Endothelial glycocalyx (GCX) has been reported as a protective factor for vascular endothelial cells (VEC) in diabetes and hypertension. However, the involvement of GCX impairment in ocular vasculopathy remains unclear. We evaluated the changes in the GCX thicknesses of the retinal and choroidal capillaries in rats with diabetes and hypertension by cationic colloidal iron staining using a transmission electron microscope. In the control group, the mean (standard error of the mean) thicknesses of retinal and choroidal GCX were 60.2 (1.5) nm and 84.3 (3.1) nm, respectively. The diabetic rats showed a significant decrease of GCX thickness in the retina, but not in the choroid, compared to controls (28.3 (0.3) nm, p<0.01 and 77.8 (1.4) nm, respectively). In the hypertensive rats, both retinal and choroidal GCX were significantly decreased compared to the control values (10.9 (0.4) nm and 13.2 (1.0) nm, respectively, both p<0.01). Moreover, we could visualize the adhesion of leukocytes and platelets on the luminal surface of VEC, at the site where the GCX was markedly degraded. These findings suggest that the GCX prevents adhesion of leukocytes and platelets to the VEC surface, and this impairment may lead to ocular vasculopathy in diabetes and hypertension.