Insulin has been detected by ELISA in the vitreous of the normal and streptozotocin-diabetic rat at levels for both about 1% of those in serum. 131I-labeled insulin, administered to conscious rats via an indwelling cannula in the right atrium, was found to cross the blood-ocular barrier into the vitreous. Autoradiographic gel analysis showed the peptide was transferred as an intact molecule. Vitreous insulin levels reflected serum levels as seen in relatively constant vitreous-to-serum insulin ratios over a wide range of serum insulin concentrations. The rate of blood-to-vitreous passage of insulin was about the same in normal as in diabetic rats (fasting serum glucose greater than or equal to 21 mM). At least a portion of vitreous insulin is therefore of pancreatic origin, and retinal tissue in the normal and diabetic animal is thus accessible to circulating hormone. The blood-ocular barrier is unaltered in streptozotocin diabetes with regard to insulin passage.