Vascular comorbidities are inherently linked to the pathogenesis of central retinal artery occlusion (CRAO) and central retinal vein occlusion (CRVO). However, the endothelial-mediated pathogenic mechanisms that precede, and therefore modulate, luminal occlusion have not been clarified. The aim of this study was to delineate the pattern of endothelial morphometric alteration in the central retinal artery and vein in patients with vascular comorbidities. Eyes with a previous history of vascular occlusion were not included in this study in order to avoid the confounding effects of post-occlusion endothelial changes. This study also sought to determine if vascular comorbidities had a disparate effect on arterial and venous endothelium in the optic nerve head. Comparisons were made between 13 human eyes from patients with vascular comorbidities and 22 control eyes from patients with no known systemic disease. Novel micro-cannulation techniques developed in our laboratory were used to label the cytoskeleton and nuclei of endothelial cells in the central retinal artery and vein following which images were captured using confocal microscopy. Endothelial and nuclear morphometric parameters were quantified in different laminar regions of the optic nerve head. F-actin stress fibre expression was also quantified. Analysis of covariance was used to determine statistical differences between the two groups. Interestingly, age did not influence endothelial morphometry, nuclear morphometry or f-actin expression in central retinal vessels. There were also no arterial endothelial differences between control and disease groups in any laminar region. Endothelial f-actin stress fibre expression increased significantly in the central retinal vein in patients with vascular comorbidities. The greatest change in these eyes was found to occur at the posterior lamina cribrosa. Increased venous endothelial f-actin stress fibre expression may reflect vascular comorbid disease-induced alterations to hemodynamic properties and coagulation cascades in the central retinal vein. The posterior lamina may be an important site for thrombus formation in CRVO as venous endothelia in this region are most influenced by the presence of vascular comorbidities. The findings of this study suggest that the role of endothelial dysfunction in CRVO and CRAO pathogenesis could be different. © 2013 Elsevier Ltd.