Abstract Background and objective Vascular endothelial growth factor (VEGF), a key mediator of intraocular neovascularization, is triggered by hypoxia and has been shown in the eyes of animal models of central retinal vein occlusion (CRVO). However, there is little information on CRVO in humans, in particular, the identity of VEGF-producing cells. Study design The study design was molecular localization of the site of VEGF production in the eyes of patients with CRVO. Participants Ten formaldehyde solution-fixed and paraffin-embedded eyes removed surgically from patients with CRVO and neovascular glaucoma were studied. Five eyes with uveal melanoma and no neovascularization served as control specimens. Methods Thin whole-eye sections were hybridized in situ with a VEGF-specific probe to identify cells producing VEGF messenger RNA (mRNA). Results All ten eyes with CRVO showed evidence of intraretinal expression of VEGF mRNA. In all eyes, the inner nuclear layer showed VEGF-upregulated expression. Upregulation of VEGF mRNA was identified in four eyes in the ganglion cell layer and in two eyes with retinal detachment in the outer nuclear layer as well. Conclusions The population of VEGF-producing retinal cells in each eye is likely to represent cells residing in ischemic regions of the retina. Hypoxia-induced VEGF is, most likely, the linking factor between retinal ischemia and iris and retinal neovascularization in CRVO.