A high-capacity membrane potential FRET-based assay for NaV1.8 channels

Area of Science:

• Neuroscience
• Pharmacology
• Molecular Biology

Background:

• Current neuropathic pain treatments targeting voltage-gated sodium channels (NaV1) can cause dose-limiting side effects due to interactions with various NaV1 subtypes.
• The NaV1.8 subtype, restricted to sensory neurons, is implicated in pain processing and represents a potential target for selective therapies.
• Selective NaV1.8 blockers are predicted to be effective for neuropathic pain with fewer adverse effects than existing treatments.

Purpose of the Study:

• To establish a functional assay for identifying selective inhibitors of the human NaV1.8 channel.
• To facilitate the discovery of novel therapeutic agents for neuropathic pain with improved safety profiles.

Main Methods:

• Stable expression of human NaV1.8 and NaV beta1 subunits in HEK293 cells.
• Development of a membrane potential, fluorescence resonance energy transfer-based assay using a FLIPR-Tetra platform.
• Validation of the assay's sensitivity to known NaV1 modulators.

Main Results:

• Successfully established a high-capacity, functional assay for human NaV1.8/NaV beta1 channels.
• The assay demonstrated biophysical properties similar to native tetrodotoxin-resistant channels.
• The assay is sensitive to known NaV1 modulators, indicating its utility for inhibitor screening.

Conclusions:

• A robust assay system for screening NaV1.8 inhibitors has been developed.
• This platform can accelerate the identification of novel, selective NaV1.8 blockers.
• The developed assay holds potential for discovering improved treatments for neuropathic pain.