OBJECTIVE We examined the efficacy of an extended-release drug delivery system, nanosponge (NS) encapsulated compounds, administered intravitreally to lower intraocular pressure (IOP) in mice. METHODS Bilateral ocular hypertension was induced in mice by injecting microbeads into the anterior chamber. Hypertensive mice received NS loaded with ocular hypotensive drugs via intravitreal injection and IOP was monitored. Retinal deposition and retinal ganglion cell (RGC) uptake of Neuro-DiO were examined following intravitreal injection of Neuro-DiO-NS using confocal microscopy. RESULTS Brimonidine-loaded NS lowered IOP 12% to 30% for up to 6 days (P < 0.02), whereas travoprost-NS lowered IOP 19% to 29% for up to 4 days (P < 0.02) compared to saline injection. Three bimatoprost NS were tested: a 400-nm NS and two 700-nm NS with amorphous (A-NS) or amorphous/crystalline (AC-NS) crosslinkers. A single injection of 400 nm NS lowered IOP 24% to 33% for up to 17 days compared to saline, while A-NS and AC-NS lowered IOP 22% to 32% and 18% to 26%, respectively, for up to 32 days (P < 0.046). Over time retinal deposition of Neuro-DiO increased from 19% to 71%; Neuro-DiO released from NS was internalized by RGCs. CONCLUSIONS A single injection of NS can effectively deliver ocular hypotensive drugs in a linear and continuous manner for up to 32 days. Also, NS may be effective at targeting RGCs, the neurons that degenerate in glaucoma. CONCLUSIONS Patient compliance is a major issue in glaucoma. The use of NS to deliver a controlled, sustained release of therapeutics could drastically reduce the number of patients that progress to vision loss in this disease.