Mechanical Stimulation on Cardiac Cells with Feedback Control System

Mechanical cues encoded in the extracellular microenvironment play essential roles in regulating cardiac rhythm and tissue-level coordination; however, systematic interrogation of when mechanical inputs are most effective remains technically challenging. While substrate stiffness and cyclic stretch are widely used to condition cardiomyocytes in vitro, most platforms lack the temporal precision required to probe phase-dependent mechanosensitivity during spontaneous beating. Here, we report a biomaterials-based mechanical stimulation platform that enables real-time, phase-specific delivery of localized stretch to autonomously beating cardiomyocyte aggregates cultured on soft polydimethylsiloxane (PDMS) substrates. By integrating live imaging-based beat detection with piezo-driven microscale actuation, mechanical stimuli were triggered at defined fractions of the intrinsic interbeat interval (IBI), allowing direct comparison of in-phase and out-of-phase mechanical perturbations. Using one-dimensional cardiomyocyte aggregates as a model system, we show that randomly timed mechanical stimulation rarely altered intrinsic rhythm (∼0.3% of beats). In contrast, phase-locked stimulation applied near the end of the contraction-relaxation cycle (∼90% of the IBI) reproducibly induced transient IBI shortening, reversed contraction-wave propagation, and shifted pacemaker activity toward the site of stimulation. Mechanical stimulation delivered at midcycle (∼50% of the IBI) produced minimal effects. Displacement-resolved analysis revealed that phase-specific stimulation redistributed mechanical output within the aggregate: contraction amplitude was maintained or enhanced near the stimulation site while being attenuated in distal regions. These results demonstrate that developing cardiac tissues exhibit narrow temporal windows of mechanical susceptibility and robust resistance to perturbation outside these phases. This platform provides a versatile biomaterials-enabled framework for dissecting dynamic mechano-electrical feedback, optimizing mechanical conditioning strategies, and engineering cardiac tissues with controllable rhythmic properties.