Rate dependency of delayed rectifier currents during the guinea-pig ventricular action potential

Abstract

1. The action potential clamp technique was exploited to evaluate the rate dependency of delayed rectifier currents (I(Kr) and I(Ks)) during physiological electrical activity. I(Kr) and I(Ks) were measured in guinea-pig ventricular myocytes at pacing cycle lengths (CL) of 1000 and 250 ms. 2. A shorter CL, with the attendant changes in action potential shape, was associated with earlier activation and increased magnitude of both I(Kr) and I(Ks). Nonetheless, the relative contributions of I(Kr) and I(Ks) to total transmembrane current were independent of CL. 3. Shortening of diastolic interval only (constant action potential shape) enhanced I(Ks), but not I(Kr). 4. I(Kr) was increased by a change in the action potential shape only (constant diastolic interval). 5. In ramp clamp experiments, I(Kr) amplitude was directly proportional to repolarization rate at values within the low physiological range (< 1.0 V s(-1)); at higher repolarization rates proportionality became shallower and finally reversed. 6. When action potential duration (APD) was modulated by constant current injection (I-clamp), repolarization rates > 1.0 V s(-1) were associated with a reduced effect of I(Kr) block on APD. The effect of changes in repolarization rate was independent of CL and occurred in the presence of I(Ks) blockade. 7. In spite of its complexity, the behaviour of I(Kr) was accurately predicted by a numerical model based entirely on known kinetic properties of the current. 8. Both I(Kr) and I(Ks) may be increased at fast heart rates, but this may occur through completely different mechanisms. The mechanisms identified are such as to contribute to abnormal rate dependency of repolarization in prolonged repolarization syndromes.

Figure 1. Rate dependency of ErgTx-sensitive current (IKr)

A and B, examples of action potential and I_ErgTx recorded at a cycle length (CL) of 1000 (A) and 250 ms (B) according to protocol A (Methods); the two CLs were tested within the same cell. C-F, comparison between cycle lengths of 1000 and 250 ms; mean repolarization current density (C), time-to-peak current (D), peak current density (E) and the ratio between I_ErgTx and total repolarizing current (I_tot, see text for definitions, F). C-F, mean data from 7 cells; *P < 0.05.

Figure 2. Rate dependency of 293B-sensitive current (IKs)

A and B, examples of action potential and I_293B recorded at a cycle length (CL) of 1000 (A) and 250 ms (B) according to protocol A; the two CLs were tested within the same cell. C-E, comparison between cycle lengths of 1000 and 250 ms for mean repolarization current density (C), time course of I293B conductance (g_293B) during the action potential (dynamic conductance, D) and the ratio between I_293B and total repolarizing current (I_tot, E). In D, mean traces (thick lines) and their confidence limits (thin lines) are shown; the arrows mark instantaneous conductance. C-E, mean data from 13 cells; *P < 0.05.

Figure 3. Dependency of ErgTx-sensitive current (I_Kr) on diastolic interval only

A, the same action potential waveform (recorded at CL of 1000 ms) was applied to measure I_ErgTx at different diastolic intervals (DI) as in protocol B (Methods); two DIs were tested within the same cell. Values of mean repolarization current density (B) and peak current density (C) measured at the two DIs are compared (n = 7).

Figure 4. Dependency of 293B-sensitive current (I_Ks) on diastolic interval only

A, the same action potential waveform (recorded at CL of 1000 ms) was applied to measure I_293B at different diastolic intervals (DI) as in protocol B (Methods); two DIs were tested within the same cell. Values of mean repolarization current density measured at the two CLs are compared in B (n = 4). *P < 0.05.

Figure 5. Dependency of ErgTx-sensitive current (I_Kr) on action potential contour only

Action potential waveforms were recorded at two CLs (A) and applied with the same diastolic interval (B) to record I_ErgTx as in protocol C (Methods); two action potential waveforms were tested within the same cell. C, time course of I_Kr gating variables; D, I_Kr amplitude predicted by applying the action potential waveforms (from A) to the numerical model. The ratio between mean currents at the two CLs is specified for measured I_ErgTx and for simulated I_Kr above the respective panels.

Figure 6. Dependency of E-4031-sensitive current (I_Kr) on the rate of repolarization

A, I_E-4031 recorded, by applying the voltage protocol shown in the inset, during ramps of variable steepness (dV/dt). B, model simulation of the experiment of A. C, peak I_E-4031 magnitude recorded from 9 cells as a function of ramp dV/dt (•); in order to eliminate differences between cells, I_E-4031 magnitude is expressed as the percentage of the value observed at dV/dt = 1.5 V s⁻¹, which was measured in all cells. The continuous line represents the relationship independently predicted by the model. D, effect of changes in the time constant of inactivation (τ) on the predicted relation between I_Kr amplitude and dV/dt (data expressed as C.

Figure 7. Dependency of E-4031 modulation of APD on repolarization rate

Effect of I_Kr inhibition (3 μm E-4031) on action potentials with repolarization rates (dV/dt)≤ 1 V s⁻¹ (A) and > 1 V s⁻¹ (B). C, E-4031-induced APD prolongation in cells with dV/dt≤ 1 V s⁻¹ and > 1 V s⁻¹. Action potential duration was modulated by constant current injection during the repolarization phase. Cells were paced at constant cycle length (2 s); I_Ks was blocked (10 μm chromanol 293B) throughout the experiment.

Figure 8. Voltage dependency of I_Kr and of its gating variables as predicted by the numerical model

A, I_Kr activated during depolarizing steps at -30 to +10 mV from a holding of -40 mV. B, voltage dependency of gating variables at steady state (E_0.5 = mid potential). C and D, voltage dependency of the time constants (τ) of activation and inactivation, respectively.