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First published online 21 September 2005
doi: 10.1242/dev.02052

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Lack of an adrenal cortex in Sf1 mutant mice is compatible with the generation and differentiation of chromaffin cells

Philipp Gut^1,*, Katrin Huber^1,*,, Jennifer Lohr¹, Barbara Brühl¹, Stephan Oberle¹, Mathias Treier², Uwe Ernsberger¹, Chaya Kalcheim³ and Klaus Unsicker¹

¹ Neuroanatomy and Interdisciplinary Center for Neurosciences (IZN), University of Heidelberg, INF 307, D-69120 Heidelberg, Germany
² European Molecular Biology Laboratory, Meyerhofstrasse 1, D-69117 Heidelberg, Germany
³ Department of Anatomy and Cell Biology, Hebrew University-Hadassah Medical School, Jerusalem 91120-PO Box 12272, Israel

View larger version (135K): [in a new window]   Fig. 1. Adrenal cortical cells are labelled by Sf1 in-situ hybridization on a transverse section (dorsal, top) of an E13.5 wild-type embryo (A). Sf1 expression is completely lacking in Sf1–/– littermates (B). In-situ hybridization for NF mRNA (blue) and TH immunofluorescence staining (red overlay) have been performed on a consecutive section of the wild-type (C; E, higher magnification) and the Sf1–/– embryo (D; F, higher magnification). Note that in Sf1–/– embryos TH+/NF– `adrenal' chromaffin cells accumulate at a location where the adrenal gland develops in wild-type mice. Scale bars: 100 µm. a, dorsal aorta; gn, gonadal anlage; sg, sympathetic chain ganglion; srg, suprarenal ganglion.

View larger version (135K): [in a new window]   Fig. 2. In-situ hybridization for Sf1 on transverse sections (dorsal, top) of an E17.5 wild-type embryo (A) and an Sf1–/– littermate (B) at the axial level of the adrenal gland. In-situ hybridization for NF mRNA (blue) and TH immunofluorescence staining (red overlay) have been performed on a consecutive section of the wild-type (C; E, higher magnification) and the Sf1–/– embryo (D; F, higher magnification). Note that in the Sf1–/– embryo TH+/NF– `adrenal' chromaffin cells are still present. Scale bars: 100 µm. a, dorsal aorta; srg, suprarenal ganglion.

View larger version (123K): [in a new window]   Fig. 3. SCG10 is expressed in the sympathetic suprarenal ganglion (srg) as shown by in-situ hybridization, but is lacking in chromaffin cells (red demarcation) in both wild-type (A) and Sf1–/– (B) mice. Chromaffin cells of both wild-type (C) and Sf1–/– (D) mouse embryos are immunoreactive for VMAT1. Only very few scattered cells express VMAT1 in the sympathetic ganglion in wild-type (not shown) and Sf1–/– mice (black demarcation). Transverse sections are shown (dorsal, top). Scale bars: 100 µm. srg, suprarenal ganglion.

View larger version (128K): [in a new window]   Fig. 4. Semithin transverse sections (dorsal, top) at the axial level of the adrenal gland of an E17.5 wild-type embryo (A) and an SF–/– littermate (B), showing the position of the right adrenal gland, the suprarenal ganglion, the upper pole of the kidney, the liver and the crus dextrum of the diaphragm. Electron micrographs have been made on tissues corresponding to the demarcated areas: red (C,D) and black (E,F). Cells with the typical ultrastructural features of chromaffin progenitor cells, which contain large dense core vesicles (arrowheads), are present in wild-type (C) and knockout (D) embryos in the areas that have been demarcated with red dashed lines (A,B). Electron micrographs (E,F) from the areas demarcated with black dashed lines (A,B) show typical sympathetic neurons of the suprarenal ganglion, which appear identical in wild-type (E) and Sf1–/– (F) embryos. Scale bars: 100 µm in A,B; and 2 µm in C-F. ag, adrenal gland; cp, chromaffin progenitor cells; d, crus dextrum of the diaphragm; k, upper pole of kidney; lv, liver; srg, suprarenal ganglion.

View larger version (100K): [in a new window]   Fig. 5. Electron micrographs of `adrenal' chromaffin tissue of an E17.5 Sf1–/– embryo showing axon terminals (arrow) abutting chromaffin cells (A) and a mitotic cell (B). Scale bars: 1 µm.

View larger version (62K): [in a new window]   Fig. 6. In-situ hybridization for SF1 (A,E), TH (B,F), PNMT (C,G) and RET (D,H) on transverse sections (dorsal, top) of the adrenal gland of E19.5 wild-type embryos (A,B,C,D) and sections of `adrenal' chromaffin tissue (red demarcation) of an Sf1–/– littermate (E,F,G,H). Note that PNMT expression is lacking in the Sf1–/– embryo (G), while RET expression is maintained (H). Scale bar: 100 µm.

View larger version (14K): [in a new window]   Fig. 7. (A) Quantification of TH+/NF– `adrenal' chromaffin cells in E13.5 and 17.5 Sf1+/+ and Sf1–/– embryos. Data are presented as mean ± s.e.m. Six `adrenal glands' from three different animals were analysed per group. Every second section was counted. (B) BrdU+/TH+ adrenal chromaffin cells in Sf1 mutant embryos and wild-type littermates at E13.5 expressed as percentage of total TH+ cells. BrdU was administered to pregnant females at day 12.5 of gestation. Data are presented as mean±s.e.m. Six `adrenal glands' from three different animals were analysed per group. Every fifth section was counted.

View larger version (36K): [in a new window]   Fig. 8. (A) Quantification of SOX10-ir, TH-ir and PHOX2B mRNA-expressing cells in the region of the developing adrenal gland in E12.5 wild-type and Sf1 mutant mice. The precise demarcations of the region analysed are described in the text. Every fifth section was counted. Four animals were analysed per group. Data are presented as mean±s.e.m. (B,C) TH (red) and SOX10 (green) double-immunolabelling on transverse sections (dorsal, top) through the region of the developing adrenal gland in wild-type and Sf1–/– mouse embryos. The area in which cells were counted is demarcated.