The early onset of Prox1 expression is revealed by β-galactosidase expression and in situ hybridization.
Whole mount β-galactosidase histochemcial reaction using X-Gal was performed in Prox1 heterozygous and nullizygous embryos. A. Starting at E11.0, a progressive upregulation of Prox1 is seen in the anterior (AC) and posterior (PC) canal cristae. B. By E13.5, expression is also detected in the horizontal canal crista (HC), the striolar region of the utricle (U), the canals and the endolymphatic duct (ED); expression in the saccule is barely detected (S). In the cochlea, upregulation of β-galactosidase expression is detected in the apex and decreases toward the base. Arrows indicate expression in anterior and posterior canal with their expression. C. Expression of β-galactosidase is identical in heterozygous and nullizygous mice with the exception that the signal is stronger in nullizygous mice. Faint β-galactosidase expression is also detected in the delaminating spiral ganglion neurons (SPG; C and insert in B,C). D. In situ hybridization shows at E14.5 expression in the canal cristae and the cochlea, but indicates a more prominent upregulation in the base at this stage. Only spiral ganglion sensory neurons are faintly positive for Prox1 in situ (SPG in D). E,F At postnatal stages, Prox1 expression remains in the canal cristae as revealed by in situ hybridization for Prox1 mRNA or X-Gal reaction, but does not show the extensive expression in the non-sensory parts of the canals as in earlier stages (insert in F). Bar, 100 µm.