OBJECTIVE: To determine the effect of single and multiple-dose 0.5% timolol maleate on intraocular pressure (IOP) and pupil size between 8 AM and 8 PM. Animals Nine female horses with normotensive eyes. Procedure IOP, horizontal and vertical pupil size were measured on a single day, between 8 AM and 8 PM at hours 0, 0.5, 1, 2, 4, 6, 8, 10, and 12. A single dose of 0.5% timolol maleate was applied to both eyes immediately after the first measurement at 8 AM. IOP and pupil size were measured at 8 AM and 4 PM in a 5-day experiment of twice-daily application of 0.5% timolol maleate. RESULTS: A significant decrease in IOP from 24.9 +/- 4.2 mmHg prior to application of timolol maleate to 20.7 +/- 3.1 mmHg (4.2 mmHg = 17%) was observed 8 h after single-dose application. A significant decrease in horizontal pupil size (2.0 mm = 11%) was present 6 h after single-dose application. In the multiple-dose experiment, a significant decrease in IOP was present on days 4 and 5 as compared to IOP measured prior to application of timolol maleate. A significant decrease in horizontal and vertical pupil size was present throughout the 5-day study as compared to the values obtained prior to treatment. CONCLUSIONS: 0.5% timolol maleate significantly decreased IOP and pupil size in normo-tensive eyes of this group of female horses in both single and multiple twice daily applications.