Evaluation of Intracellular Motion in Living Cardiomyocytes by a Multi-Microsphere 3D Sensing System

Precise quantification of cardiomyocyte mechanical motion has substantially advanced the cardiac physiology research. Dysregulated rhythmic contractions in cardiomyocytes are predominantly driven by intracellular structural perturbations. However, high-resolution three-dimensional (3D) mapping of intracellular motion within cardiomyocytes remains a technical challenge. We herein developed an innovative intracellular 3D motion-sensing system integrated with a stable motion-tracking algorithm to quantify intracellular contraction trajectories. Using this system, we observed that the propagation velocity of the intracellular motion decreases from the cardiomyocyte center to the periphery. These velocity changes correlate with constraints imposed by the cell membrane and cytoskeleton, enabling our method to detect subtle cellular alterations. To validate this system, we induced cytoskeletal depolymerization with cytochalasin D, resulting in a marked reduction in the cellular motion velocity. Collectively, these findings demonstrate that our intracellular motion sensing system offers a novel platform for in vitro single-cell multiposition motility assessment, toxicity screening, and disease-associated phenotype detection.