suggests that all placodal precursors start with the same initial fate, a lens fate (Bailey, et al., 2006); this is also supportive of the hypothesis of a commonality of organization and processes underlying the formation of all cranial placodes.

Lens, the ground state of all sensory placodes

Recently, it has been proposed that the entire pre-placodal region is initially specified as lens (Bailey, et al., 2006), the simplest form of all ectodermal placodes, giving rise to only two cell types (Jacobson, 1966). When antero-posterior segments of the pre-placodal ectoderm are cultured in isolation in a defined medium, all segments form lens structures and express lens-specific genes, even cells that normally never contribute to the lens, like the most posterior segmentsof pre-placodal ectoderm. Moreover, none of these explants express markers specific for olfactory, trigeminal or otic placodes. All placodal cells, regardless of their final fate have an initial lens character, which represents the ground state for all sensory placodes (Bailey, et al., 2006). This model implies that in normal placode development the acquisition of a specific identity requires an initial step of repression of lens character, prior to or concomitant to the activation of a program specific for olfactory, trigeminal or otic fate. Since lens placode is non-neurogenic, non-lens placodes also need to acquire neurogenic properties in the process.

Among the possible candidate molecules repressing lens character in the pre-placodal region are members of Fibroblast Growth Factors (Fgf) family. As recently described, exposure of the presumptive lens ectoderm to Fgf8 blocks expression of the lens marker Pax6 and promotes olfactory placode character. On the other hand, markers for adenohypophyseal, trigeminal or otic placodes are not induced under these conditions (Bailey, et al., 2006). These results indicate that Fgf8 signaling represses lens fate and is sufficient to elicit olfactory placode development. This work also suggests that a different signal (or combination of signals) might be required to specify other placodes. Alternatively, because several members of the Fgf family (including Fgf3, Fgf10 and Fgf19) are implicated at least in adenohypophyseal, otic and epibranchial placodes formation in several species (see below), other Fgf ligands besides Fgf8, might be implicated in the specification of other placodes.

Inducing factors in the generation of cranial placodes

The initial overlap between different placode precursors within the pre-placodal ectoderm has made it very challenging to identify the specific inductive signals for the different placodes. Several classes of signaling molecules are implicated in the induction of the pre-placodal region and its subsequent sub-division into domains with distinct placodal identities. A number of recent review articles have discussed some of these findings (Baker and Bronner-Fraser, 2001; Bailey and Streit, 2006; Schlosser, 2006; McCabe and Bronner-Fraser, 2009); here we will summarize the activity of these molecules focusing more specifically on the Bone Morphogenetic Protein (Bmp), Fibroblast Growth Factor (Fgf) and Wnt signaling pathways.

Bone Morphogenetic Protein Signaling

While unperturbed Bmp signaling in the ectoderm converts cells into epidermis, attenuation of Bmp activity in the ectoderm results in emergence of neural, neural crest and placodal cells (for review Sasai and De Robertis, 1997; Stern, 2005). Studies in frog and fish indicate the neural plate border forms in region of the ectoderm where Bmp signaling is partially attenuated by Bmp antagonists, such as Chordin, Noggin and Follistatin, derived from the axial mesoderm (Marchant et al., 1998; Nguyen et al., 1998; Tribulo et al., 2003). In Xenopus a balance of Bmps and their antagonists is in part responsible for positioning the pre-placodal ectoderm (Brugmann et al., 2004; Glavic et al., 2004). Xenopus animal caps treated with different concentrations of the Bmp antagonist, Noggin, form epidermis in the presence of high levels of Bmp activity, while neural crest and pre-placodal cells are generated at intermediate levels and neural plate at low levels (Wilson, et al., 1997; Tribulo, et al., 2003; Brugmann, et al., 2004; Glavic, et al., 2004). Zebrafish embryos with mutations in distinct components of the Bmp signaling pathway show expanded neural crest domain while the placode territory is somewhat displaced, but not expanded (Neave, et al., 1997; Nguyen, et al., 1998).

