Mannitol is used widely to decrease intracranial pressure (ICP); however, the mechanism by which this effect occurs is unclear. This study was designed to examine the effects of mannitol on cerebrospinal fluid (CSF) formation rate (Vf), resistance to reabsorption of CSF (Ra), and brain tissue water content (BTWC). Eighteen New Zealand White rabbits were allocated into one of three groups and studied at baseline and at two sequential doses of 20% mannitol: 0.75 g/kg followed by 4.4 mg·kg−1·min−1, and 2.0 g/kg (1.25 g/kg added to the initial dose of 0.75 g/kg) followed by 8.6 mg·kg−1·min−1. In Group 1, closed ventriculocisternal perfusion (VCP) was performed to determine changes in ICP due to mannitol. In Group 2, the increase in CSF osmolality due to mannitol was determined. In Group 3, mock CSF was used for open VCP to determine Vf and Ra. At the conclusion of each study, brain tissue samples were taken for determination of BTWC. Mannitol increased CSF and plasma osmolality. Ra was increased by 104% with the low dose of mannitol and not significantly changed by the high dose. Mannitol decreased BTWC, Vf (by 49% with the high dose), ICP, and hematocrit. The authors conclude that two of the mechanisms contributing to decreased ICP with mannitol are: 1) decreased CSF volume as indicated by decreased Vf, and 2) decreased brain tissue volume as indicated by decreased BTWC.