Topical administration of prostaglandin F2 alpha (PGF2 alpha) produced a reduction in intraocular pressure in eyes of rabbits, cats, and cynomolgus monkeys. In rabbit eyes at 5 or 6 hr, 50 micrograms, 100 micrograms, or 250 micrograms of PGF2 alpha caused a significant intraocular pressure reduction with a small miotic effect. Treatment with 500 micrograms, 750 micrograms, or 1000 micrograms of PGF2 alpha lowered intraocular pressure significantly in cat eyes for at least 24 hr with the development of profound pupillary constriction. Administration of 500 micrograms, 750 micrograms, or 1000 micrograms of PGF2 alpha produced a significant reduction of intraocular pressure in monkey eyes lasting at least 24 hr, with an initial hypertensive phase and a small decrease in pupillary diameter in the treated eyes. Tonography revealed an increased facility of outflow simultaneous with the reduction of intraocular pressure in the eyes of cats and monkeys. These increases of outflow facility could not explain completely the reductions in intraocular pressure. The aqueous humor flow measured by fluorophotometry was unaltered in both species, and possible reasons for this finding are discussed. Anterior chamber aqueous humor protein was significantly higher in cat eyes topically treated with 750 micrograms of PGF2 alpha than in the diluent-treated fellow eyes.