Abstract Body: Calcium ion (Ca²⁺) is one of key components in the heart excitation and conduction, and its overload is known to induce abnormal impulse propagation and generation, enabling to lead lethal arrhythmias. However, unknown is how large area of Ca²⁺-overload in a cardiac tissue is required to provoke such abnormalities.
By utilizing UV-photolysis of caged Ca²⁺, we investigated whether the increase of Ca²⁺ concentration ([Ca²⁺]) at the local on a cardiomyocytes monolayer alters its spatiotemporal impulse patterns, and also assessed its dependence on area-sizes of the Ca²⁺-overload. The monolayers of cardiomyocytes isolated from neonatal rats were prepared on quartz-substrates (φ27 mm) with 2 days incubation, then treated with a fluorescent indicator of Ca²⁺ (Fluo-8/AM: 5 µM) and a caged Ca²⁺ (DMNPE-4/AM: 5 - 50 µM). Its excitation manner and impulse propagation were monitored as fluorescence images by a macro-zoom microscope (image size: ~1,500 mm², frame rate: 400 frame/s). Under monitoring, UV light (λ: 365 nm) for flash photolysis of caged Ca²⁺ was irradiated to a limited area (diameters of 3 - 9 mm) on the cell layer, with and without electrical pacing.
Under periodical excitation by pacing, the delay of impulse propagation was detected at UV-irradiated area on the monolayer. The delay was enhanced with an increase of [Ca²⁺] under the photolysis, but gradually dissolved with a decay of [Ca²⁺] after stopping UV irradiation. Under non-pacing, on the other hand, it was found that abnormal impulses were generated from the UV-irradiated area as a result of increase of [Ca²⁺]. Probabilities about the abnormal impulses showed a linear relationship with the area size of UV-photolysis. Also, we confirmed that the abnormal impulses by the UV-photolysis provoked a reentry-like excitation manner when the electrical conductivity of cardiomyocytes was further degraded by an administration of carbenoxolone.
In conclusion, we demonstrated altering impulse propagation/generation of cardiomyocytes monolayer by the local frash photolysis of caged Ca²⁺, and investigated its dependence on spatial conditions of the overload region. The target of this study will be shifted to whole hearts as one on next steps in order to provide the statistical insight in arrhythmogenesis.
This research was partially supported by JSPS(18K19459, 22K18174) and JST-CREST(JPMJCR 1925).