Selective inhibition of LY294002 across cloned human cardiac ion channels: implications for atrium-selective therapy in atrial fibrillation

Figures

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  FIGURE 1. Workflow for assessing the selective inhibition of LY294002 on I Kur. (A) Schematic overview of the experimental workflow: a panel of major cardiac ion channels is depicted; CHO cells heterologously expressing Kv1.5 were established and cultured for subsequent whole-cell patch-clamp recordings. (B) Whole-cell patch-clamp recording and dosing paradigm: LY294002 was applied using a cumulative application protocol, and current changes over time/concentration were recorded to generate inhibition profiles. (C) Bar plot comparing IC50 values across ion channels: LY294002 showed the lowest IC50 toward I Kur (Kv1.5), whereas markedly higher IC50 values were observed for other cardiac ion channels, indicating a relatively selective inhibitory effect on I Kur. (D) Molecular docking: LY294002 adopted a stable binding pose within the channel pocket with favorable predicted binding energy and was predicted to engage in hydrogen bonding, hydrophobic/covalent contacts, and π–π conjugation interactions, providing a molecular rationale for its inhibitory activity.

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  FIGURE 2. The inhibitory effect of LY294002 on I Ks currents in CHO cells. (A) Superimposed I Ks current traces elicited on repolarization to −50 mV following 2 s depolarization from a holding potential −80 mV to +30 mV in the presence of various LY294002 concentrations (6, 12.5, 25, 50 and 100 µM) in a cumulative manner. (B) The concentration-response relation for the LY294002 inhibition on I Ks. The solid line represents the fit of the data to the Hill equation, with an IC50 and n H shown in the inset.

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  FIGURE 3. The inhibitory effect of LY294002 on I Kr currents in CHO cells. (A) Superimposed I Kr current traces elicited by applying 2 s depolarizing pulses from a holding potential of −80 mV to +10 mV and then repolarization to −60 mV, in the presence of various LY294002 concentrations (15, 20, 30, 50, 70 and 100 µM). (B) The concentration-response relation for the LY294002 inhibition on I Kr. The solid line represents the fit of the data to the Hill equation, with an IC50 and n H shown in the inset.

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  FIGURE 4. The inhibitory effect of LY294002 on I to currents in CHO cells. (A) Superimposed I to current traces elicited by applying 300 ms depolarizing pulses from a holding potential of −80 mV to +50 mV in the presence of various LY294002 concentrations (25, 50, 100 and 150 µM). (B) Concentration-response relation for the inhibition of I to peak currents by LY294002. The solid line represents the fit of the data to the Hill equation, with an IC50 and n H shown in the inset.

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  FIGURE 5. The inhibitory effect of LY294002 on I K1 currents in CHO cells. (A) Superimposed I K1 current traces evoked by applying a 150 ms square pulse from a holding potential of −80 mV to −120 mV in the presence of LY294002 at various concentrations (50, 100, 150, 200 and 300 µM). (B) Concentration-response relation for the inhibition of I K1 currents by LY294002. The solid line represents the fit of the data to the Hill equation, with an IC50 and n H shown in the inset.

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  FIGURE 6. The inhibitory effect of LY294002 on I Na currents in CHO cells. (A) Superimposed I Na current traces elicited by applying 200 ms depolarizing pulses from a holding potential of −80 mV to +20 mV in the presence of various LY294002 concentrations (25, 35, 70, 100 and 150 µM). (B) Concentration-response relation for the inhibition of I Na currents by LY294002. The solid line represents the fit of the data to the Hill equation, with an IC50 and n H shown in the inset.

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  FIGURE 7. The inhibitory effect of LY294002 on I Ca-L currents in CHO cells. (A) Superimposed I Ca-L current traces elicited by 300 ms depolarizing voltage to 0 mV from a holding potential of −80 mV in the presence of various LY294002 concentrations (15, 25, 50, 100, and 150 µM). (B) Concentration-response relation for the inhibition of I Ca-L currents by LY294002. The solid line represents the fit of the data to the Hill equation, with an IC50 and n H shown in the inset.

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  FIGURE 8. Molecular docking of LY294002 within the tetrameric Kv1.5 pore cavity.