Wnt/β-catenin signaling regulates VE-cadherin-mediated anastomosis of brain capillaries by counteracting S1pr1 signaling

Canonical Wnt signaling is crucial for vascularization of the central nervous system and blood-brain barrier (BBB) formation. BBB formation and modulation are not only important for development, but also relevant for vascular and neurodegenerative diseases. However, beyond the early requirement of Wnt signaling for brain capillary development, there is little understanding of how Wnt signaling further contributes to brain angiogenesis and BBB formation. By combining high resolution in vivo imaging with temporally and spatially controlled manipulation of Wnt signaling, we were able to dissect different requirements for Wnt signaling during brain angiogenesis and BBB formation. In the absence of Wnt signaling, premature Sphingosine-1-phosphate receptor (S1pr) signaling leads to a reduction of VE-cadherin and Esama at cell-cell junctions. Wnt signaling most likely suppresses S1pr signaling during angiogenesis to enable the dynamic junction formation during anastomosis, whereas later S1pr signaling regulates BBB maturation and VE-cadherin stabilization. Our data provides a novel link between brain capillary angiogenesis and BBB formation and identifies Wnt signaling as coordinator of the timing and as regulator of anastomosis. Isolation of the zebrafish CtA endothelial cells from 36hpf-old Tg(fli1a:Gal4)ubs3;(UAS:Kaede)rk8 embryos treated with IWR-1 or DMSO