Chemical and Biological Tools for the Study of Voltage-Gated Sodium Channels in Electrogenesis and Nociception

The malfunction and misregulation of voltage-gated sodium channels (Na_Vs) underlie in large part the electrical hyperexcitability characteristic of chronic inflammatory and neuropathic pain. Na_Vs are responsible for the initiation and propagation of electrical impulses (action potentials) in cells. Tissue and nerve injury alter the expression and localization of multiple Na_V isoforms, including Na_V1.1, 1.3, and 1.6–1.9, resulting in aberrant action potential firing patterns. To better understand the role of Na_V regulation, localization, and trafficking in electrogenesis and pain pathogenesis, a number of chemical and biological reagents for interrogating Na_V function have been advanced. The development and application of such tools for understanding Na_V physiology are the focus of this review.     

Trifunctional Saxitoxin Conjugates for Covalent Labeling of Voltage-Gated Sodium Channels**

Voltage-gated sodium ion channels (Na_Vs) are integral membrane protein complexes responsible for electrical signal conduction in excitable cells. Methods that enable selective labeling of Na_Vs hold potential value for understanding how channel regulation and post-translational modification are influenced during development and in response to diseases and disorders of the nervous system. We have developed chemical reagents patterned after (+)-saxitoxin (STX) – a potent and reversible inhibitor of multiple Na_V isoforms – and affixed with a reactive electrophile and either a biotin cofactor, fluorophore, or ‘click’ functional group for labeling wild-type channels. Our studies reveal enigmatic structural effects of the probes on the potency and efficiency of covalent protein modification. Among the compounds analyzed, a STX-maleimide-coumarin derivative is most effective at irreversibly blocking Na⁺ conductance when applied to recombinant Na_Vs and endogenous channels expressed in hippocampal neurons. Mechanistic analysis supports the conclusion that high-affinity toxin binding is a prerequisite for covalent protein modification. Results from these studies are guiding the development of next-generation tool compounds for selective modification of Na_Vs expressed in the plasma membranes of cells.