Fig. 3 S1

Vegfr2 is responsible for Ca²⁺ responses before ISVs sprouting from the DA.

(A) 3D-rendered time-sequential images of Tg(fli1:Gal4FF);(UAS:GCaMP7a);(fli1:H2B-mC) embryos treated with ki8751 to inhibit Vegfr before ISV sprouting. The embryos were treated with ki8751 from 17 ss and time-lapse imaged at 18 ss using a light sheet microscope. Yellow dashed lines indicate positions of somite boundaries. (B) The DA is subdivided into three regions (Region 1–3) between two somite boundaries (SBs) as illustrated in the schematics (left). The fluorescence changes in GCaMP7a (ΔF/F₀) of individual ECs from A are shown as separated graphs (Region 1–3) as in Figure 3B. Where individual EC belong to is determined by the location based on the position of the nucleus at the start of time-lapse imaging. A representative graph of two ECs at each region is shown. (C) 3D-rendered time-sequential images of Tg(fli1:Gal4FF);(UAS:GCaMP7a);(fli1:H2B-mC) embryos before ISV sprouting injected with vegfr2 (kdrl) morpholino (MO) (18 ss). (D) The fluorescence changes in GCaMP7a (ΔF/F₀) of individual ECs from C are shown as separated graphs (Region 1–3) as in B. (E) 3D-rendered time-sequential images of Tg(fli1:Gal4FF);(UAS:GCaMP7a) embryos before ISV sprouting injected with vegfr2 MO and UAS:Vegfr2,NES-mC plasmid which drives the expression of full length Vegfr2 and NES-mC simultaneously in ECs in a mosaic manner (19 ss). Note that Ca²⁺ oscillations are recovered in NES-mC-positive ECs (arrowheads). (F) The fluorescence changes in GCaMP7a (ΔF/F₀) of individual ECs expressing NES-mC from E indicated by arrowheads (green, blue, orange and magenta) at the left panel are shown as a graph. Scale bars, 10 µm in A, C and E. DA, dorsal aorta; SB, somite boundary.