H_(2)S promotes developmental brain angiogenesis via the NOS/NO pathway in zebrafish

Background Hydrogen sulphide(H_(2)S)is considered as the third member of the gasotransmitter family,along with nitric oxide(NO)and carbon monoxide.H_(2)S has been reported to induce angiogenesis by promoting the growth,migration and tube-like structure formation of endothelial cells.Those studies were conducted in conditions of cell culture,mouse Matrigel plug assay model,rat wound healing model or rat hindlimb ischaemia model.Recent in vivo studies showed the physiological importance of H_(2)S in muscle angiogenesis.However,the importance of endogenous H_(2)S for brain angiogenesis during development remains unknown.We therefore aimed at determining the role of H_(2)S in brain vascular development.Methods and results Both knockdown and knockout of H2S-producing enzymes,cystathionineβ-synthase(cbs)and cystathionineγ-lyase(cth),using morpholino oligonucleotides and clustered regularly interspaced short palindromic repeats/Cas9-mediated mutation,impaired brain vascular development of larval zebrafish.Incubation with the slow-releasing H_(2)S donor GYY4137 alleviated the defects of brain vascular development in cbs and cth morphants.Quantitative analysis of the midbrain vascular network showed that H_(2)S enhances angiogenesis without affecting the topological structure of the brain vasculature.Mechanically,nitric oxide synthase 2a(nos2a)expression and NO production were decreased in both cbs and cth morphants.Overexpression of nos2a by coinjection of cbs or cth MO with full-length zebrafish nos2a mRNA alleviated the brain vascular developmental defects in cbs and cth morphants.Conclusion We conclude that H_(2)S promotes brain developmental angiogenesis via the NOS/NO pathway in zebrafish.