Decrease in peak Nav amplitude during 10-pulse trains applied at different frequencies is associated with accumulation of channels in slow recovery pathways. (A) A standard single 5-ms step to 0 mV was used to produce inactivation (pulse 1, P1), with recovery at −80 mV preceding a test step (pulse 2, P2) to 0 mV. Dotted lines highlight amplitude following 10, 100, and 3,000 ms of recovery. (B) From the same cell as in A, a 4-Hz, 10-pulse train of 5-ms steps to 0 mV was applied before the standard recovery step to −80 mV with dotted lines reflecting recovery amplitudes in A. (C) From the same cell as in A and B, a 10-Hz train preceded the recovery steps. (D) The decrement in peak Nav current amplitude is plotted for 1-, 4-, 10-, and 66.7-Hz trains of 10 pulses applied to 0 mV. Mean ± SD. (E) Time course of recovery from inactivation following different stimulus trains. 12 cells were used for 4- and 10-Hz trains, which also included a control set of recovery determinations with the standard single pulse (1P) protocol. 13 different cells were used for the 1-Hz protocol, which were also tested with the 1P protocol. 19 cells (same as in Fig. 7) were used with the 66.7-Hz 10P protocol, which also included the 1P protocol as a control. Fit values for 1P, 4-Hz, and 10-Hz cells were as follows. For 1P: A_f = 0.55 ± 0.02, τ_f = 14.7 ± 0.8 ms, A_s = 0.44 ± 0.02, and τ_s = 354.3 ± 17.2 ms; for 4 Hz, A_f = 0.37 ± 0.02, τ_f = 22.1 ± 2.5 ms, A_s = 0.64 ± 0.02, and τ_s = 453.2 ± 20.7 ms; and for 10 Hz, A_f = 0.28 ± 0.02, τ_f = 25.8 ± 3.1 ms, A_s = 0.69 ± 0.04, and τ_s = 433.7 ± 26.3 ms. For 1P and 1-Hz cells, 1P: A_f = 0.50 ± 0.02, τ_f = 16.1 ± 1.0 ms, A_s = 0.50 ± 0.02, and τ_s = 367.3 ± 13.0 ms; and for 1 Hz, A_f = 0.49 ± 0.02, τ_f = 16.1 ± 1.3 ms, A_s = 0.49 ± 0.02, and τ_s = 403.2 ± 20.3 ms. For 1P and 66.7-Hz cells, 1P: A_f = 0.50 ± 0.02, τ_f = 19.6 ± 1.9 ms, A_s = 0.51 ± 0.02, and τ_s = 404.3 ± 51.6 ms; and for 66.7 Hz, A_f = 0.24 ± 0.03, τ_f = 45.2 ± 8.6 ms, A_s = 0.75 ± 0.03, and τ_s = 493.3 ± 49.9 ms. (F) Means, standard errors, and individual values for the fitted fast component amplitude for all individual cells for the indicated stimulus protocols. Kolmogorov–Smirnov test P values for 12 cells compared with 1P, 4-Hz, and 10-Hz protocols: 1P versus 4 Hz, 0.000; 1P versus 10 Hz, 0.000; 4 Hz versus 10 Hz, 0.066. For 13 cells compared with 1P and 1 Hz protocols: 1P versus 1 Hz, 0.881. For 19 cells, compared with 1P and 66.7-Hz protocols: 1P versus 66.7 Hz, 0.000. (G) Mean ± SD and individual determinations of slow and fast time constants are displayed for the indicated conditions. For cells tested with 1P, 4 Hz, and 10 Hz protocols, Kolmogorov–Smirnov P values, for the fast recovery time constant, were 0.019 (1P versus 4 Hz), 0.001 (1P versus 10 Hz), and 0.433 (4 Hz versus 10 Hz). For the 1P versus 1 Hz train comparison, P = 0.638. For the 1P versus 66.7 Hz train comparison, P = 0.000. For slow time constants, for 1P versus 4 Hz, P = 0.001; for 1P versus 10 Hz, P = 0.186; for 4 Hz versus 10 Hz, P = 0.186. For the 1P versus 1 Hz train comparison, P = 0.341. for the 1P versus 66.7 Hz comparison, P = 0.462.