Perivascular Neurons Instruct Three-Dimensional Retinal Vascular Lattice Formation via Direct Neurovascular Contact

Abstract

Purpose : The retinal vasculatures are assembled into the 3D lattice network of laminar vascular beds to provide nutrients and oxygen for accurate visual functions. However, the mechanisms controlling 3D vascular lattice formation are largely unknown in contrast to 2D vascular growth. We hypothesized that retinal neuron subsets play instructive roles in guiding the vertical vascular penetrance and supporting the 3D vascular lattice architecture along the neural plexiform.

Methods : We used retro-orbital (R.O) injection of the engineered AAV2 virus to visualize the subset of retinal ganglion cells (RGCs) with close contact with blood vessels. Then, we identified the molecular profiles of our targeted RGC types. We carried out a mutant analysis for vascular development. We combined immunohistochemistry, electron microscopy, 3D cell reconstruction, multiphoton imaging, and fluorescein fundus imaging to analyze these mutants. Furthermore, we applied multiple established experimental optic neuropathy models to analyze these mutants.

Results : Retro-orbital delivery of AAV2-BR1-GFP revealed a subset of Fam19a4/Nts-positive retinal ganglion cells (Fam19a4/Nts-RGCs) that physically contact endothelial cells with unique perisomatic endfeet. Genetic ablation of Fam19a4/Nts-RGCs led to disorganized vascular sprouts close to the RGC layer, disrupting the planar organization of the vascular bed. We identified Piezo2 as a Fam19a4/Nts-RGCs-enriched protein during retinal vascular development. Pan-neuronal and Fam19a4/Nts-RGC specific deletion of Piezo2 led to the loss of Fam19a4/Nts-RGC-vascular contacts and phenocopied the vascular defects upon the Fam19a4/Nts-RGC ablation. These abnormal 3D vascular lattice formations lead to reduced vessel perfusions and global hypoxia in the adult retina. Furthermore, these mutants displayed accelerated retina neuron losses and declined visual functions upon transient ischemic challenge.

Conclusions : We uncovered a specific and unexpected neuron-type role in instructing 3D vascular lattice formation via direct neurovascular interaction in the retina. Furthermore, we also demonstrated that Piezo2 is a key structural mediator for this mechanism.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.