UC Berkeley

Coordinated development of neurons, glia, and the vasculature is essential for formation of a functional nervous system, yet the mechanisms that coordinate these processes remain unclear. One possibility is that neural activity directly instructs vascular development, while an alternative model proposes that glial and vascular development proceed through a partially parallel, activity-independent program. Here, I investigated how neural activity, Müller glia, and the developing vasculature are coordinated during retinal angiogenesis in the mouse retina.Using quantitative confocal imaging from postnatal day 5 to eye-opening, we mapped the emergence of the superficial, intermediate, and deep vasculature layers and found that they emerged normally in mice lacking the b2-containing nicotinic acetylcholine receptors, despite a dramatic reduction in cholinergic signaling. Tip cell density and overall vessel growth were unchanged, indicating cholinergic wave activity is not required for the emergence of retinal vasculature.We next defined the developmental timeline of Müller glia–vascular physical interactions. Sparse labeling and immunohistochemistry revealed that Müller glial lateral processes closely associate with endothelial tip cells during intermediate- and deep-layer angiogenesis and establish Aquaporin-4–enriched endfeet at vascular contact sites from the earliest stages of growth, even when vessel trajectories are perturbed.Finally, two-photon calcium imaging combined with simultaneous electrophysiology demonstrated that Müller glial endfeet exhibit robust, compartmentalized calcium transients during development. Although a subset of events was temporally correlated with retinal waves, enhancing neurotransmitter spillover increased wave-associated activity in glial stalks but also in endfeet and other lateral processes. Despite this enhancement, endfoot calcium transients remained weakly coupled to spontaneous neuronal activity, indicating that signaling at the glial-vascular interface is not strictly driven by retinal waves. Together, our results support a model in which Müller glia engage growing vessels largely through an activity-independent, parallel developmental program that provide instructive cues for retinal angiogenesis.