Ground-penetrating radar (GPR) has a wide range of applications, from geologic mapping to concrete inspection. A recently emerging GPR application is deployment in biological investigations as a non-invasive technique. Geophysical mapping of features such as tree roots and turtle burrows has proved valuable for the understanding of these subsurface systems for ecological, environmental, or engineering purposes. Four case histories of GPR investigations pertaining to animal burrows are described: cutter ants in Brazil, groundhogs in Michigan, and groundhogs, and burrowing bees in Pennsylvania. Cutter ants (Atta spp.) in Amazonian Brazil are known to construct burrows of nearly the same dimensions as groundhogs as they excavate galleries up to 7 m deep for leaf storage. Cutter ant burrows are hazardous to heavy equipment and may also cause loss of mud circulation during rotary drilling. Groundhogs (Marmota monax), found throughout the United States, cause unseen hazards, particularly for equestrian facilities where a sudden collapse can cause severe injuries to both horse and rider. Burrowing bees (Colletes inaequalis) are common in the northeastern United States. The size of the bee burrows is significantly smaller than that of the cutter ants and the groundhogs. The data for these surveys were collected over a twenty-year span, crossing several generations of survey equipment and processing techniques. Together, these four case histories highlight the historic and current capabilities of GPR systems applied to mapping subsurface burrow systems. These examples demonstrate the important impact near surface heterogeneities have in altering ecological, environmental, or engineering systems and the utility of GPR for mapping such heterogeneities.