In the healthy kidney, specialized cells called podocytes form a sophisticated blood filtration apparatus that allows excretion of wastes and excess fluid from the blood while preventing loss of proteins such as albumin. To operate effectively, this filter is under substantial hydrostatic mechanical pressure. Given their function, it is expected that the ability to apply mechanical force is crucial to the survival of podocytes. However, to date, podocyte mechanobiology remains poorly understood, largely because of a lack of experimental data on the forces involved. We perform quantitative, continuous, nondisruptive, and high-resolution measurements of the forces exerted by differentiated podocytes in real time using a recently introduced functional imaging modality for continuous force mapping. Using an accepted model for podocyte injury, we find that injured podocytes experience near-complete loss of cellular force transmission but that this loss of force is reversible under certain conditions. The observed changes in force correlate with F-actin rearrangement and reduced expression of podocyte-specific proteins. By introducing robust and high-throughput mechanical phenotyping and by demonstrating the significance of mechanical forces in podocyte injury, this research paves the way to a new level of understanding of the kidney. In addition, in an advance over established force mapping techniques, we integrate cellular force measurements with immunofluorescence and perform continuous long-term force measurements of a cell population. Hence, our approach has general applicability to a wide range of biomedical questions involving mechanical forces.