Skip to Content
Merck

Defining the Functional Role of NaV1.7 in Human Nociception.

Neuron (2019-02-24)
Lucy A McDermott, Greg A Weir, Andreas C Themistocleous, Andrew R Segerdahl, Iulia Blesneac, Georgios Baskozos, Alex J Clark, Val Millar, Liam J Peck, Daniel Ebner, Irene Tracey, Jordi Serra, David L Bennett
ABSTRACT

Loss-of-function mutations in NaV1.7 cause congenital insensitivity to pain (CIP); this voltage-gated sodium channel is therefore a key target for analgesic drug development. Utilizing a multi-modal approach, we investigated how NaV1.7 mutations lead to human pain insensitivity. Skin biopsy and microneurography revealed an absence of C-fiber nociceptors in CIP patients, reflected in a reduced cortical response to capsaicin on fMRI. Epitope tagging of endogenous NaV1.7 revealed the channel to be localized at the soma membrane, axon, axon terminals, and the nodes of Ranvier of induced pluripotent stem cell (iPSC) nociceptors. CIP patient-derived iPSC nociceptors exhibited an inability to properly respond to depolarizing stimuli, demonstrating that NaV1.7 is a key regulator of excitability. Using this iPSC nociceptor platform, we found that some NaV1.7 blockers undergoing clinical trials lack specificity. CIP, therefore, arises due to a profound loss of functional nociceptors, which is more pronounced than that reported in rodent models, or likely achievable following acute pharmacological blockade. VIDEO ABSTRACT.

MATERIALS
Product Number
Brand
Product Description

Millipore
Monoclonal Anti-HA−Agarose antibody produced in mouse, clone HA-7, purified immunoglobulin, PBS suspension
Sigma-Aldrich
TGF-β RI Kinase Inhibitor VI, SB431542, TGF-β RI Kinase Inhibitor VI, SB431542, CAS 301836-41-9, is a cell-permeable inhibitor of SMAD2 phosphorylation. Inhibits the activity of ALK4 and ALK5 (IC₅₀ = 140 nM and 94 nM, respectively).
Sigma-Aldrich
Anti-Brn-3a Antibody, Chemicon®, from rabbit
Sigma-Aldrich
Monoclonal Anti-β-Actin antibody produced in mouse, clone AC-74, ascites fluid
Sigma-Aldrich
LDN193189 hydrochloride, ≥98% (HPLC)
Sigma-Aldrich
Anti-Rat IgG (H+L), highly cross-adsorbed, CF405M antibody produced in goat, ~2 mg/mL, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Naloxone hydrochloride dihydrate, ≥98% (TLC and titration), powder
Sigma-Aldrich
Monoclonal Anti-S-100 (β-Subunit) antibody produced in mouse, clone SH-B1, ascites fluid
Sigma-Aldrich
DAPT, ≥98% (HPLC), solid
Sigma-Aldrich
CHIR99021, ≥98% (HPLC)