After perfusion, each mouse was killed by CO2 inhalation. One drop of fixative containing 2% formaldehyde and 0.5% glutaraldehyde was then instilled on the perfused eye. Next, the mouse was transcardially perfused through the left ventricle with 0.1 M PBS followed by fixative containing 2% formaldehyde and 0.5% glutaraldehyde in PBS (pH 7.4). After enucleation, the eyes were postfixed in the same fixative solution for 1 hour and then cryoprotected by passage through 10%, 20%, and 30% sucrose in PBS. After placing the tissue into molds containing tissue-freezing medium (TFM; Triangle Biomedical Sciences, Durham, NC), the tissue was snap frozen by placing the mold into a slurry of acetone and dry ice. Cryostat sections (10 μm thick) were cut, and the sections were placed on gelatin-coated glass slides. The slides were analyzed by fluorescence microscopy (Eclipse E800; Nikon Inc., Melville, NY), and photographs were taken with a cooled digital camera (SPOT Digital Camera System; Diagnostic Instruments, Inc., Sterling Heights, MI). To identify the tracer within the anatomic structures and to differentiate tracer signal from the autofluorescence of the tissue, the sections of the perfused eye were compared with sections obtained from the nonperfused control eye.