NADPH oxidases (NOX) are activated in ischemic conditions leading to increases in reactive oxygen species (ROS) and neurotoxicity. The aim of the present study was to investigate the role of NOX in the development of retinal pathologies, associated with excitotoxicity and the evaluation of NOX inhibitors as putative therapeutic agents. Sprague-Dawley rats were used for the induction of the in vivo retinal model of (RS)-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid hydrobromide (AMPA) excitotoxicity. Rats were intravitreally administered with PBS, AMPA (42 nmoles) or AMPA + NOX inhibitors, VAS2870 (pan-NOX inhibitor, 10-6-10-4 M), ML171 (NOX1 inhibitor, 10-5, 10-4 M), and GLX7013114 (NOX4 inhibitor, 10-4 M). Immunohistochemical studies were performed using antibodies raised against nitrotyrosine, a ROS/oxidative stress marker, bNOS, a neuronal marker for nitric oxide synthase and the macro and microglia markers, glial fibrillary acidic protein and ionized calcium-binding adaptor molecule-1, respectively. VAS2870 and ML171 showed neuroprotective and anti-inflammatory actions reversing the AMPA induced reduction of bNOS expressing amacrine cells and attenuating macro/microglial activation. GLX7013114 (10-4 M) did not protect bNOS expressing amacrine cells, but it did attenuate the AMPA induced increase in nitrotyrosine positive cells and activation of glial cells. These results suggest that NOX1, NOX4 and possibly NOX2 (due to the actions of VAS2870) play an important role in the pathophysiology of the retina and that NOX inhibitors are putative neuroprotective and anti-inflammatory agents against retinal abnormalities caused by excitotoxicity.