Abstract This proposal investigates the cornea-trigeminal neural circuit with particular emphasis on the relationship between the cornea epithelium and the intraepithelial nerves to investigate the hypothesis that ocular surface pathologies that involve transient and/or prolonged corneal epithelial barrier disruption will cause greater nociceptor stimulation, discomfort, pain and a compensatory reduction in epithelial and stromal nerve density. Single cell RNA and epigenetic profiling and in situ hybridization will be performed in corneal and trigeminal ganglion cells that project to the cornea in homeostatic conditions and after epithelial barrier disruption due to dry eye and corneal epithelial debridement. Data from these assays will be integrated and single cell atlases of the cornea and trigeminal ganglion under normal and stressed conditions will generated that identify cells based on unique expression profiles. Cells will be spatially resolved in these tissues to identify communications between corneal cells and nerve axons and between trigeminal neuronal and non-neuronal cells. In the ganglion, temporally and spatially resolved transcriptomic data will be associated with the changes in corneal epithelial nerve morphology and sensitivity that occur after barrier disruption to identify genes that maintain the corneal epithelial nerves in health and promote the pathological changes after barrier disruption. Three specific aims are proposed to investigate the cornea-trigeminal circuit. Aim 1 will investigate the effects of barrier disruption from dry eye and epithelial debridement on transcriptomic profiles, corneal epithelial nerve morphology and nerve function. Aim 2 will investigate the effects of preserving epithelial barrier in the MMP-9 knockout strain in dry eye and epithelial wounding on these parameters, and Aim 3 will investigate the effects of two neural sensitizing chemokines on the circuit in normal and dry eyes. The proposed experiments investigate the cornea epithelial-trigeminal sensory circuit at a coordinated deep level to identify factors that impact the intraepithelial nerves in health and disease. At the conclusion of this project, we will have a better understanding of the mechanisms by which corneal epithelial cells and intact barrier suppress nerve activation and the factors produced by cornea cells in barrier disrupted corneas that cause nerve degeneration and heightened sensitivity. The benchmark accomplishments of this project will be a fundamental new understanding of the impact of corneal epithelial barrier disruption that occurs in common corneal diseases on the integrity and function of the corneal epithelial nerve network.