Fig. 4.

Dynamic action potential-clamp experiment implementing WT, R853Q, L1563V, or R1882Q I_Na. (A, i) Schematic representation of the dynamic clamp technique used to effectively replace the in silico I_Na of the virtual AIS compartment model with I_Na expressed in a mammalian (CHO) cell. (ii) In dynamic clamp (DC) mode, I_Na is recorded from a CHO cell, digitized (A/D), scaled (Fs), and continuously applied to the virtual (model) cell as an external current input. The model cell is in current clamp (CC) mode and its V_m is computed in real-time by the PC-controlled ADwin system. The computed V_m is converted into an analog signal (D/A), sent back to the amplifier, and applied as a voltage clamp (VC) command to the CHO cell. Action potential firing in the model cell is triggered either by step stimulus currents (I_st) or synaptic current (g_e:g_i). The set-up enables switching between dynamic clamp and conventional voltage clamp modes. (B) Dynamic action potential-clamp experiments reveal Na_v1.2 channel gain-of-function or loss-of-function. Action potential firing with model cell incorporating wild-type, R853Q, L1563V, or R1882Q I_Na in response to increasing I_st, in the range between 0 and 16 pA, in 2-pA increments. Representative V_m changes elicited by 4- or 10-pA step currents are shown. (C) Input−output showing the I_st dependence of action potential firing. Data are mean ± SEM; n, number of experiments between parentheses; Note the altered action potential firing of the model cell in the presence of mutant I_Na compared with wild-type (*P < 0.05). (D) Rheobase (*P < 0.05, compared with wild-type); n, same as in C.