As essential components of gene expression networks, transcription factors regulate neural circuit assembly.GS homeobox 1(gsx1) is expressed in the developing visual system; however, no studies have examined its role in visual system differentiation. In zebrafish, retinal ganglion cell (RGC) axons terminate in ten arborization fields (AFs) in the optic tectum (TeO) and pretectum (Pr). Pretectal AFs (AF1-AF9) mediate distinct and essential visual behaviors, yet we understand very little about their development compared to AF10 in the TeO. Usinggsx1zebrafish mutants, immunohistochemistry, and transgenic lines, we observed thatgsx1is required for vesicular glutamate transporter,slc17a6b, expression in the Pr, but not overall neuron number.gsx1mutants have normal eye morphology, yet exhibit impaired vision, and a significantly reduced volume of RGC axons innervating the Pr and TeO, including loss of AF7. Consistent with this, prey capture is reduced ingsx1mutants. Timed laser ablation ofslc17a6b-positive neurons reveals that they aide directly in AF7 formation. This work is the first to implicategsx1in establishing cell identity and functional neural circuits in the visual system.