'Use Dependence' Without a Ball and Chain - Inhibition of Bacterial Sodium Channels by μ-Contoxins

“Use-” or “state-” dependent block of voltage-gated sodium channels is characteristic of therapeutic agents used to moderate electrical activity in pathologically hyperactive tissues and combat certain forms of epilepsy, cardiac arrhythmia, myotonia, and pain. Externally acting Na channel pore blockers are not commonly used as therapeutic drugs, but intraperitoneal μ-conotoxin (μCTX) KIIIA can act as an effective analgesic against inflammatory pain in mice, at concentrations that do not cause obvious systemic side effects (Zhang et al 2007 J Biol Chem 42:30699). This action appears to result from highly specific targeting to neuronal Na channels, without essential reliance on the use-dependent potency that contributes to the efficacy of clinical local anesthetics. μCTXs, although highly selective among closely related eukaryotic Na channels, have recently been found to block bacterial Nav channels with high affinity (Chen & Chung 2012 Biophys J 102:483; Finol-Urdaneta et al 2013 Biophys J 104:136a). In addition, the block was potentiated at more depolarized holding potentials. Here, we explore the dependence of μCTX PIIIA block of NaChBac and NavSp1 on varying patterns of conditioning depolarization. Briefly, in the low picomolar range, PIIIA application was associated with faster inactivation decay during a test depolarization, and block of the peak current. The speeding of inactivation precedes the block of current following PIIIA application, and this kinetic effect was obvious at more negative holding potentials than was the decrease in peak current. In NaChBac, both activation and inactivation were shifted in the hyperpolarizing direction by ∼25-30 mV in the presence of 5pM PIIIA. The data suggest that PIIIA binding is conformation dependent, with high affinity for non-conducting, depolarization-induced states. At present, the relative importance of “pre-activated” and “in-activated” states in potentiating inhibition by PIIIA is unclear.