Purpose. To characterize a technique that concurrently assesses all aqueous humor hydrodynamic parameters in mouseeyes. Methods. Mouse outflow facility (C) was determined by multiple flow-rate infusion and episcleral venous pressure (Pe) measured by manometry. The animals were then euthanatized, eliminating aqueous formation rate (Fin) and Pe. C was determined again (Cdead) while uveoscleral outflow (Fudead) and Fin were deduced. To assess whether Fudead would remain the same as Fulive, the animals were perfused with FITC-dextran and Fu determined. The effects of IOP-lowering drugs on the parameters of aqueous hydrodynamics were also evaluated. Results. Under the conditions tested, Fulive (0.012 ± 0.003 /xL/min) was not different from Fudead (0.015 ± 0.003 fxL/min; P = 0.472). In anesthetized mice, IOP = 11.4 ± 0.2 mm Hg (mean ± SEM, n = 8), C = 0.018 ± 0.0006 fxL/min/mm Hg, Pe = 5.4 ± 0.2 mm Hg, Fin = 0.14 ± 0.0007 fxL/min, and Fu = 0.029 ± 0.005 fxL/min. Cdead was not different from C (P = 0.317). Latanoprost reduced IOP by increasing C by 0.009 ± 0.0003 fxL/min/mm Hg (P < 0.001), without affecting Fin or Fu. Betaxolol reduced Fin by 0.075 ± 0.021 fxL/min (P = 0.009). Brimonidine increased C by 0.005 ± 0.0005 fxL/ min/mm Hg (P < 0.001) and Fu by 0.013 ± 0.003 fxL/min (P = 0.007).Conclusions. In this study, a unique technique was developed to concurrently assess IOP, C, Pe, Fin, and Fu in the mouse eye. This experimental approach should be useful to evaluate effects of pharmacologic agents or genetic manipulations on aqueous humor dynamics in mice and other animal models.