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    Cardiovascular Engineering

    (G-260) Optogenetic stimulation can accelerate, slow, or suppress spontaneous beating in PSC-CMs: Proof-of-concept from computational simulations

    Saturday, October 14, 2023
    9:30 AM – 10:30 AM PDT
    Location: Exhibit Hall - Row G - Poster # 260
    Introduction::
    Myocardial infarction results in extensive cardiac remodeling that can lead to heart failure, morbidity, and mortality. Human pluripotent stem cell-derived cardiomyocyte (PSC-CM) injection can improve contractile function, potentially alleviating heart failure [1]. However, administration of this treatment is associated with ventricular arrhythmia in animal models, hindering progress toward clinical trials in humans [2]. Optogenetics offers a means of controlling cardiac electrophysiological activity via light-sensitive proteins (opsins). Light stimulation can be used to elicit bioelectric currents in opsin-expressing cardiac cells or tissue. Here, we present a computational study to establish the feasibility of using optogenetic stimulation to modulate the spontaneous beating rate of PSC-CMs. The ability to precisely control pacemaker-like PSC-CM activity could provide an important tool for reducing incidence and frequency of ventricular arrhythmias associated with cell therapy.


    Materials and Methods::
    We conducted simulations in the openCARP software package [3] using a modified version of a published human induced PSC-CM model with a fast spontaneous beating rate (~1.9 Hz) and a minimum diastolic potential of ~–72 mV [4]. In these virtual cells, we modeled expression of three different opsins: ChR2 (non-selective cation channel; reversal potential [Erev] ≈ 10 mV) [5], GtACR1 (chloride-conducting channel; Erev ≈ –40 mV) [6], and WiChR (potassium-conducting channel; Erev ≈ –90 mV) [7]. ChR2 and GtACR1 were simulated using published photocurrent models [5], [8]. To represent WiChR, we devised a “WiChR-like” model by adjusting the key parameters of the GtACR1 photocurrent model (Erev changed to –90mV, τoff changed to 350 ms, and responsive wavelength changed to blue light) to reflect published WiChR characteristics [7]. 

     

    To probe the feasibility of optogenetically modulating spontaneous beating rate in PSC-CMs, we imposed continuous 4000 ms-long light stimuli; these were compared to a control condition, in which the virtual cells were left completely unstimulated. For each opsin models, we simulated light at the appropriate excitation wavelength (488 nm for ChR2 and WiChR-like; 515 nm for GtACR1) and irradiance was calibrated to the working range for optogenetic stimulation (ChR2: 1-100 μW/mm2; GtACR1: 0.01-1 μW/mm2; WiChR-like: 0.001-1 μW/mm2). For light configurations that resulted in suppression of spontaneous activity, we delivered electrical stimuli to the optogenetically-silenced PSC-CM model to ascertain whether it was still possible to elicit action potentials.


    Results, Conclusions, and Discussions::
    Optogenetic stimulation of simulated PSC-CMs expressing either ChR2 or GtACR1 accelerated spontaneous beating from the baseline rate (1.9 Hz; Fig. 1A). For the examples shown, the hastened rates were 2.1 Hz (10 μW/mm2 ChR2; Fig. 1B) and 2.2 Hz (0.05 μW/mm2 GtACR1; Fig. 1C), respectively. Notably, the irradiances (Ee) used in these cases were relatively dim, below the threshold for optogenetically eliciting new action potentials [8]. Use of higher Ee values in GtACR1-expressing PSC-CMs suppressed spontaneous beating, but caused repolarization failure (i.e., the cells did not return to resting levels).  Simulated illumination of PSC-CMs expressing the WiChR-like opsin suppressed spontaneous excitation for Ee values above 0.1 μW/mm2 (e.g., 0.5 μW/mm2 WiChR-like; Fig. 1D) without inducing repolarization failure. As shown in Fig. 1E, weaker Ee values in WiChR-like simulations prompted slowed spontaneous PSC-CM excitation; the same figure shows the Ee dependency of light-induced acceleration for the ChR2 and GtACR1 cases.

     

    In the context of light-induced suppression of spontaneous activity in simulated PSC-CMs expressing the WiChR-like opsin, we confirmed that electrical stimuli could still elicit normal action potentials (Fig. 1F; same Ee as 1D). These action potentials had morphology that qualitatively resembled that of normal human ventricular excitation and had comparable electrophysiological features (APD90 ≈ 308 ms vs. normal human myocardial APD ≈ 330 ms [9]). This suggests that optogenetic activation of WiChR could fully suppress spontaneous beating in PSC-CMs while maintaining relatively unimpaired electrophysiological excitability and cellular function.