Bmp signals are also implicated in the specification of olfactory and lens placodes in the chick embryo. At gastrula stages Bmp2 and Bmp4 are expressed in the pre-placodal region. In an explant assay using placodal progenitor cells, short exposure to Bmp signaling promotes specification of olfactory fate, while prolonged exposure to Bmp signals promotes formation of lens cells at the expense of placodal cells. Bmp signaling is also sufficient to promote olfactory and lens progenitors in forebrain explants (Sjodal et al., 2007).

Bmp4-/- mouse embryos lack lens placodes, but expression of Six3 and Pax6 is detected in the prospective lens ectoderm and the olfactory placode appears normal (Furuta and Hogan, 1998). These results suggest that placodal progenitor cells are induced without Bmp4, which may reflect functional redundancy with other Bmp family members such as Bmp7. These results also indicate that Bmp4 activity is required for differentiation of lens but not olfactory placodal cells after the initial specification of placodal progenitors. Bmp4 alone is not sufficient to promote lens development suggesting that it may act with other inducers (Furuta and Hogan, 1998). Bmp7 protein is present in the head ectoderm at the time of lens placode induction. Inhibition of Bmp7 signaling at the time of lens placode induction significantly decreases the frequency of lens formation in an organ culture system. The expression of the lens placode marker Sox2 was also severely affected in Bmp7-/- mutant embryos (Wawersik, 1999).

Fibroblast Growth Factor Signaling

In the chick, several observations implicate Fgfs as one of the key factors initiating the formation of the border region: misexpression of Fgf8 induces ectopic expression of neural plate border-specific genes (Streit and Stern, 1999; Litsiou et al., 2005). However, in chich and frogs Fgf alone is not sufficient to generate neural crest and placode fates (Mayor, et al., 1997; LaBonne and Bronner- Fraser, 1998; Monsoro-Burq, et al., 2003; Ahrens and Schlosser, 2005; Litsiou, et al., 2005; Hong et al., 2008). There is good evidence that Fgf8 may be required in concert with Bmp signaling to promote placodal fate. In Xenopus, Fgf8 positively regulates expression of the pre-placodal specific gene Six1 when Bmp is inhibited in the ectoderm (Ahrens and Schlosser, 2005). Fgf8 is expressed in the paraxial mesoderm and anterior neural ridge in frogs (Christen and Slack, 1997), and morpholino-mediated knockdown of Fgf8a results in a broad loss of neural crest and pre-placodal genes (Hong and Saint-Jeannet, 2007). However, the loss neural crest in these experiments is likely to be indirect since Fgf8a is required to activate Wnt8 in the mesoderm (Hong et al., 2008). As described above, in chick embryos Fgf8 is also hypothesized to be the factor required to repress lens fate in the pre-placodal ectoderm and to promote specification of olfactory cells (Bailey et al., 2006).

Several members of the Fgf family are implicated in otic placode specification (reviewed in Schimmang 2007; Schneider-Maunoury and Pujades, 2007). Different species appear to use different combinations of these molecules. Fgf19 expressed in the paraxial mesoderm and Fgf8 derived from the endoderm, are implicated in otic placode induction in the chick (Ladher et al., 2000; Ladher et al., 2005). In zebrafish otic placode induction depends primarily on Fgf3 and Fgf8 (Philipps et al., 2001; Maroon et al., 2002; Leger and Brand, 2002; Liu et al., 2003). In the mouse, paraxial mesoderm-derived Fgf8 and Fgf10 are involved in this process (Wright and Mansour, 2003; Ladher et al., 2005; Zelarayan et al., 2007).

