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Fig. 7
Spatio-temporal fate map of the larval prethalamic eminence/prospective vENT and adjacent areas. Images and accompanying schematics show Kaede photoconversion areas in lateral views of 48 hpf Tg(lhx5:Kaede)b1204 fish (A,A′,C,C′,E,E′) and location of Kaede cells by 72 hpf in lateral view (B-B′′,D-D′′,F-F′′) and transverse sections after 3D rendering (B′,D′,F′). Dotted arch and lines at 48 hpf mark the dorsal limit of the preoptic area and prethalamic eminence respectively. Photoconversion in the prethalamic eminence (n = 8). (A-B′′) Cells were photoconverted from green to red (magenta in the images) at the diencephalic prethalamic eminence of Tg (lhx5:Kaede)b1204 embryos at 48 hpf (arrowhead in A, red cells in schematic A′′) and their location was then imaged by 72 hpf in lateral view (B and red cells in B′′). Note in (A) that area of photoconversion by 48 hpf is located at the level of the tract of the habenular commissure (marked with a star). (B′) is transverse section after 3D rendering. Note that 24 h after photoconversion, red Kaede expressing neurons have expanded from the photoconversion area rostrally into the telencephalon (yellow arrow) and caudally into hypothalamic areas (white arrow). These red Kaede cells send afferent projections to ventral (arrow in B inset) and dorsal halves (arrowhead in B inset) of the habenula and were identified as vENT. Transverse section shows that the vENT occupies a lateral position within the forebrain (B′). Photoconversion in preoptic regions (PO) (n = 9). (C-D′′) Cells were photoconverted in the presumed preoptic area of Tg (lhx5:Kaede)b1204 embryos at 48 hpf (C,C′). Note that 24 h after photoconversion, most red Kaede cells maintain their relative rostro-caudal positions within the telencephalon and diencephalon (D), and are located ventro-medially relative to the vENT when viewed in transverse section (D′). Most conversions of presumptive PO (n = 6/9) did not result in labeling of habenula afferents. Photoconversion of the prethalamus (PTh; n = 3). (E-F′′) Cells were photoconverted in the presumed prethalamic area of Tg (lhx5:Kaede)b1204 embryos at 48 hpf (E,E′). Twenty four hours after photoconversion, most red Kaede cells maintain their positions within the diencephalon and do not expand into the telencephalon (F). A few red Kaede cells expand into caudal areas (white arrow). Transverse section shows that these cells occupy a dorsomedial position (F′). (G) Summary of the results of the fate map experiments with Tg(lhx5:Kaede)b1204, with the areas of Kaede photoconversion in 48 hpf. Open circles denote cells that were not fate mapped. Level of transverse section is shown in the 72 hpf lateral view of the brain. Scale bars: All 100 µm.