Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

Placodes are ectodermal thickenings of the embryonic vertebrate head. Their descendants contribute to sensory organ development, but also give rise to sensory neurons of the cranial nerves. In mammals, the signaling pathways which regulate the morphogenesis and neurogenesis of epibranchial placodes, localized dorsocaudally to the pharyngeal clefts, are poorly understood. Therefore, we performed mouse whole embryo culture experiments to assess the impact of pan-fibroblast growth factor receptor (FGFR) inhibitors, anti-FGFR3 neutralizing antibodies or the pan-bone morphogenetic protein receptor (BMPR) inhibitor LDN193189 on epibranchial development. We demonstrate that each of the three paired epibranchial placodes is regulated by a unique combination of FGF and/or bone morphogenetic protein (BMP) signaling. Thus, neurogenesis depends on fibroblast growth factor (FGF) signals, albeit to different degrees, in all epibranchial placodes (EP), whereas only EP1 and EP3 significantly rely on neurogenic BMP signals. Furthermore, individual epibranchial placodes vary in the extent to which FGF and/or BMP signals (1) have access to certain receptor subtypes, (2) affect the production of Neurogenin (Ngn)2+ and/or Ngn1+ neuroblasts, and (3) regulate either neurogenesis alone or together with structural maintenance. In EP2 and EP3, all FGF-dependent production of Ngn2+ neuroblasts is mediated via FGFR3 whereas, in EP1, it depends on FGFR1 and FGFR3. Differently, production of FGF-dependent Ngn1+ neuroblasts almost completely depends on FGFR3 in EP1 and EP2, but not in EP3. Finally, FGF signals turned out to be responsible for the maintenance of both placodal thickening and neurogenesis in all epibranchial placodes, whereas administration of the pan-BMPR inhibitor, apart from its negative neurogenic effects in EP1 and EP3, causes only decreases in the thickness of EP3. Experimentally applied inhibitors most probably not only blocked receptors in the epibranchial placodes, but also endodermal receptors in the pharyngeal pouches, which act as epibranchial signaling centers. While high doses of pan-FGFR inhibitors impaired the development of all pharyngeal pouches, high doses of the pan-BMPR inhibitor negatively affected only the pharyngeal pouches 3 and 4. In combination with partly concordant, partly divergent findings in other vertebrate classes our observations open up new approaches for research into the complex regulation of neurogenic placode development.

The Role of the Placodes in the Development of the Glossopharyngeal, Vagal, and Trigeminal Ganglia

The epibranchial placodes combine with the neural crest to form the inferior and superior ganglia of the glossopharyngeal and vagal cranial nerves, respectively. By comparison, the single trigeminal ganglion is composed of both neural crest and placodal cells. The steps that lead up to these events include gastrulation and the embryology of the notochord, neural crest, and the placodes. Each of these steps is reviewed in some detail. In previous reviews in this series, the embryology related to the olfactory, otic, and lens placodes, and to the geniculate ganglia has been discussed.1-3 However, the somewhat unusual embryology of the 2 ganglia of cranial nerves IX and X was only briefly mentioned as was the development of the trigeminal ganglion.4 This present review revisits these events and specifically focuses on how these ganglia develop.Learning Objective: The reader will learn the unusual development of the superior and inferior glossopharyngeal and the vagal ganglia as well as review the varied steps in the embryology that proceeds these events. By comparison, the development of the single trigeminal ganglion is presented and the differences in its development from that of the ganglia of cranial nerves IX and X are emphasized.

An Eya1-Notch axis specifies bipotential epibranchial differentiation in mammalian craniofacial morphogenesis

Craniofacial morphogenesis requires proper development of pharyngeal arches and epibranchial placodes. We show that the epibranchial placodes, in addition to giving rise to cranial sensory neurons, generate a novel lineage-related non-neuronal cell population for mouse pharyngeal arch development. Eya1 is essential for the development of epibranchial placodes and proximal pharyngeal arches. We identify an Eya1-Notch regulatory axis that specifies both the neuronal and non-neuronal commitment of the epibranchial placode, where Notch acts downstream of Eya1 and promotes the non-neuronal cell fate. Notch is regulated by the threonine phosphatase activity of Eya1. Eya1 dephosphorylates p-threonine-2122 of the Notch1 intracellular domain (Notch1 ICD), which increases the stability of Notch1 ICD and maintains Notch signaling activity in the non-neuronal epibranchial placodal cells. Our data unveil a more complex differentiation program in epibranchial placodes and an important role for the Eya1-Notch axis in craniofacial morphogenesis.