Intravitreal injections are currently used in the rat to introduce a therapeutic factor in the eye, especially for experimental treatments of retinal degenerations. The injected volume and its location can influence the quantification of results. We have investigated the quantitative effect of a single intravitreal injection in rats at different ages and for different volumes. Albinos rats aged three weeks or two months received intravitreal injections of 1, 3, 5 or 10 microl China ink. Animals were sacrificed immediately after injection, eyes were enucleated, fixated, embedded in paraffin and microtomy was performed in a sagittal plane. Regularly spaced sections were analyzed to reconstruct the vitreous and injected dye volumes. The measured vitreous volume was 6.76 +/- 0.37 mm(3) in three weeks old rats and 13.36 +/- 0.64 mm(3) in two months old rats. Mean intravitreal ink volumes immediately after injection were 0.8 mm(3) for 1 ml injections, 2 mm( 3) for 3 ml, 2.3 to 2.6 mm(3) for 5 microl and 3.2 mm(3 ) for 10 microl. The percentage of vitreous volume involved by the injection ranged from 4.4% to 33.2%. The injected volume is limited by the large lens size of the rat. Extraocular loss of injected solution increases for higher injected volumes, with larger standard deviations. In this model, the dye tends to localize behind the lens. A 3 or 5 microl volume appears to have the best reproducibility with minimum loss of solution.