Table 1.
Fig 1.
CART is expressed throughout normal pituitary development.
CART protein is not detected in e12.5 pituitary gland at e12.5 (A). PROP1 protein is detected in Rathke’s pouch of the e12.5 pituitary gland (B). CART protein was detected in paraffin sections of pituitary glands with a specific antibody (green fluorophor) in Prop1+/+ (C, E, G) and Prop1df/df (D, F, H) anterior pituitary lobes at e14.5 (C, D), e16.5 (E, F), and P1 (G, H). At P8 CART is expressed abundantly in the anterior pituitary lobe (I), but it is absent in the Prop1df/df anterior pituitary lobe (J). CART protein is present in the Pou1f1+/+ (normal) (K) and Pou1f1dw/dw anterior lobe of the pituitary gland (L). Images A-F were taken at 200X and scale bar is 200 μm. Images G-L were taken at 100X and scale bar is 200 μm.
Fig 2.
CART expression the pituitary hormones.
Co-localization of CART with specific pituitary hormones was determined with a CART-specific antibody (green) and hormone specific antibodies (red) in sections from pituitary glands collected at P8. The hormone specific antibodies were TSHb (A), GH (B), ACTH (C), FSHb (D), LHb (E), and PRL (F). Co-localization is indicated with yellow arrows (A, B). In one CART immunostaining experiment all hormone antibodies were applied together and detected with the same fluorophore (G, H). A few CART positive cells do not express a detectable level of hormone (green arrow) in the Pou1f1+/+ pituitary (G), but almost all CART positive cells do not co-localize with any hormones in the somatotrope deficient Pou1f1dw/dw anterior pituitary (H). The CART protein (red) does not co-localize with PROP1 (green) in the developing pituitary gland at e14.5 (I). Images A-I were taken at 400X and scale bar is 100 μm.
Fig 3.
CART is co-expressed with several cell specific transcription factors.
Immunostaining with antibodies specific for CART (red, cytoplasmic) and lineage specific transcription factors (green, nuclear) was carried out on paraffin sections of pituitaries collected at P8 from Pou1f1+/+ mice (A, C, E) and Pou1f1dw/dw mutants (B, D, F). The lineage specific transcription factors were POU1F1 (A, B), TPIT (C, D), and NR5A1 (E, F). All images were taken at 400X and scale bar is 100 μm.
Fig 4.
CART is expressed in the proliferating cells of the pituitary gland.
Immunohistochemical staining was used to detect CART (red) and the proliferative markers Ki67 (green) or BrdU (green) in sections of pituitaries collect from Pou1f1+/+ and Pou1f1dw/dw mice at P8. Co-localization is indicated by white arrows, and CART immunofluorescence in Ki67 or BrdU negative cells is indicated with arrow heads. There are many CART cells in both pituitary genotypes that do not co-localize with any proliferating cells (arrow heads). All images were taken at 400X, and scale bar is 100 μm.
Fig 5.
CART co-localizes with thyrotropes in the adult pituitary.
Immunohistochemical staining was used to detect co-localization of CART (green) and hormones (red) in sections prepared from adult pituitary glands. The hormone specific antibodies were GH (A), PRL (B), TSHβ (C), LHβ (D), ACTH (E), and FSHβ (F). Representative cells (Inset panel C) exhibited co-localization of CART with TSHβ (yellow arrow), or lacked TSH expression (green arrow), and some TSHβ positive cells lacked CART staining (red arrow). All images were taken at 200X and scale bar is 100 μm. Inset of image C was taken at 400X and scale bar is 100 μm.