We aimed to evaluate the effect of the topical application of a strong corticosteroid, difluprednate, on the levels of inflammatory and angiogenic cytokine in the vitreous and aqueous humor after laser photocoagulation. Pigmented rabbits were treated with retinal laser photocoagulation and divided into 4 groups, namely control (no additional treatment), topical application of difluprednate 0.05%, betamethasone sodium phosphate 0.1%, and sub-Tenon injection of triamcinolone acetonide (STTA). Samples of vitreous and aqueous humor were collected on posttreatment days 0, 1, 7, and 14. The levels of intraocular vascular endothelial growth factor (VEGF), interleukin-6 (IL-6), intercellular adhesion molecule-1 (ICAM-1), and monocyte chemotactic protein-1 (MCP-1) were measured using an immunoassay. Intraocular pressure (IOP) was monitored in each group. VEGF, IL-6, ICAM-1, and MCP-1 were significantly elevated on day 1 and were reduced in both the vitreous and aqueous humor following topical application of difluprednate and STTA. Topical betamethasone reduced their levels in the aqueous humor but not in the vitreous. A significant increase of IOP induced by difluprednate returned to control levels after withdrawal of administration. Although the elevation of IOP was an expected, manageable side effect, topical application of difluprednate was more effective than STTA and betamethasone for reducing inflammatory cytokine levels after laser treatment.