Three-dimensional studies of Prop1-expressing cells in the rat pituitary primordium of Rathke's pouch

Pituitary embryonic development progresses daily toward terminal differentiation exhibiting quantitative and qualitative alterations regulated by signal molecules and transcription factors expressed under temporospatial control. In this study, we analyzed the heterogeneity of the cells in the pituitary primordium of embryonic day (E) 13.5. The three-dimensional structure of the Rathke's pouch was built up from measurements taken from multiple DAPI-stained sections and cell populations positive to stem/progenitor marker SOX2 and pituitary-specific transcription factor PROP1 were analyzed. The pituitary primordium (Rathke's pouch) of E13.5 showed a flattened discoid shape of about 500 μm in diameter and 200 μm depth in a dorsoventral axis and consisted in about 5,800 cells. Immunohistochemistry revealed that 0.3% of the cells in Rathke's pouch were SOX2-negative in the lateral region, whereas all cells at E12.5 were SOX2-positive. On E13.5, the shape and size of their nuclei showed a location-specific divergence: ellipsoid morphology in the median region and round morphology in the lateral region. Moreover, on E14.5, adrenocorticotropic-hormone-positive cells (the first hormone-producing cells appearing in the pituitary) contained round nuclei. These data suggest that differentiation to pituitary-hormone-producing cells from SOX2-negative cells starts in the lateral region between E12.5 and E13.5 and that the onset of differentiation is preceded by a change in nuclear shape.