KairoSight-3.0: A validated optical mapping software to characterize cardiac electrophysiology, excitation-contraction coupling, and alternans

Figures

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  Fig. 1. KairoSight-3.0 overview. A) Illustration of KairoSight-3.0 workflow. B) KairoSight-3.0 graphical user interface. Action potentials on the right correspond to selected pixels (colored dots) on the epicardial surface. A time window of interest is selected (red bars) to highlight a single action potential. C) Example of KairoSight-3.0 analysis features, including a masking tool to select a region of interest (left), and the generation of optical maps (right) corresponding to action potential duration (APD), calcium transient duration (CaD), conduction velocity (CV), action potential alternans coefficient (AltAP), calcium coefficient from extrasystolic stimulation (ΔCaD50), action potential-calcium transient coupling coefficient (ACC). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

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  Fig. 2. Validation of semi-automated software analysis. Optical signals were analyzed using KairoSight-3.0, and the accuracy was compared with manual measurements. A-C) Maps of APD and CaD derived manually, using KairoSight-3.0 software, and the percent accuracy of the automated measurement. D–F) Analysis of alternans from sequential APD measurements (odd, even beats) at a specified repolarization phase (50 %, “fixed”), with corresponding coefficient measurement (AltAP). Results derived manual, using KairoSight-3.0 software, and the percent accuracy of the automated measurement. G–I) Analysis of alternans from sequential APD measurements (odd, even beats) at the maximum detected repolarization phase between consecutive beats (“dynamic”), using manually analysis, KairoSight-3.0 software, and the percent accuracy of the automated measurement. Note the longer duration time compared to “fixed” approach indicates repolarization >50 %. An example of calcium alternan coefficient (AltCa) map is also shown. Optical maps derived from an intact rat heart loaded with a voltage- or calcium-sensitive dye. Anatomical orientation denoted (RV: right ventricle, LV: left ventricle).

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  Fig. 3. Impact of signal quality and filtering on data analysis. A) Reducing the excitation light intensity (e.g., 40–100 %) results in a lower signal-to-noise ratio (note difference in scale). Post-processing can be applied (e.g., temporal filter, spatial averaging) to aid in the measurement of B) conduction velocity, C) APD at 30 % repolarization, and D) APD at 80 % repolarization. Example of optical maps derived from an intact, rat heart loaded with a voltage-sensitive dye. Anatomical orientation denoted (RV: right ventricle, LV: left ventricle).

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  Fig. 4. Mapping of cardiac alternans. A) AltAP maps generated from two sequential action potentials using the “fixed” alternan tool at varying phases of repolarization (20–60 % APD). Top: AltAP at a slower pacing rate (200 ms PCL); Bottom: AltAP at faster pacing rate (100 ms PCL). B) APD measurements for sequential (odd, even) action potentials and the calculated AltAP using the “dynamic” alternan tool. Variables measured at a slower (200 ms PCL) and faster rate (100 ms PCL). C) AltCa maps generated from two sequential calcium transients using the “dynamic alternans” tool. AltCa derived at a slower (200 ms PCL) and faster rate (100 ms PCL). Optical maps derived from an intact, guinea pig heart loaded with a voltage- or calcium-sensitive dye. AltAP = AP alternans coefficient, AltCa = calcium alternans coefficient. Anatomical orientation denoted (RV: right ventricle, LV: left ventricle).

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  Fig. 5. Effect of extrasystolic stimulation on cardiac metrics. A) Top: example of optical action potential recordings resulting from regular (S1: 200 ms PCL) and extrasystolic stimulation (S2: 140 ms PCL). Bottom: corresponding optical maps for APD50 during S1, S2, and the map of the resulting difference. B) Top: example of optical action potential recordings resulting from regular (S1: 200 ms PCL) and extrasystolic stimulation (S2: 140 ms PCL). Bottom: Corresponding optical maps for CaD50 during S1, S2, and the map of the resulting difference. C) Corresponding optical maps for CaT amplitude during S1, S2, and the map of the resulting difference. D) Difference maps of APD50 as S2 PCL is decremented (180–110 ms). Note different scale bar at faster frequencies. E) Difference maps of CaD50 as S2 PCL is decremented (180–110 ms). Fluorescent signals collected from guinea pig hearts. Anatomical orientation denoted (RV: right ventricle, LV: left ventricle).

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  Fig. 6. Conduction velocity measurements according to pacing location. A) Top: Illustration of septal pacing site. Middle: examples of electrical propagation in the mid, longitudinal, and transverse directions. Bottom: example conduction velocity measurements collected from six vectors. B) Top: illustration of apical pacing site. Middle: example conduction velocity measurements half-way across the epicardial surface. Bottom: example conduction velocity measurements from apex to base. C) Top: illustration of pacing site at the left ventricular base (LVB). Middle: example conduction velocity measurements half-way across the epicardial surface. Bottom: example conduction velocity measurements across the entire heart. D) Top: illustration of pacing site at the right ventricular base (RVB). Middle: example conduction velocity measurements half-way across the epicardial surface. Bottom: example conduction velocity measurements across the entire heart. Scale bar denotes activation time; fluorescent signals collected from rat hearts.

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  Fig. 7. Action potential-calcium transient coupling coefficient (ACC). A) Illustration of ACC measurements, with overlay of optical action potentials (green) and intracellular calcium transients (red). B) ACCact maps generated in response to slow (200 ms PCL) or fast pacing rate (120 ms PCL). C) Multiple ACCrep maps derived at different phases of repolarization (30–80 %), in response to slow (200 ms PCL) or fast pacing rate (120 ms PCL). ACCact = action potential-calcium transient coupling coefficient during activation, ACCrep = action potential-calcium transient coupling coefficient during repolarization. D) Effect of atrial removal on action potential-calcium transient coupling during activation (ACCact) and repolarization (ACCrep30%, ACCrep70%). Anatomical orientation denoted (RV: right ventricle, LV: left ventricle). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)