Orderly and specific patterns of neural wiring are critical for proper behavioral outcomes. Most studies on long- range neuronal connections have utilized invertebrate models or analyzed periphery-to-brain connectivity. Those studies have uncovered the basic mechanisms responsible for establishing long-range neuronal connections, such as axon pathfinding, topographic mapping and laminar-specific connectivity. However, our understanding of the other connectivity-based phenomena, e.g., axon-target nuclei selection, remains rather limited. Axon-target nuclei selection is the process, by which growing axons choose their final targets while avoiding the adjacent ones. However, how such selection is achieved within complex mammalian brain, is not well understood. In this application, we propose to examine mechanisms of axon-target matching, focusing on connections between superior colliculus (SC) and thalamus in the mammalian brain. SC is a midbrain center controlling head and eye movements in response to a sensory stimulation. SC also mediates visual cue- triggered defense responses, such as freezing and escaping. The superficial layer of SC (sSC) receives visual inputs from the retina and cortex and contains neurons that project to the deep layers of the SC and to the other subcortical areas. The sSC projections to thalamus modulate visual information to elicit appropriate behavioral responses to specific stimuli. Moreover, some studies have reported that projections to a specific thalamic nucleus comprise the second visual pathway that responds to sensory cues in the absence of the primary visual cortex. However, little is known about the molecular mechanisms that regulate the development of neuronal connections between sSC and thalamus. Recently, we discovered that expression of retinoid- related orphan receptor β (Rorβ) is highly enriched within sSC. Based on the layer-restricted expression of Rorβ, we hypothesize that Rorβ+ sSC neurons project axons to distinct thalamic nuclei. Using innovative genetic strategies, we will examine the role of Rorβ in the development of specific sSC circuits. We will also define underlying molecular mechanisms that regulate axonal projections of Rorβ+ neurons to distinct thalamic nuclei. This project will help us understand molecular basis of neuronal connections between sSC and thalamus that are involved in specific visual responses and fear-related behaviors. Proposed studies will also improve our understaninding of how long-range axon-target nuclei selection is regulated in mammalian brain. The novel genetic approach employed in this proposal should be broadly applicable for studying long-range projections throughout mammalian nervous system.