Epilepsy-Induced High Affinity Blockade of the Cardiac Sodium Current INa by Lamotrigine; A Potential for Acquired Arrythmias

Figure 1. LTG blockade of INa is increased during epilepsy. (A) Inset: voltage clamp protocol. INa currents were activated by 25 ms voltage steps from −70 to 10 mV, in 5 mV increments, from a −100 mV holding potential. Representative INa recordings from sham and epileptic rat ventricular cardiomyocytes in control conditions (solid line), and in presence of 10 µmol/L of LTG (dash-dot line). (B) Current–voltage relationships (I/V) for sham (filled symbol) and epileptic (open) animals in control conditions (circle), 10 µmol/L (square) and 100 µmol/L of LTG (triangle). (C) Normalized INa activation curves for control and 10 µmol/L of LTG. Data points (±SEM) were fitted to a Boltzmann equation (solid lines). (D) LTG (10 µmol/L) depolarized INa voltage threshold (INa,Th) only in epileptic animals, while 100 µmol/L of LTG significantly depolarized INa,Th in both groups. Each bar represents average values (±SEM) in control conditions and upon addition of 10 and 100 µmol/L of LTG. (E) LTG decreased Na+ channel conductance (GNa,Max) in epileptic animals. Average values of GNa,Max in same conditions as (D). Tukey test using one-way ANOVA (ctrl vs. LTG), * indicates p ˂ 0.05, *** indicates p ˂ 0.001.…

Figure 2. Epilepsy increased the affinity of LTG for INa. (A) Dose response curves for sham (filled symbol) and epileptic (open symbols) animals. The curves were constructed by averaging the ratio between the INa control value (INa,Max) and its amplitude at each LTG concentration tested. (B) IC50 values estimated by fitting single- and dual-dose response equations (solid lines) to sham (filled) and epileptic (open) animal data, respectively. Number of cells: Sham (ctrl, LTG): 7–14. Epileptic (ctrl, LTG): 8–13.

Figure 3. LTG hyperpolarized INa steady state-inactivation. (A) Representative INa inactivation recording obtained during a 15 ms test pulse to −30 mV following a series of 500 ms conditioning potentials from −120 to −40 mV in 5 mV increments (top). (B) Steady-state inactivation curves for sham (filled symbol) and epileptic (open symbol) animals in control conditions (circles) and 10 µmol/L of LTG (squares). Data points (±SEM) were fitted using a Boltzmann function (solid line). Number of animals: 3/group (ctrl and LTG). Number of cells: Sham (ctrl, LTG):15, 11; Epileptic (ctrl, LTG): 12, 19.

Figure 4. LTG slows INa recovery from inactivation. (A) Representative INa recording during recovery from inactivation. INa was elicited by two step pulses, S1 (100 ms) and S2 (20 ms) to −35 mV, separated by increasing intervals at the resting membrane potential, in increments of 5 ms. Holding membrane potential was −100 mV. (B) Recovery from inactivation curves for sham (filled symbol) and epileptic rats (open symbol) in control (circles), and 10 µmol/L of LTG (squares). Data (±SEM) were fitted to two exponential functions (solid: control, short-dot line: LTG) to estimate the time constants. Number of animals: 3/group (ctrl and LTG). Number of cells: Sham (ctrl, LTG):19, 9; Epileptic (ctrl, LTG): 9, 15.

Figure 5. LTG prolonged cardiac action potential refractory period. (A,C) Representative action potential recordings from sham and epileptic rats in control (ctrl) and in presence of 1 µmol/L of LTG. Cardiomyocytes were held at membrane potentials of −80 or −100 mV and stimulated at a pacing cycle length of 2 ms (sham) and 10 ms (epileptic) for control experiments, and 5 ms (sham) and 50 ms (epileptic) to 1 µmol/L of LTG. For simplicity, only representative records of cells held at −80 mV are shown in the figure. However, as indicated above, the recordings were also performed on the same cell at −100 mV. (B,D) Action potential (AP) recovery time was measured as the time needed to recover a full amplitude action potential. Each bar represents the average value (±SEM). Tukey test using one-way ANOVA (ctrl vs. LTG), *** indicates p ˂ 0.001, and § indicates p ˂ 0.001 (sham vs. epileptic). Number of animals: 3/group (ctrl and LTG). Number of cells: Sham (ctrl, LTG):14, 14; Epileptic (ctrl, LTG): 14, 12.

Figure 6. LTG blocked INaL current in cardiomyocytes from epileptic animals but had no effect in sham rats. (A) Representative INaL recordings from sham and epileptic rat cardiomyocytes in control conditions (ctrl, black) and in presence of 10 µmol/L/L of Lamotrigine (LTG, gray). Currents were elicited by a voltage ramp protocol from −120 to 60 mV with a 0.04 mV/ms rate. (B) IV-curves showing the average of current density values for sham (left) and epileptic animals (right) in control conditions (filled circles) and to 10 µmol/L/L of LTG (empty circles). Number of animals: 3/group (ctrl and LTG). Number of cells: Sham (ctrl, LTG):12, 12; Epileptic (ctrl, LTG): 14, 16.