Vascular endothelial growth factor (VEGF) is one of the major regulatory molecules in diabetic retinopathy (DR). In our previous study, we demonstrated that succinate levels were elevated in the retinas of diabetic rats and that the knockdown of the succinate receptor, G-protein-coupled receptor 91 (GPR91), inhibited the release of VEGF and attenuated retinal vascular disorder in the early stages of DR. In the present study, we examined the signaling pathways involved in the GPR91-dependent release of VEGF in the retinal ganglion cell line, RGC-5. The cells were infected with a lentiviral small hairpin RNA (shRNA) expression vector targeting GPR91 (LV.shGPR91). Immunofluorescence staining revealed that GPR91 was predominantly localized in the cell bodies of the RGC-5 cells. RT-qPCR, western blot analysis and ELISA indicated that succinate exposure upregulated VEGF expression, activated the extracellular signal-regulated protein kinase (ERK)1/2, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) signaling pathways and led to the release of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2). The knockdown of GPR91 inhibited ERK1/2 and JNK activity, but did not inhibit the activation of the p38 MAPK pathway. The increase in COX-2 expression and the release of PGE2 were inhibited by transduction with LV.shGPR91 and ERK1/2, JNK and COX-2 inhibitors. The expression and release of VEGF showed similar results. Cell Counting Kit-8 (CCK-8) assays revealed that the shRNA-mediated knockdown of GPR91 decreased the proliferation of RF/6A cells cultured in succinate-conditioned medium. Our data suggest that GPR91 modulates the succinate-induced release of VEGF through the MAPK/COX-2/PGE2 signaling pathway.