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. 2021 Sep 2;9(9):1141. doi: 10.3390/biomedicines9091141

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© 2021 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

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Recovery course from slow-inactivation and affinity of cannabidiol to the slow-inactivated Na_v1.4 channel. (A) Patched cells expressing the Na_v1.4 channel were held at −120 mV for 5 s. An inactivating pulse at −10 mV for 18 s was applied with a subsequent repolarization to −120 mV for a certain time to eliminate the channels with fast inactivation. The second test pulse of 0 mV was applied after an intervening gap at −120 mV for different lengths of time (0.003, 0.006, 0.009, 0.012, 0.015, 0.018, 0.021, 0.024, 0.027, 0.11, 0.12, 0.31, 0.51, 1, 2, 3, 4, and 5 s). Representative currents were overlaid according to the time sequence of examination. Time scales were applied for current kinetics only. The relative current, defined as the normalized peak current in the second test pulse (to the peak current in the first test pulse), was plotted against the length of intervening gap to obtain the time course of recovery from slow-inactivation in control (n = 5) Na_v1.4 channels and those treated with different concentrations of CBD (n = 5). Note the close-up view of the first 500 ms of data shows a significant dose-dependent change in the recovery of slow-inactivation after CBD treatment (inset figure is a closed-up view of Figure 5A; * p < 0.05). (B) Approaches and analysis protocols were the same as in Figure 5A, except that the inactivating pulse was set at −80 mV for 18 s. (C) Difference between the area in control and upon different concentrations of CBD in Figure 5A to the maximal difference giving the relative difference in the area, which is plotted against the CBD concentration (see Supplementary Data Figure S1). The line is the fit of data points using the following formula: relative difference in area = ([CBD]/31.5)/(1 + [CBD]/31.5), where [CBD] denotes CBD concentration in μM [41]. (D) Analysis protocols were the same as those of Figure 5B, except that the inactivating pulse was set at −80 mV for 18 s. The line is the fit using the following formula: relative difference in the area = ([CBD]/127.3)/(1 + [CBD]/127.3), where [CBD] denotes CBD concentration in μM [41].