β-catenin in retinal interneurons is important for maintaining stability of the retinal microvasculature

Abstract

Purpose : Functional interactions between neurons, vasculature, and glia occur within neurovascular units and are critical for retinal homeostasis. We have previously shown that amacrine cells are required for generating and maintaining the intermediate plexus through precise regulation of hypoxia-inducible and pro-angiogenic factors. Besides VEGF, Wnt/β-catenin and transforming growth factor (TGF)β1 signaling are important for regulating the stability and permeability of vascular networks. In this study we used various transgenic approaches to delineate the role of these signaling pathways in amacrine and horizontal cells.

Methods : Transgenic mice expressing Cre recombinase specifically in amacrine and horizontal cells (Ptf1a-Cre mice) were mated with floxed VEGF, VHL, β-catenin, and TGF-β mice to generate conditional knockouts. The status of the intraretinal vascular plexuses was assessed in retinal flat mounts and by ultrastructural analyses. Fluorescein isothiocyanate (FITC)-conjugated bovine serum albumin (BSA) extravasation assays were performed to measure vascular permeability in mutants and control littermates.

Results : The number of branching points were similar in ptf1a-Cre; β-catenin ^f/+ and ptf1a-Cre; β-catenin ^f/f (intermediate plexus P=0.1766, deep plexus P=0.1268). However, confocal and electron microscopy revealed distinct anatomical differences in the intermediate and deep plexuses (but not the superficial plexus) in the mutants compared with controls. No significant leakage of FITC-BSA was observed in the three plexuses in controls, but a statistically significant increase in vascular permeability was observed in the outer layers of IPL and OPL in ptf1a-Cre; β-catenin^f/f mice at P23 (P=0.001996; β-catenin^f/f versus β-catenin^f/f; ptf1a-Cre; 5.9 × 10⁴ versus 6.9 × 10⁴ pixels). No abnormal leakage was observed in ptf1a-Cre; VEGF^f/f, ptf1a-Cre; VHL^f/f, or ptf1a-Cre; TGFβ1^f/f mice.

Conclusions : We show previously undescribed functions for amacrine and horizontal neurons in the regulation of the inner blood-retinal barrier. Learning to regulate these pathways in amacrine and horizontal cells may inform future therapies for diabetic retinopathy or other vasoproliferative diseases.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.