Selective Inhibition of Cardiac Late Na+ Current Is Based on Fast Offset Kinetics of the Inhibitor

Figures

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  Figure 1. Averaged analogue current records exhibited by 1 µM GS967, 20 µM mexiletine and 10 µM quinidine under action potential voltage clamp conditions in isolated canine ventricular myocytes (A). The stimulation cycle length was 700 ms. The command AP is shown above the current traces. Dashed lines indicate zero voltage and current levels. (B) Current densities measured at 50% duration of APD90 of the command AP. (C) Current integrals from which the initial 20 ms period was excluded. Symbols and bars are the mean ± SEM, small dots represent individual data, and numbers in parentheses indicate the number of myocytes studied.

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  Figure 2. Representative superimposed analogue records (left) and average INaL densities, measured at 50 ms, (right) demonstrating the effects of 1 µM GS967 (A), 20 µM mexiletine (B) and 10 µM quinidine (C) on INaL under conventional voltage clamp conditions using test pulses of 2 s duration clamped to −20 mV from the holding potential of −120 mV. At the end of each experiment, the cells were superfused with 20 µM TTX to dissect the remaining INaL. Dashed lines indicate zero current level. Symbols and bars are the mean ± SEM, small dots represent individual data, numbers in parentheses indicate the number of myocytes studied, and asterisks denote significant differences between the pre-drug and post-drug values.

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  Figure 3. Effects of 1 µM GS967 (A), 20 µM mexiletine (B) and 10 µM quinidine (C) on the maximal rate of depolarization (V+max) measured in canine right ventricular trabeculae under steady-state conditions. Symbols and bars are the mean ± SEM, numbers in parentheses indicate the number of preparations studied, and asterisks denote significant differences between the pre-drug control and post-drug values.

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  Figure 4. Left: Determination of the offset kinetics of GS967 (A), mexiletine (B) and quinidine (C) as indicated by the time-dependent restitution of V+max. Solid lines were obtained by monoexponential fitting of the data from 0 to 5 s for GS967 and mexiletine, and from 0 to 60 s for quinidine. Right: Determination of the onset kinetics of V+max block for GS967 (D), mexiletine (E) and quinidine (F) using trains of 2.5 Hz applied following a 1 min stimulation-free period. Solid lines were obtained by monoexponential fit. Symbols and bars are the mean ± SEM, and numbers in parentheses indicate the number of preparations studied. The experimental protocols are shown in the insets (top).

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  Figure 5. Relationship between the magnitude of the V+max block and inhibition of INaL plotted as a function of the respective offset time constant obtained for GS967, mexiletine and quinidine, as documented in Figure 4. (A) Results obtained from sharp microelectrode and conventional voltage clamp experiments (Figure 2 and Figure 3), where the ratio of the percentage V+max block was normalized to the percentage inhibition of INaL density and its integral. (B) Results obtained from sharp microelectrode and APVC experiments (Figure 1 and Figure 3), where the absolute magnitude of the V+max block was divided by the density and integral of the excised INaL. Open circles and filled triangles represent data regarding the current densities and integrals, respectively.