Pharmacological modification of sodium channel Nav1.9 activity

Steup, Stefanie Nicole; Spehr, Marc (Thesis advisor); Lampert, Angelika (Thesis advisor)

Aachen : RWTH Aachen University (2022, 2023)
Dissertation / PhD Thesis

Dissertation, RWTH Aachen University, 2022

Abstract

Voltage-gated sodium channels (Navs) generate the sodium current that is responsible for the rapid upstroke of the action potential. Therefore, they are a very important component of the nervous system. The voltage-gated sodium channel 1.9 (Nav1.9) plays an important role in chronic pain. Gain-of-function (GOF) mutations of Nav1.9 are linked to both chronic pain disorders and congenital insensitivity to pain (CIP). By contrast, loss of Nav1.9 does not cause an overt phenotype in rodent models. However, investigation of Nav1.9 function is difficult: Nav1.9 is hard to express heterologously, i.e. in cell lines that do not express Nav1.9 under physiological conditions, and there are no known, selective Nav1.9 modulators. Recently, the development of a Nav1.9/Nav1.4 chimera (Nav1.9_C4) led to progress on the first issue by increasing Nav1.9 expression. The chimera is a cross of the human Nav1.9 containing the C-terminus of rat Nav1.4. In this thesis, I address the second issue, the lack of Nav1.9 modulators - more specifically Nav1.9 activators. I examined the effect of three known Nav activators on Nav1.9_C4 heterologously expressed in ND7/23 cells using whole-cell patch-clamp electrophysiology: the β-scorpion toxins Ts1 and Tf2 and the insecticide deltamethrin, a type II pyrethroid. β-scorpion toxins activate Navs by hyperpolarizing the voltage-dependence of Nav activation, although some also have a depressant effect of reducing current density. Deltamethrin prolongs the opening of Navs by slowing down fast inactivation and deactivation kinetics. Pyrethroids are supposedly safe for humans, however, they have also been linked to the gulf-war syndrome, a neuropathic pain condition that can develop following exposure to certain chemicals. Because deltamethrin is a general Nav activator, and Nav1.7, Nav1.8, and Nav1.9 have all been linked to chronic pain conditions, I additionally examined the effects of deltamethrin on Nav1.7 and Nav1.8, in comparison to Nav1.9_C4. I found that all three activators act on Nav1.9_C4. Both Ts1 and Tf2 cause a small hyperpolarizing shift in activation, but fast inactivation and deactivation remain unchanged. Ts1 additionally reduces current density. Deltamethrin application lead to increased persistent and tail currents in Nav1.7, Nav1.8 and Nav1.9_C4. Surprisingly, deltamethrin induced an enhanced slow inactivation in all three Nav subtypes. An enhanced slow inactivation is contrary to the prolonged opening caused by pyrethroids and has not been described for deltamethrin or any other pyrethroid before. In summary, all three activators act on Nav1.9_C4, but also on other Nav subtypes with similar sensitivities. My findings support a novel mechanism whereby deltamethrin enhances slow inactivation of voltage-gated sodium channels, which may, depending on the cellular resting membrane potential (RMP), reduce neuronal excitability and counteract the well-described pyrethroid effects on channel activation.