An alternative method for preparation of tissue sections from the rat mammary gland

In toxicopathological studies of the rat mammary glands, the guidelines of the Registry of Industrial Toxicology and Animal Data (RITA) recommend transverse sections of the inguinal mammary gland. However, occasionally limited amounts of mammary gland tissue are found in transverse sections. We compared transverse sectioning with an alternative method comprising horizontal sections of the rat mammary glands. Normal cycling female Sprague Dawley rats were sacrificed in proestrous, estrous, metestrous and diestrous, and samples from all mammary glands were collected. Transverse sections were prepared from the left-sided glands, and horizontal sections were cut from the right-sided glands. Sections were stained with HE, and epithelial and myoepithelial cells were visualized by immunohistochemical staining of cytokeratin 18 and alpha smooth muscle actin, respectively. Area of the mammary fat pad, mammary epithelium and connective tissue were measured in randomly sampled vision fields from each section. Horizontal sections contained a significantly larger area of mammary fat pad as well as glandular and connective tissue. No differences in tissue densities of epithelial or myoepithelial cells or connective tissue were observed between transverse sections and horizontal sections. Interestingly, densities of epithelium and connective tissue varied between cranial and caudal glands as well as the phases of the estrous cycle. In conclusion, horizontal histological sections of the rat mammary gland provided significantly larger samples of mammary gland tissue with no difference in tissue composition compared to transverse sections, which are recommended in the RITA guidelines.