O2-sensitive microelectrodes were used to measure PO2 as a function of depth through the retina and choroid of anesthetized and artificially ventilated guinea pigs. The guinea pig retina is of particular interest, because it has a typically mammalian structure but no retinal circulation; it relies totally on choroidal delivery of O2 and other nutrients. Measurements of intraretinal O2 distribution in an avascular mammalian retina have not previously been reported. Under normal ventilation conditions, PO2 decreased monotonically from the choroid (33.6 +/- 2.9 mmHg, n = 11) to near zero (0.4 +/- 0.1 mmHg) at the retina-vitreous boundary. The inner half of the retina had an average PO2 of only 0.6 +/- 0.1 mmHg. Stepwise increases in inspired O2 (from 20 to 40 to 60 to 80 to 100%) had surprisingly little effect on choroidal PO2. Rapid changes (20-100%) produced overshoot-type responses in the choroid before recovery to levels only slightly above those found in normoxia. This indicates the presence of an active O2-regulatory mechanism in the guinea pig choroid. Addition of CO2 (5%) to O2 ventilation appeared to break down this control mechanism and led to dramatic and sustained increases in PO2 throughout the retina and choroid. The demonstration of an O2-regulating mechanism in the guinea pig choroid that maintains choroidal PO2 well below that in the systemic arterial blood, coupled with the observation of very low O2 levels throughout the inner retina, suggests that the O2 requirement of the inner retina in the guinea pig is small and that O2 levels in the choroid are deliberately constrained.