In a pilot developmental neurotoxicity study, a protocol was designed to utilize three-dimensional magnetic resonance (MR) images for linear and volumetric measurements of the developing rat brain. MR imaging, because of its non-destructive nature, provides a complement to traditional optical microscopy. Sprague-Dawley dams received 0, 1.25, 4.0 or 7.5mg/kg methylazoxymethanol acetate (MAM) by intraperitoneal injection during gestation days 13-15. At postnatal days (PND) 23 and 60, brains from representative male and female rats from two dams in each dose group were fixed with 10% neutral buffered formalin by transcardial perfusion for in situ MR imaging. A 7T small animal magnet system was used to obtain isotropic images at 100 microm resolution for PND 23 and 150 microm resolution for PND 60. Data from a rapid screening method based on midpoint MR slices of whole brain, cerebrum, cerebellum, and hippocampus showed a dose-related decreased volume of whole brain, cerebrum, and hippocampus in treated rats. Subsequent volumetric estimates using the Cavalieri method confirmed these findings. The brains were subsequently removed and processed for conventional histologic examination of hematoxylin and eosin-stained sections. It is concluded that MR imaging in rat developmental neurotoxicity studies offers the advantages of in situ volumetric measurements of brain structures while preserving the samples for conventional optical microscopy.