A role for Fgf3 and Fgf8 ligands is reported in epibranchial placode specification in zebrafish (Nechiporuk, et al., 2007; Nikaido, et al., 2007; Sun, et al., 2007). In the Fgf8 mutant embryos, acerebellar (ace), placodal expression of Sox3 was disrupted. This phenotype was rescued by implantation of an Fgf8 bead near the prospective hindbrain. This requirement for Fgf signaling was further demonstrated using a soluble FgfR inhibitor, which resulted in reduced expression of the epibranchial markers Sox3 and Phox2a (Nikaido, et al., 2007). In contrast, other experiments using morpholinos against both Fgf3 and Fgf8 suggested a dual requirement of Fgf3 and Fgf8, rather than Fgf8 alone (Sun et al., 2007; Nechiporuk et al., 2007).

Fgf3 is prodiced by the ventral diencephalon and is required for the expression of early adenohypophysis markers (Herzog et al., 2004). In the mouse, loss of FgfR2b or deletion of its ligand, Fgf10, leads to early defects in the adenohypophysis (Ohuchi et al., 2000) also suggesting an important function of Fgf signaling in adenohypohysis placode formation.

Wnt Signaling

At the neural plate border, canonical Wnt signaling is required in conjunction with Bmp attenuation to specify the neural crest (reviewed in Knecht and Bronner-Fraser, 2002; Huang and Saint-Jeannet, 2004). Interestingly, the pre-placodal ectoderm appears to have a different requirement with regard to Wnt signaling compared to the neural crest, because inhibition of the canonical Wnt signaling pathway favors placodal tissue at the expense of neural crest fate (Brugmann et al., 2004; Litsiou et al., 2005). Recent work using Xenopus animal caps suggest that inhibition of canonical Wnt signaling is more specifically required to define the most anterior domain (olfactory) of the pre-placodal ectoderm (Park and Saint-Jeannet, 2008). Expression of Fgf8a promotes olfactory placode formation (based on Dmrt4 expression) in animal caps injected with Noggin. However, simultaneous activation of canonical Wnt signaling in these explants inhibited Dmrt4 expression and promoted otic placode fate based on Pax8 expression. These results indicate that cranial placodes along the antero-posterior axis have different requirements with regard to Wnt signaling. The most anterior cranial placode (olfactory, at least) requires Wnt inhibition while more posterior placodes (otic) depend on active canonical Wnt signaling (Park and Saint-Jeannet, 2008).

There is also strong evidence in multiple species that canonical Wnt signaling is involved in otic placode induction. In chick, presumptive otic ectoderm showed a stronger induction of the otic marker gene Pax2 when cultured in the presence of Fgf19 and Wnt8C compared to explants cultured with Fgf19 alone (Ladher et al., 2000). A subsequent study in zebrafish showed that Wnt8 depletion or overexpression of a Wnt inhibitor (Dkk1) did not prevent otic vesicle formation (Phillips et al., 2004). Mouse work suggests that the presumptive otic ectoderm is exposed to Wnt signals early, as the activity of a TCF/Lef-LacZ reporter is detected in the pre-otic ectoderm (Ohyama et al., 2006). Conditional knockout of β-catenin results in a smaller than normal otic vesicle, and conditional stabilization of β-catenin expands the placodal domain. In the mouse canonical Wnt signaling is likely to mediate the decision between otic and epidermal fate (Ohyama et al., 2006; 2007). In Xenopus, canonical Wnt signaling appear to cooperate with Fgf signals (Fgf3 and Fgf8) to specify the otic placode (Park and Saint-Jeannet, 2008).

Wnt molecules are also implicated in trigeminal placode formation in the chick embryo. Blocking canonical Wnt signaling prevented the targeted cells to adopt or maintain an ophthalmic trigeminal placodal fate, based on the expression of Pax3 and Eya4. In contrast, activation of the Wnt pathway was not sufficient to elicit Pax3 expression, suggesting that other signaling cues are also required to promote ophthalmic trigeminal placodal fate (Lassiter et al., 2007; Dude et al., 2009).

