Quantitative analysis of subretinal injections in the rat

Background: Experimental therapeutic approaches to retinal degenerations of ten require the subretinal injection of a therapeutic agent. The injected v olume and the age of the animal can influence the proportion of the retinal surface affected by the subretinal injection. We have investigated the qua ntitative effect of a single injection in the subretinal space. Methods: No rmal and Royal College of Surgeons rats aged 1 week, 3 weeks or 2 months re ceived subretinal transscleral injections of 1, 3, 5 or 10 mu l China ink. After 24 h, animals were killed, injected eyes were enucleated and fixated, and the retinas flattened. An image analyzing program was used to measure the total retinal surface and the retinal surface affected by the dye. Resu lts: The mean retinal surface affected by the injection ranged from 5.24+/- 2.76 mm(2) to 14.8+/-2.3 mm(2), depending on animal age and injected volume . The injection affected 8.79+/-0.89 to 36.9+/-8.13% of total retinal surfa ce. There was no statistically significant difference between normal and Ro yal College of Surgeons rats. Intravitreal leakage of the dye was more freq uent with increasing injection volumes. Conclusion: The retinal sur-face af fected by a single subretinal injection increases with the injected volume, but this increase is not proportional. Higher volumes lead to a loss of in jected solution, either in the vitreous body or through the sclerotomy. In 2-month-old rats, a 3-mu l subretinal injection appears to have the best re producibility, with 20-30% of retinal surface covered by the injected dye.