Purpose: Our previous studies have confirmed that the catecholamine norepinephrine (NE) promoted the corneal infection and progression of Pseudomonas aeruginosa (P. aeruginosa) keratitis. Here we explored the effects of adrenoceptor (AR) blockers on the severity of P. aeruginosa keratitis in mice. Methods: A total of 630 C57BL/6 mice were used and ocularly inoculated with P. aeruginosa (ATCC 19660). Six AR blockers were topically administrated 6 h before or combined with Tobrex 12 h after bacterial inoculation. Clinical scores, neutrophil infiltration and neutrophil extracellular traps (NETs), proinflammatory factors and bacterial virulence factors expression, bacterial load and biofilm formation were evaluated in vivo. The growth inhibitory and bactericidal activity of ICI118551 was measured in vitro. Results: Among various blockers, the selective β2-AR blocker ICI118551 showed the most significant improvement in disease severity. Prophylactic administration of ICI118551 attenuated the clinical scores, neutrophil infiltration, NETs formation, proinflammatory factors expression of infected corneas, accompanied with the reduction of bacterial load, virulence factors expression, and biofilm formation. When adjunctive treatment with Tobrex, ICI118551 exhibited apparent therapeutic effects with the reduced severity after 12 h of bacterial inoculation. Moreover, ICI118551 inhibited bacterial growth and possessed bactericidal activity in vitro. In addition, ICI118551 had no significant influence on ocular barrier function and intraocular pressure. Conclusions: The selective β2-AR blocker ICI118551 attenuated the severity of P. aeruginosa keratitis in mice, which may represent the potential prophylactic and therapeutic approach to control the P. aeruginosa corneal infection.