Interstitial fluid pressure (Pi) has been measured in vivo in ocular tissues of rats and rabbits, and in vitro in corneas of cows and humans with micropipettes connected to a servocontrolled counterpressure system. The in vivo measurements were performed after anaesthesia and careful immobilization of the head. In all species Pi became gradually more negative from the limbus to the cornea centre; in rabbits the change was from -17.8 mmHg 0.5 mm centrally to the limbus to -42.7 mmHg in central cornea. Cornea and sclera interstitial fluid was isolated by centrifugation, and was used for analysis of colloid osmotic pressure (both tissues) and Na(+)-concentration and electrophoresis (cornea only). During low speed centrifugation (3000 rpm), a wet weight fraction of 2-4% was isolated from cornea, having a colloid osmotic pressure not significantly different from that of plasma (18.9 mmHg). Na+ analysis suggested that the isolated fluid did not derive from corneal cells. Electrophoresis showed a protein pattern in cornea fluid similar to that of plasma except for a band with MW corresponding to 39,000 found in cornea only. The present study demonstrates a pressure gradient from sclera to cornea, and along cornea from periphery to centre, and suggests that proteins in cornea fluid contribute significantly to swelling pressure.