Connexin 43-Enriched Vesicles Improve Synchronization in hiPSC-Derived Cardiomyocytes

Human induced pluripotent stem cell-derived cardiomyocytes are valuable for studying cell-cell communication and synchronization, but remain immature and often lack robust electrical and mechanical coupling. To address this, we investigated gap junction-mediated communication and developed plasma membrane vesicles enriched in functional connexin hemichannels, termed Connectosomes, to enhance intercellular coupling. Connectosomes display properly oriented connexins and enrich the Cx43 expression at cell-cell borders between cardiomyocytes. Through mathematical modeling and experimental validation, we demonstrate that Connectosome incorporation reinforces endogenous gap junctions, promotes synchronous calcium transients, and improves spatial coordination of beating across networks. Mechanistic studies using engineered cell lines with tagged connexin-43 confirm that channel orientation and functionality are critical, supporting a model in which Connectosomes contribute to gap junction coupling. These results show that Connectosomes can synchronize the beating of immature cardiomyocytes by boosting electrochemical communication, laying the groundwork for future therapeutic advances.