Other Signaling Pathways

In addition to the three major signaling pathways discussed above other signaling molecules are also implicated in various aspects of placode development. While very little is known about the factors inducing lateral line placodes in amphibia and fish, recent studies are focuse on the signaling molecules regulating posterior lateral line primordium migration in zebrafish. Chemokines of the CXCL class and their receptors (CXCR) are known for promoting the directional migration of leukocytes during inflammation (Zlotnik and Yoshie, 2000). In zebrafish, this signaling pathway is essential to provide directionality during migration of the posterior lateral line primordium. Moreover the cooperation between chemokine, Wnt and Fgf signaling appears to regulate the mechanisms by which the primordium maintains its integrity during migration despite the periodic deposition of cells during formation of the lateral line system (Dambly-Chaudiere et al., 2007; Aman and Piotrowski, 2008; Nechiporuk and Raible, 2008). The same class of molecules is also implicated in olfactory placode development. In zebrafish, Cxcl12/Cxcr4 signaling mediates assembly of olfactory placodal precursors into a compact cluster to form the olfactory placode (Miyasaka et al., 2007). Later in development, Cxcr4-mediated chemokine signaling is required for assembling the trigeminal sensory neurons into a ganglion (Knaut et al., 2005).

Platelet Derived Growth Factor (Pdgf) signaling is implicated in the induction of the ophthalmic lobe of the trigeminal placode in the chicken (McCabe and Bronner-Fraser, 2008). Pdgf receptor β is detected in the cranial ectoderm at the time of trigeminal placode formation, and the ligand Pdgfd is expressed in neural folds of the midbrain. In recombinants explants of quail ectoderm with chick neural tube, which normaly promote trigeminal placode fate, blocking Pdgf signaling results in loss of Pax3 and CD151 expression, two early markers of the trigeminal placode. Conversely, microinjection of exogenous Pdgfd increases the number of Pax3-expressing cells in the trigeminal placode and neurons in the condensing ganglia.

Shh is one of the primary factors involved in adenophypophysis placode induction. During development Shh is expressed throughout the oral ectoderm, but it is excluded from Rathke’s pouch as soon as this structure forms. In Shh-deficient mouse embryos, formation of the diencephalon is severely disrupted which has made it difficult to assess adenohypophysis development. However, Rathke’s pouch formation was completely arrested in transgenic animals expressing a specific hedgehog inhibitor (Hip) throughout the oral ectoderm (Treier et al., 2001). In the talpid3 chicken mutant Shh signaling is reduced, and formation of the pituitary is severely disrupted (Lewis et al., 1999). Large ectopic lenses form as diverticula from the roof of the mouth, at a position corresponding to the adenohypophyseal placode in normal embryos (Ede and Kelley, 1964). The zebrafish double mutants for shh and tiggy winkle hedgehog (twhh), which encode partially redundant hedgehog ligands, have a complete loss of anterior pituitary fates (Herzog et al., 2003). In contrast overexpression of shh causes induction of an excessive number of pituitary cells at the expense of lens precursors (Dutta et al., 2005; Herzog et al., 2003; Sbrogna et al., 2003; reviewed in Pogoda and Hammerschmidt, 2009)

Retinoic acid (RA) is primarily implicated in morphogenesis and patterning of the otocyst. Its activity is presumably indirect mediated through its ability to regulate several target genes in the hindbrain, among which are Hox genes that provide rhombomere identity in the hindbrain (reviewed in Romand et al., 2006). In zebrafish, application of a dose of retinoic acid that does not perturb patterning of the anterior neural plate leads to increased otic induction, a process dependent on Fgf signaling (Hans et al., 2007). In vitamin A-deficient quail embryos, Rathke’s pouch fails to develop, suggesting an important role of retinoic acid in adenohypophyseal placode development in birds. However, this phenotype might be secondary to the loss of other signaling molecules since Bmp2, Shh and Fgf8 are downregulated in these animals (Maden et al., 2007).

Image fig1
Copyright © 2010 by Morgan & Claypool Life Sciences.
Bookshelf ID: NBK53176

Views

  • PubReader
  • Print View
  • Cite this Page

Other titles in this collection

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...