bioRxiv [Preprint]. 2026 Feb 13:2026.02.11.705466. doi: 10.64898/2026.02.11.705466.
ABSTRACT
Human iPSC-derived engineered heart tissues (EHTs) and cardiac organoids are increasingly used to model cardiac physiology and drug responses, yet it remains unclear whether they can reproduce tissue-scale mechanisms underlying ventricular arrhythmias. Advanced electrophysiological characterization of EHTs has been limited by the lack of a recording framework compatible with small, perfused preparations, despite the availability of mapping hardware. We present a reproducible workflow that couples milliPillar-based EHT fabrication with high-resolution (22 µm spatial, 1 ms temporal) dual channel optical mapping using RH-237 for voltage and Rhod-2 AM for Ca²⁺. Baseline electrophysiological measurements align with published data from human and animal cardiac tissues, showing rate-dependent restitution of action potential and Ca²⁺ transient duration (APD and CaD), conduction slowing at higher pacing rates, and physiologic AP-Ca²⁺ activation latency. Selective hERG blockade with E-4031 prolongs APD, confirming pharmacological sensitivity. To interrogate mechanisms underlying ventricular tachycardia (VT), we utilized an established proarrhythmic perturbation widely validated in animal models of acquired long-QT syndrome, combining hERG inhibition with hypokalemia and hypomagnesemia; electrolyte disturbances commonly encountered in clinical settings. Treated EHTs (VT group) displayed tachyarrhythmic contractile bursts with marked beat-to-beat instability, whereas controls responded synchronously to field stimulation. Beat-resolved optical mapping revealed progressive diastolic interval shortening, APD dispersion with transient localized long-short APD zones, and regional depression of excitability that together formed spatial conduction barriers precipitating wavebreak and reentry. Early afterdepolarizations contributed to triggered activity and created localized repolarization barriers at long-short APD zones, leading to rotor formation. Phase singularity tracking identified short-lived rotors localized predominantly in the heads of VT EHTs and absent in controls. A minority of tissues exhibited multiple simultaneous rotors and wavelets generating chaotic-like activation. Although tissue acceleration promoted rotor formation, these events were brief, likely due to the spatial limitations of EHTs, and treated tissues more closely resembled VT than Torsades de Pointes or sustained fibrillation. Our comprehensive studies demonstrate that human iPSC-derived EHTs can recapitulate the tissue-scale VT mechanisms associated with acquired long QT syndrome - APD dispersion with long-short APD zones, triggered activity, conduction block, wavebreak and reentry - which had previously been assessed only in intact hearts. The presented platform thus provides a scalable, non-animal system for mechanistic arrhythmia research.
PMID:41727060 | PMC:PMC12919079 | DOI:10.64898/2026.02.11.705466