The heart is a contractile organ, without which its purpose is lost. In a number of disease states, this contractility is either reduced or completely eliminated, often in regional ways. Studying contractility at the level of the whole organ can only tell us its output and may hide a number of subtler changes occurring at the intra-organ level: regional ischemia; apex-base axis differences in catecholamine sensitivity; or endo- to epicardial changes in calcium transient duration. Therefore, when we have the opportunity in the pre-clinical world to study the whole heart, efforts must be made to study regional differences, as we know occurs in human disease. This methods collection will highlight novel methods of studying regional differences in the animal model heart, including echocardiography, optical mapping, and single-cell studies requiring the careful isolation of different heart regions.