The inner and outer blood-retina barriers (BRBs) are formed by tight junctions between adjacent endothelial or retinal pigment epithelial (RPE) cells. Breakdown of BRBs is a major pathological change in age-related macular degeneration, diabetic retinopathy, retinopathy of prematurity, and uveitis. While previous studies have yielded a better understanding about the roles of the inner BRB under physiological and pathological conditions, surprisingly little is known about the regulation and pathophysiology of the outer BRB. Since the outer BRB is responsible for ~85% of blood circulation to the retina, it is unimaginable that ischemia-induced pathological change in the outer BRB plays an insignificant role in the overall pathology of retinochoroidal vascular diseases. To investigate the mechanisms of outer BRB breakdown and to test the concept of inhibiting outer BRB permeability as a therapeutic strategy for retinochoroidal vascular diseases, we have prepared gene knockout systems for the mouse RPE and M?ller glia, cells that regulate the function of both inner and outer BRBs through vascular endothelial growth factor (VEGF-A). Using these conditional gene knockout systems, we have disrupted VEGF and its receptor (VEGFR2) in the mouse RPE and have generated mice with VEGF disruption in the M?ller cells. In Specific Aim 1, we will test our hypothesis that outer BRB breakdown is a significant contributor to diabetes/ischemia-induced overall retinal vascular leakage through autocrine VEGF/VEGF-R2 signaling in the RPE by measuring the total retinal vascular leakage, the number of significant breakpoints in the outer BRB, and the quantity of outer BRB-specific leakage in the RPE-specific VEGF and VEGFR2 knockout mice after inducing ischemia or diabetes. In Specific Aim 2, we will test the concept of inhibiting outer BRB permeability as a therapeutic strategy for uveitis by examining the total retinal vascular leakage, the number and severity of retinal detachment, the quantity of outer BRB-specific leakage, and the expression of inflammatory biomarkers in the RPE-specific VEGFR2 knockout mice after inducing uveitis. As a control for inner BRB breakdown, we will also measure the same parameters in uveitic M?ller cell- specific VEGF knockout mice. In Specific Aim 3, we will determine the molecular mechanism of diabetes/ischemia-induced outer BRB breakdown by investigating the biochemical pathway governing the regulation of tight-junction proteins.