Imaging technologies are regularly used in biomedical research to study processes in living animals in a noninvasive manner. But imaging procedures can affect animal physiology, and the need to anesthetize animals for imaging entails potential health risks. In addition, certain imaging modalities require the use of ionizing radiation or the administration of contrast agents or imaging biomarkers, which also have consequences for animal physiology. Finally, procedures associated with imaging, such as animal preparation (e.g., fasting, premedication) and blood sampling, can also affect physiology and animal welfare. Here, the authors review the imaging modalities commonly used for rodents in biomedical research and their associated considerations for animal welfare.