    In conclusion, our simulations establish that optogenetic modulation of PSC-CM spontaneous beating rate is theoretically feasible: we predict that ChR2- and GtACR1-based stimulation can hasten spontaneous beating while light-induced activation of WiChR can slow spontaneous beating or, at sufficiently elevated Ee, suppress it altogether. We also predict that optogenetic suppression of spontaneous beating with a WiChR-like opsin would not impair cellular excitability (e.g., normal activation by a propagating wavefront from host myocardium into engrafted PSC-CMs), as normal action potentials were elicited by electrical stimuli at the cellular scale.


    Acknowledgements (Optional): : This work was facilitated through the use of advanced computational, storage and networking infrastructure provided by the Hyak supercomputer system at the University of Washington. Funding support for this project was provided by NIH grant R01HL158667, by an NSF Graduate Research Fellowship under Grant No. DGE-2140004. Fellowship support came from the UW Institute for Stem Cell and Regenerative Medicine, the UW Mary Gates Endowment, and the UW Levinson Scholars Program.

    References (Optional): :

    [1] J. Chong, X. Yang, C. Don, E. Minami, B. Van Biber, and C. Murry, “PW281 Human Pluripotent Stem Cell Derived Cardiomyocytes Regenerate Infarcted Hearts Of Non-Human Primates,” Global Heart, vol. 9, no. 1, pp. e315–e316, Mar. 2014, doi: https://doi.org/10.1016/j.gheart.2014.03.2364.


    [2] Y.-W. Liu et al., “Human embryonic stem cell–derived cardiomyocytes restore function in infarcted hearts of non-human primates,” Nature Biotechnology, vol. 36, no. 7, pp. 597–605, Jul. 2018, doi: https://doi.org/10.1038/nbt.4162.


    [3] G. Plank et al., “The openCARP simulation environment for cardiac electrophysiology,” Computer Methods and Programs in Biomedicine, vol. 208, p. 106223, Sep. 2021, doi: https://doi.org/10.1016/j.cmpb.2021.106223.


    [4] C. E. Gibbs, S. Marchianó, K. Zhang, X. Yang, C. E. Murry, and P. M. Boyle, “Graft–host coupling changes can lead to engraftment arrhythmia: a computational study,” The Journal of Physiology, vol. 601, no. 13, pp. 2733–2749, Apr. 2023, doi: https://doi.org/10.1113/jp284244.


    [5] J. C. Williams et al., “Computational Optogenetics: Empirically-Derived Voltage- and Light-Sensitive Channelrhodopsin-2 Model,” PLoS Computational Biology, vol. 9, no. 9, p. e1003220, Sep. 2013, doi: https://doi.org/10.1371/journal.pcbi.1003220.


    [6] E. G. Govorunova et al., “The Expanding Family of Natural Anion Channelrhodopsins Reveals Large Variations in Kinetics, Conductance, and Spectral Sensitivity,” Nature Scientific Reports, vol. 7, no. 1, Mar. 2017, doi: https://doi.org/10.1038/srep43358.


    [7] J. Vierock et al., “WiChR, a highly potassium-selective channelrhodopsin for low-light one- and two-photon inhibition of excitable cells,” Science Advances, vol. 8, no. 49, Dec. 2022, doi: https://doi.org/10.1126/sciadv.add7729.


    [8] A. R. Ochs, T. V. Karathanos, N. A. Trayanova, and P. M. Boyle, “Optogenetic Stimulation Using Anion Channelrhodopsin (GtACR1) Facilitates Termination of Reentrant Arrhythmias With Low Light Energy Requirements: A Computational Study,” Frontiers in Physiology, vol. 12, Aug. 2021, doi: https://doi.org/10.3389/fphys.2021.718622.


    [9] M. Franz, C. D. Swerdlow, L. B. Liem, and J. Schaefer, “Cycle length dependence of human action potential duration in vivo. Effects of single extrastimuli, sudden sustained rate acceleration and deceleration, and different steady-state frequencies.,” The Journal of Clinical Investigation, vol. 82, no. 3, pp. 972–979, Sep. 1988, doi: https://doi.org/10.1172/jci113706.