β-adrenergic stimulation augments transmural dispersion of repolarization via modulation of delayed rectifier currents IKs and IKr in the human ventricle

Figures

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  Figure 1. Experimental Preparations. Section of the left ventricular marginal area from a donor human hearts was dissected into a coronary perfused wedge preparation. (a) Orientation of the preparation was labeled accordingly. Color squares indicate recordings sites of the representative optical traces, illustrating that the tissue was fully perfused on the transmural surface. (b) Representative traces of how different repolarization % changes APD and ∆APD between conditions were calculated. (c) A third order sigmoidal curve was fitted to ∆APD80 vs Log (nM) for the dose response of JNJ-303. Log(IC50) was obtained as 1.82 ± 0.13 (±S.E.). 95% confidence interval for IC50 in nM was between 35.4 and 128.4 nM. Best fit value was determined to be 66.7 nM.

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  Figure 2. Percent Repolarization. Different pharmacologic interventions targeting specific ion channels led to changes in APD morphology. (a) Box-and-whisker (B&W) plots of % ∆APD80 from baseline to isoprenaline indicate that APD did not alter among different % repolarization. (b) IKs blockade with JNJ-303 linearly increased % ∆APD80 as % repolarization increased, indicating that IKs was more prominent at the latter phases of AP. (c) IKr block with E-4031 increased % ∆APD80 as % repolarization was increased, peaking at approximately 80–85% to indicate that IKr contributes the most at 80–85% repolarization.

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  Figure 3. Conduction Properties. Conduction velocity (CV) was calculated from the transmural surface of the preparation. (a) A typical elliptical activation pattern was observed. Directions of activation (longitudinal, transverse) indicate the fastest and slowest conduction, respectively. (b) A representative activation map from protocol B shows that the activation pattern was the same both before and after application of JNJ-303. (c) Representative activation maps from protocol C indicate that conduction was increased with the application of isoprenaline. This change in activation was maintained with the addition of JNJ-303 and E-4031. (d) No significant difference was observed between transverse and longitudinal conduction velocities (CVT, CVL) in protocol B, and the anisotropy ratio was not altered (d). (f) Both CVT and CVL increased significantly with isoprenaline, but no other significant changes from previous conditions were observed. However, significant differences to the baseline were apparent. (g) Conduction anisotropy increased with isoprenaline, slightly decreased with JNJ-303, and remained unchanged with E-4031.

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  Figure 4. APD80 Comparisons. APD was measured at 80% of repolarization. (a) APD80 maps for both conditions in protocol B show that JNJ-303 increased APD80 slightly (but not significantly), as indicated by the red color. (b) Representative traces show changes in APD80 distribution after application of JNJ-303. Blue indicates baseline and red indicates the pharmacologic intervention. (c) APD80 maps for all conditions in protocol C highlight that isoprenaline decreased APD80, JNJ-303 increased APD80 above baseline, and E-4031 dramatically increased APD80. (d) Representative traces for each condition in protocol C show distribution histograms for three pairs of comparisons: baseline vs. isoprenaline, isoprenaline vs. JNJ-303, and JNJ-303 vs. E-4031. Bars in blue are representative of previous treatment, while bars in red are representative of post treatment. (e) B&W plots indicate a slight but not significant increase in APD80 with IKs block. (f) Significant reduction of APD80 was observed with application of isoprenaline, and an increase of APD80 was observed with both IKs and IKr block. (g,h) ∆APD80 in both ms and % of pre and post JNJ-303 with or without isoprenaline stimulated β-ARs.…

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  Figure 5. Restitution Properties. Using an S1S1 pacing protocol, restitution curves show that APD80 decreased as pacing cycle length decreased. This relationship was fitted using a two phase exponential curve. (a) The restitution curve shifted minimally with the addition of JNJ-303 in isolation. (b) Restitution downshifted with isoprenaline, upshifted with IKs block, and further upshifted with IKr block. (c) Functional refractory period was reached when S1S1 pacing lost capture. The refractory period significantly decreased with isoprenaline, and increased with both IKs and IKr blockers. (d–f) Restitution curves were plotted for endo, mid, and epi areas of the transmural surface indicating similar changes across all three areas.

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  Figure 6. Transmural Dispersion. Absolute ∆APD80 was calculated in milliseconds from epi, mid, and endo areas of the transmural surface. (a) No significant difference was observed from ∆APD80 across the transmural surface post IKs blockade in protocol B. (b) ∆APD80 was calculated as the difference between post and pre-treatment. APD80 reduction from isoprenaline was significantly greater in the epicardium as compared to the endocardium. On the other hand, APD80 under IKr blockade increased more significantly in the midmyocardium and endocardium as compared to the epicardium. (c) Normalized ∆APD80 was calculated as the difference between post treatment and baseline. The transmural gradient was increased by both isoprenaline and E-4031, a trend which was also observed in comparing pre- and post-treatments.

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  Figure 7. Computational Modeling of IKs and IKr Block with β-AR Stimulation. Protocol C was repeated in a computational model. The top panel displays AP traces obtained by sequential application of various pharmacologic interventions in the model. Baseline experimental AP traces were accurately reproduced by the model. β-AR stimulation was produced by an increased phosphorylation of all ion channels. The Model was also able to reproduce experimental data for IKs block at 80% current blockade. However, IKr blockade could not be reproduced using the modified O’Hara-Rudy model. When IKr block exceeded 20% early afterdepolarizations (EAD) were observed. EADs were not observed in any preparation even after saturated IKr block.