DISCOVERING CORRELATIONS BETWEEN GENETIC MODIFICATIONS AND STRUCTURAL CHANGES IN ADULT ZEBRAFISH EYE USING OPTICAL COHERENCE TOMOGRAPHY

Abstract

Visual impairments, such as refractive errors and cataracts, present a growing global health challenge, driving the critical need for advanced diagnostic imaging and genetically tractable animal models. The zebrafish (Danio rerio) has emerged as a premier model for vision research; however, translating clinical Optical Coherence Tomography (OCT) to adult zebrafish presents significant optical challenges regarding tissue penetration and lateral resolution. This thesis presents an interdisciplinary approach bridging mechanical engineering and vision biology, aiming to optimize a customized OCT imaging framework to uncover the structural consequences of targeted genetic modifications in the adult zebrafish eye. To overcome inherent optical limitations, a dual Spectral-Domain OCT (SD-OCT) platform was engineered and optimized. A custom 1310 nm system was tailored to maximize tissue penetration depth, enabling comprehensive whole-eye biometry from the anterior cornea to the posterior retina. Concurrently, an 880 nm system equipped with a specialized probe was optimized for high-resolution microstructural retinal imaging. Paired with rigorous image processing, including multi-volume despeckle averaging and index-matching protocols applied to freshly euthanized specimens, this dual-system approach successfully achieved measurement-grade structural clarity and mitigated scattering artifacts. This validated imaging framework was subsequently applied to investigate the roles of gap junction (connexin) and hemichannel (pannexin) proteins in ocular development and structural maintenance. Quantitative biometry revealed distinct, channel-specific phenotypes. Depletion of the connexin gjd2b/Cx35.1 resulted in significantly reduced axial length and hyperopic shifts, whereas closely related Cx27.5 knockouts exhibited no structural biometric alterations. Investigations into the pannexin family demonstrated that both Panx1b and Panx2 deficiencies led to increased axial lengths and myopic shifts, with Panx2 knockouts also displaying marked lens epithelial malformations. Furthermore, a comprehensive longitudinal study of Panx1a mutants revealed a progressive structural decline characterized by axial myopia, age-related cataract-like lens texture change, epithelium lens defects, and significant retinal ganglion cell layer thinning. Ultimately, this research establishes a robust, non-invasive quantitative phenotyping methodology for small animal models. By directly linking optical instrumentation design to biological discovery, this work provides critical insights into the genetic regulation of ocular growth and integrity, establishing a strong structural foundation for future translational investigations into human visual diseases