Objectives The aim of this study was to assess the effect of topical tetracaine hydrochloride 0.5% on intraocular pressure (IOP) in ophthalmologically normal cats. Methods Twenty domestic shorthair cats (40 eyes) were used in this study. Each cat was randomly allocated to one of two groups (treatment or control). Baseline IOP (T0) was measured in each cat, and then one drop of tetracaine hydrochloride 0.5% or artificial tears was administered into a randomly chosen eye of each cat in the treatment and control groups, respectively. Repeat IOP measurements were performed at 2 mins (T2), 5 mins (T5), 15 mins (T15) and 30 mins (T30) with a rebound tonometer. Results Mean baseline IOP in all eyes was 20.6 ± 2.5 mmHg. After the unilateral administration of tetracaine, mean IOP decreased significantly in the treated eye at T2 ( P = 0.01). Mean IOP returned to baseline values at T15. The mean IOPs in the treated eyes at T0, T2, T5, T15 and T30 were 20.6 ± 3.3 mmHg, 18.2 ± 2.5 mmHg, 18.2 ± 3.4 mmHg, 20.2 ± 3.2 mmHg and 19.8 ± 2.7 mmHg, respectively. A significant difference in IOP was found at all time points between the tetracaine and control groups (P <0.03). Conclusions and relevance The results of the present study showed a statistically significant reduction in mean IOP 2 mins after the administration of tetracaine hydrochloride 0.5% in the treated eyes of the cats.