Potentiation by hydrogen peroxide (H2O2) of persistent sodium current in rat peripheral sensory axons has characteristics of a TRPA1-mediated response but is not secondary to Ca2+ entry.

A tetrodotoxin (TTX)-sensitive, persistent sodium current (PSC) can be detected as a persistent extracellular hyperpolarizing DC potential that is provoked when TTX is applied to axons in isolated nerves (1). PSCs revealed in this way in isolated human sural and rat saphenous nerves are potentiated by H2O2 (2). We have studied the mechanism of this phenomenon in rat axons using a grease-gap recording method that has been described previously (3). We measured PSCs in isolated, desheathed rat (Wistar, either sex, 200 - 400 g) saphenous and sciatic nerves superfused with a HCO3- -buffered physiological solution at 20 - 22 oC, gassed with 95% O2 / 5% CO2. Responses to TTX (1 µM) or vehicle (superfusate) control were measured as the integrated DC potential change recorded during a 3 min period (from 1 min after drug or vehicle application) in control nerves or after conditioning by a previous application of either H2O2 (5 mM, 3 min), lidocaine (3 mM, 15 min), mustard oil (MO, 0.2 µM, 9 min), or capsaicin (0.1 - 0.2 µM, 3 min). Conditioning stimuli were followed by at least 15 min washing before TTX or vehicle was applied. Data were compared using 2-way ANOVA followed by a Tukey test.

Application of vehicle did not cause a response in saphenous nerve (-1.02 ± 0.54 mV.s, n = 31), but, in agreement with previously reported data (2), TTX provoked a significant hyperpolarizing response (-7.56 ± 1.57 mV.s, n = 8; P < 0.05) in the nerve. The response to TTX was increased following a conditioning application of H2O2 (-17.48± 3.16mV.s, n = 8; P < 0.01). In sciatic nerve, application of vehicle did not provoke a response (0.26± 0.67mV.s, n = 6) but TTX provoked a significant hyperpolarizing response (-9.17 ± 2.70 mV.s, n = 12; P < 0.05) that was similar to that observed in isolated saphenous nerve. In sciatic nerve the response to TTX was not increased significantly following a conditioning application of H2O2 (-12.65 ± 4.32 mV.s, n = 6; P > 0.05). Since the saphenous nerve is almost exclusively sensory whilst the sciatic nerve is predominantly motor in character these data suggest that the potentiating effect of H2O2 is selective for sensory axons.

In saphenous nerve, a conditioning application of lidocaine or MO increased the TTX response (-17.58 ± 3.41 mV.s after lidocaine, n = 7; P < 0.05 and -18.18 ± 2.18 mV.s after MO, n = 10; P < 0.01) but capsaicin did not (-5.00 ± 1.60 mV.s after capsaicin, n = 6; P > 0.05). Since H2O2, lidocaine and MO are all TRPA1 agonists but capsaicin is not, these data are consistent with activation of TRPA1 as a mechanism of potentiation of the PSC.

In saphenous nerve, when the extracellular Ca2+ concentration was reduced to 50 µM during the conditioning application of H2O2 and then Ca2+ was returned to normal (1.5 mM) before TTX was applied, the potentiation of the PSC was unaffected (-14.78 ± 1.91 mV.s, n = 9; P > 0.05). It is unlikely therefore, that potentiation of the PSC in sensory axons is dependent on Ca2+ entry into the axons via TRPA1 ion channels.

References:

(1) Stys PK et al. (1993). Proc Natl Acad Sci USA 90: 6976-6980.

(2) Docherty RJ et al (2013). http://www.pA2online.org/abstracts/Vol11Issue3abst210P.pdf

(3) Docherty RJ et al (2013). Pain 154: 1569-1577.