Spatiotemporal characterization of periocular mesenchyme heterogeneity during anterior segment development

Establishment of the ocular anterior segment (AS) is a critical event during development of the vertebrate visual system. Failure in this process leads to Anterior Segment Dysgenesis (ASD), which is characterized by congenital blindness and predisposition to glaucoma. The anterior segment is largely formed via a neural crest-derived population, the Periocular Mesenchyme (POM). In this study, we aimed to characterize POM behaviors and identities during zebrafish AS development. POM distributions and migratory dynamics were analyzed using transgenic zebrafish embryos (Tg[], Tg[], Tg[], Tg[], and Tg[] throughout the course of early AS development (24-72hpf). imaging analysis revealed unique AS distribution and migratory behavior among the reporter lines, suggesting AS mesenchyme (ASM) is a heterogenous population. This was confirmed using double hybridization. Furthermore, we generated ASM transcriptomic profiles from our reporter lines and using a four-way comparison analysis uncovered unique ASM subpopulation expression patterns. Taken together, our data reveal for the first time that AS-associated POM is not homogeneous but rather comprised of several unique subpopulations identifiable by their distributions, behaviors, and transcriptomic profiles.