A thermosensitive mutation alters the effects of lacosamide on slow inactivation in neuronal voltage-gated sodium channels, NaV1.2

Abstract

Epilepsy is a disorder characterized by seizures and convulsions. The basis of epilepsy is an increase in neuronal excitability that, in some cases, may be caused by functional defects in neuronal voltage gated sodium channels (NaVs). The C121W mutation of the β1 subunit, in particular, gives rise to the thermosensitive generalized epilepsy with febrile seizures plus (GEFS+) phenotype. Lacosamide is used to treat epileptic seizures and is distinct from other anti-seizure drugs by targeting NaV slow-inactivation. We studied the effects of a physiologically relevant concentration of lacosamide on the biophysical properties of NaV1.2 channels associated with either WT-β1 or the mutant C121W-β1 subunit. Biophysical parameters were measured at both normal (22°C) and elevated (34°C) temperatures to elicit the differential temperature-sensitivity of C121W. Lacosamide was more effective in NaV1.2 associated with the WT-β1 than with C121W-β1 at either temperature. There is also a more potent effect by lacosamide on slow inactivation at elevated temperatures. Our data suggest a modulatory role is imparted by the β1 subunit in the interaction between the drug and the channel.

Keywords: C121W-β1; GEFS+; WT-β1 (wild-type); lacosamide; slow inactivation; thermosensitive; voltage-gated sodium channels.

Figure 1

Macroscopic currents. Current amplitude plotted verses time duration (ms). These traces were elicited by a set of alternating test pulses that range from −80 mV to +60 mV in increments of +10 mV.

Figure 2

Steady-state fast inactivation. Normalized current is plotted against a range of prepulse potentials (mV). Steady-state fast inactivation was elicited by both 20 ms prepulse durations (filled circles) and 500 ms prepulse durations (filled squares). For both WT at 22°C and 34°C (A,B) and C121W at 22°C and 34°C (C,D). The means of control (shown in black) were compared to 100 μM Lacosamide (shown in red). In panel (D), an inset shows the pulse protocol used to measure steady-state fast inactivation.

Figure 3

Fast inactivation recovery. Normalized Current is plotted against a range of recovery durations (ms). Filled circles shows fast inactivation recovery elicited by 20 ms prepulse durations; whereas closed triangles represent prepulse durations of 500 ms. For both WT at 22°C and 34°C (A,B) and C121W at 22°C and 34°C (C,D). The means of control (shown in black) were compared to 100 μM lacosamide (shown in red). Panel (D) shows an inset of the double pulse protocol measuring fast inactivation recovery.

Figure 4

Slow inactivation onset. Normalized Current is plotted against a range of onset durations (s). We show two different recovery pulse durations: 100 ms (filled squares) and 2 s (filled circles) in control (black) and 100 μM lacosamide (red). For both WT at 22°C and 34°C (A,B) and C121W at 22°C and 34°C (C,D). Inset in (D) shows the pulse protocol.

Figure 5

Steady-state slow inactivation. Normalized Current is plotted against the potential (mV). Control is shown in black circles and drug in red triangles. For both WT at 22°C and 34°C (A,B) and C121W at 22°C and 34°C (C,D). Panel (D) shows an inset of the pulse protocol used to measure slow inactivation. The difference from the protocol used to measure steady-state fast inactivation is that the prepulse duration is longer (50 s) and followed by a brief recovery pulse that recovers channels from fast inactivation prior to the test pulse.

Figure 6

Slow inactivation recovery. Normalized Current is plotted over the various recovery durations (s). Recovery was measured using both 500 ms prepulse durations (closed circles) and also 8 s (closed squares). Control (black) is compared to 100 μM Lacosamide (red). For both WT at 22°C and 34°C (A,B) and C121W at 22°C and 34°C (C,D). Panel (D) shows a pulse protocol inset used to measure slow inactivation recovery.