hERG1b consists of heteromeric channels rather than two separate pools of homomeric hERG1a and hERG1b channels. A hERG1 Markov Model Reproduces the Experimental Data The Markov model of hERG1, developed by Fink et al. [15], is able to reproduce hERG1 currents recorded both at room temperature as well as at physiological temperature. The relative changes in kinetics determined from the oocyte experiments reflecting different ratios of hERG1a and hERG1b were incorporated into the Markov model to investigate whether the model could reproduce the experimental findings. The factors by which the rate constants were changed are given in table S2. Like most hERG1 Markov models, the present model uses a single- exponential function to describe the transition from the open to the first closed state (O to C, transition rate b ) while the 3 deactivation process experimentally is most commonly best fit with a double-exponential function. Changes in the model deactivation kinetics were based on the relative changes in the fast component of deactivation obtained from the experiments. The model was subjected to the same voltage clamp protocol that was used in the in vitro experiments (see inset in Figure 2A). The resulting simulated currents are shown in figure 4A. Increasing the rate constants of activation, deactivation and recovery from inactiva- tion by the same factors as obtained from the oocyte experiments gradually shifted the ‘peak potential’ from 248.4 mV (represent- ing hERG1a) to 228.4 mV (representing hERG1b). The overall shape of the currents were very similar to the recordings obtained for hERG1 channels expressed in both mammalian cells and oocytes (compare figure 4A to figures 2A and 3A). The peak amplitude is plotted against the ‘peak potential’ in figure 4B. The changes in peak amplitude followed a pattern similar to the one observed in the oocyte experiments (compare to figure 3B). Action Potential Characteristics Are Modulated by the Relative Abundance of hERG1a and hERG1b To examine the potential functional effects of changes in the relative abundance of hERG1a and hERG1b on the cellular level we substituted the original Hodgkin-Huxley formulation of IKr in the TT model of the human ventricular cardiomyocytes [16] for the Fink Markov model of the hERG1 current. We refer to the modified model as TTF. After adjustment of the maximum conductance of IKr (GKr,max) there were only minor differences in the shape of IKr when comparing the two models (data not shown). Steady-state APD90 (BCL 1000ms) was 303 ms and 302 ms for the TT and the modified TTF models respectively. As before, the hERG1 Markov model was modified to reflect the different ratios of hERG1a and hERG1b based on the factors given in table S2. Steady-state action potentials (BCL 1000 ms) are shown in figure 5A. As the relative abundance of hERG1b was increased the APD90 was gradually shortened from 302 ms (hERG1a) to 275 ms (hERG1b). For 20, 40, 60 and 80% hERG1b the APD90 was 292, 285, 285 and 279 ms respectively. In figure 5B the IKr corresponding to the action potentials in figure 5A are shown. The IKr peaked earlier and the peak amplitude became greater as the relative abundance of hERG1b was increased. Figures 5C and D show the proportion of channels in the open and inactivated states respectively during the action potential. The increase in recovery rate allowed the channels to escape the inactivated state earlier during the action potential (figure 5D). Although the maximum proportion of channels in the open state was decreased as the relative abundance of hERG1b was increased (figure 5C) the driving force on these channels were much greater Role of Cardiac hERG1 Isoforms Figure 4. The experimental data is reproduced by a hERG1 Markov model. A, The effect of changes in transition rates, representing a gradual increase in the relative abundance of hERG1b, on simulated hERG1 currents. The transition rates of hERG1 channel activation (a3), deactivation (b3) and recovery from inactivation (b4) were modified to represent a gradual increase in the relative abundance of hERG1b (see table S2 for details). The model was subjected to the same voltage protocol as in figure 2A. Only currents recorded during the ramp are shown. For comparison, currents were normalized to the maximum peak current during the ramp and plotted against the command voltage. The dotted lines indicate the potential where the currents peaked (‘peak potential’) and are also used to indicate the isoform ratio (in % of hERG1b abundance). B, The variation in ‘peak potential’ amplitude is shown as a function of ‘peak potential’. The data were not normalized as the number of channels in the model remained constant. doi:10.1371/journal.pone.0009021.g004 due to the more depolarized membrane potential at this earlier phase resulting in the increased IKr amplitude. The shortening of APD in response to greater abundance of hERG1b is therefore mainly due to the increase in recovery rate. The TT model has been shown to reproduce data on human restitution kinetics [16]. We wanted to know whether the changes in transition rates would also affect the restitution properties. Action potential restitution was measured using a standard S1-S2 protocol. At steady-state pacing (BCL 1000 ms) an additional stimulus (S2) was introduced at successively shorter coupling intervals and the corresponding APD90 was determined. The relationship between APD90 and DI could be described by a PLoS ONE | www.plosone.org 5 February 2010 | Volume 5 | Issue 2 | e9021