Sensory neuron differentiation is regulated by notch signaling in the trigeminal placode

Abstract

Trigeminal sensory neurons develop from the neural crest and neurogenic placodes, and have been studied as a principle model of sensory neuron formation. While the Notch pathway has been extensively characterized in central nervous system development and other developmental processes, it has not been well characterized in sensory neurogenesis. Here we studied the functional role of Notch signaling in the trigeminal ophthalmic (opV) placode, a prime model of sensory neurogenesis. To establish a good spatiotemporal description of Notch pathway genes in the chick trigeminal placode, a stage-specific expression analysis was conducted, showing that expression of most Notch pathway genes and effectors are expressed in the placode, with expression primarily being confined to ectodermal cells. Expression was highest at stages of peak neuronal differentiation. To test the function of Notch signaling in opV placode cell differentiation, Notch receptor cleavage was blocked using the gamma-secretase inhibitor, DAPT, or signaling was activated by misexpression of the Notch intracellular domain (NICD). Notch activation resulted in a significant reduction in sensory neurogenesis. Cells remained in the ectoderm and did not differentiate. Expression of the opV specification marker Pax3 was also lost in targeted cells. DAPT exposure resulted in a dramatic increase in neurogenesis without increasing proliferation, where many differentiated cells were found in the mesenchyme and, surprisingly, within the ectoderm. This is the first result clearly showing prolific neuronal differentiation in the ectoderm of the trigeminal placodes after experimental manipulation of a molecular signaling pathway, thus identifying Notch signaling as a primary regulator of the sensory neuron fate in the opV placode.

Notch pathway regulation of neural crest cell development in vivo

“…The reduction in glial cells in Wnt1Cre;RBP-J f/f mice suggests that endogenous canonical Notch signaling is required to promote gliogenesis in the DRG, consistent previous reports (Taylor et al, 2007; Hu et al, 2011). In cultured avian NCCs, Notch activation suppresses neurogenic differentiation of NCCs and accelerates glial differentiation via promotion of Sox10 expression, but this has not been demonstrated previously in vivo (Lassiter et al, 2010; Morrison et al, 2000; Wakamatsu et al, 2000). Our data demonstrate that increased Notch signaling in NCCs promotes gliogenesis through the promotion of Sox10, while inhibiting neurogenesis in cranial ganglia and DRG, in vivo in mice.…”

Signaling mechanisms controlling cranial placode neurogenesis and delamination

“…Neurogenesis begins within these restricted zones and the primary morphological event of the placode is delamination of neuroblasts from the specified epithelium (Graham et al, 2007; Lassiter et al, 2010; McCabe et al, 2009). Sensory neurons are derived from both the cranial placodes and the neural crest migratory cell populations; however, placodal delamination differs markedly from that of neural crest.…”

“…The timing of terminal differentiation occurs within the epithelium prior to delamination, as seen in the ophthalmic trigeminal (McCabe et al, 2009), or shortly after delamination during their migration, observed in mmV, epibranchial, and otic neurons (Begbie et al, 2002; Blentic et al, 2011). Interestingly, epibranchial placode cells appear to be mitotically quiescent during the delamination process (Graham et al, 2007), while neuronal differentiation of placode-derived cells initiates prior to migration (Blentic et al, 2011; Lassiter et al, 2010). Placode neurogenesis begins within the epithelial niche and these cells are committed as neurons upon delamination from the ectoderm, indicating that neuronal cell selection and changes in cell adhesion leading to delamination may be coupled.…”

“…In the trigeminal placode, Notch LOF carried out in chick head explant cultures resulted in a dramatic increase in neurogenesis within placodal epithelium and in the mesenchyme, in both the opV and mmV trigeminal placodes (Lassiter et al, 2010). Inhibition of Notch led to a substantial increase of ectodermal cells expressing Isl1 and NF, as well as a vast sheet of neuroblasts delaminating from the epithelium with an abundant amount of differentiating neurons also observed in the mesenchyme.…”

“…Interestingly, this significant enhancement in neurogenesis and delamination did not extend beyond the normal specified placode domain suggesting that Notch signaling is regulating a neuronal fate choice at this developmental time point and is not involved in specifying the competent neurogenic domain. In the trigeminal placodes, blocking Notch activity with the gamma-secretase inhibitor DAPT resulted in precocious neurogenesis, including ectopic neuronal differentiation within the epithelium (Lassiter et al, 2010). The spatiotemporal expression patterns of Delta/Notch pathway components were also described in this study, and most expression was observed only within the placodal ectoderm and not in migratory cells.…”