Retinal O2 utilization was studied to identify the O2 consuming processes in the retina and the spatial distribution of those processes. Neural retina, retinal pigment epithelium and choroid were dissected from the toad eye and superfused with an oxygenated Ringer's solution. Double-barreled microelectrodes were used to measure O2 and local voltage simultaneously within the retina in both light and dark adaptation. The profile of PO2 was measured during a withdrawal of the electrode tip across the retinal pigment epithelium and through the neural retina. The PO2 decreased through the distal retina, reaching a minimum in the inner segment or outer nuclear layer, and then increased steadily through the proximal retina. From fitting PO2 profiles measured in the dark-adapted retina to a three-layer diffusion model, O2 consumption was found to be 1.0 +/- 0.4 and 0.4 +/- 0.3 ml O2 (100 g min)-1 in the outer and inner halves of the retina, respectively. Light decreased consumption in both halves of the retina. In steady illumination (500 nm) that saturated the ERG b- and c-waves, O2 utilization decreased significantly to 48% and 68% of the dark values in the outer and inner retina, respectively. When Na+ was removed from the superfusate to inhibit the photoreceptor Na+/K+ pump, O2 consumption in the outer retina decreased by about the same amount as in light, but O2 consumption in the inner retina was not significantly affected.