Neural Innervation Invigorates Yolk Sac Biological Functions beyond Nutrient Reservoir during Zebrafish Embryo Development

The zebrafish yolk sac (YS) is traditionally viewed as a nutrient reservoir. By reconstructing the complete progression of embryonic neural development via live-cell imaging, we previously uncovered an intriguing involvement of YS after it is innervated by the neurons from the brain, hinting its uncharacterized functional roles beyond nutrient storage. Using transgenic lines and long-term live imaging, we characterized a dynamic neuro-vascular-metabolic interface on the YS surface. We observed that peripheral neural networks expand radially and mature through hierarchical integration, sharing the same structural and dynamical features as those in the brain and spinal cord. To many unexpected, elavl3-positive cells on the YS exhibit collective calcium flashes, suggesting primitive functional communication. Furthermore, we characterized activity-dependent neuronal pruning and stress-induced lipid droplet crystallization as indicators of developmental refinement and homeostatic collapse, respectively. Finally, we identified directional blood flow occurring before the formation of endothelial tubes, indicating a pre-vascular transport mechanism. These findings demonstrate that, empowered by neural innervation, the YS serves as a coordinated developmental hub, facilitating complex crosstalk between neural, vascular, and metabolic systems during early vertebrate embryogenesis.