Retinal ganglion cells (RGCs) serve as a vital connection between the eye and the brain with damage to their axons resulting in loss of vision and/or blindness. Retinal organoids are three-dimensional structures derived from human pluripotent stem cells (hPSCs) which recapitulate the spatial and temporal differentiation of the retina, providing a valuable model of RGC development in vitro. The working hypothesis of these studies is that hPSC-derived RGCs are capable of extensive outgrowth and display target specificity and pathfinding abilities. Initial efforts focused on characterizing RGC differentiation throughout early stages of organoid development, with a clearly defined RGC layer developing in a temporally-appropriate manner expressing a compliment of RGC-associated markers. Beyond studies of RGC development, retinal organoids may also prove useful to investigate and model the extensive axonal outgrowth necessary to reach post-synaptic targets. As such, additional efforts aimed to elucidate factors promoting axonal outgrowth. Results demonstrated significant enhancement of axonal outgrowth through modulation of both substrate composition and growth factor signaling. Furthermore, RGCs possessed guidance receptors that are essential in influencing outgrowth and pathfinding. Subsequently, to determine target specificity, aggregates of hPSC-derived RGCs were co-cultured with explants of mouse lateral geniculate nucleus (LGN), the primary post-synaptic target of RGCs. Axonal outgrowth was enhanced in the presence of LGN, and RGCs displayed recognition of appropriate targets, with the longest neurites projecting towards LGN explants compared to control explants or RGCs grown alone. Generated from the fusion of regionally-patterned organoids, assembloids model projections between distinct regions of the nervous system. Therefore, final efforts of these studies focused upon the generation of retinocortical assembloids in order to model the long-distance outgrowth characteristic of RGCs. RGCs displayed extensive axonal outgrowth into cortical organoids, with the ability to respond to environmental cues. Collectively, these results establish retinal organoids as a valuable tool for studies of RGC development, and demonstrate the utility of organoid-derived RGCs as an effective platform to study factors influencing outgrowth as well as modeling long-distance projections and pathfinding abilities.