[3H]-lifarizine, a high affinity probe for inactivated sodium channels

A. C. MacKinnon, K. M. Wyatt, J. G. McGivern, R. D. Sheridan, C. M. Brown

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    18 Citations (Scopus)


    1 [3H]-lifarizine bound saturably and reversibly to an apparently homogeneous class of high affinity sites in rat cerebrocortical membranes (K(d) = 10.7 ± 2.9 nM; B(max) = 5.10 ± 1.43 pmol mg-1 protein). 2 The binding of [3H]-lifarizine was unaffected by sodium channel toxins binding to site 1 (tetrodotoxin), site 3 (a-scorpion venom) or site 5 (brevetoxin), Furthermore, lifarizine at concentrations up to 10 µM had no effect on [3H]-saxitoxin (STX) binding to toxin site 1. Lifarizine displaced [3H]- batrachotoxinin-A 20-a-benzoate (BTX) binding with moderate affinity (pIC50 7.31 ± 0.24) indicating an interaction with toxin site 2. However, lifarizine accelerated the dissociation of [3H]-BTX and decreased both the affinity and density of sites labelled by [3H]-BTX, suggesting an allosteric interaction with toxin site 2. 3 The binding of [3H]-lifarizine was voltage-sensitive, binding to membranes with higher affinity than to synaptosomes (pIC50 for cold lifarizine=7.99 ± 0.09 in membranes and 6.68 ± 0.14 in synaptosomes). Depolarization of synaptosomes with 130 mM KCl increased the affinity of lifarizine almost 10 fold (pIC50=7.86 ± 0.25). This suggests that lifarizine binds selectively to inactivated sodium channels which predominate both in the membrane preparation and in the depolarized synaptosomal preparation. 4 There was negligible [3H]-lifarizine and [3H]-BTX binding to solubilized sodium channels, although [3H]-STX binding was retained under these conditions. 5 The potencies of a series of compounds in displacing [3H]-lifarizine from rat cerebrocortical membranes correlated well with their affinities for inactivated sodium channels estimated from whole-cell voltage clamp studies in the mouse neuroblastoma cell line, N1E-115 (r=0.96). 6 These results show that [3H]-lifarizine is a high affinity ligand for neuronal sodium channels which potently and selectively labels a site, allosterically linked to toxin binding site 2, associated with inactivated sodium channels.

    Original languageEnglish
    Pages (from-to)1103-1109
    Number of pages7
    JournalBritish Journal of Pharmacology
    Issue number6
    Publication statusPublished - 1995


    • BTX
    • Lifarizine
    • Ligand binding
    • N1E-115 cells
    • Patch-clamp
    • Rat cortex
    • RS-87476
    • Sodium channels
    • Sodium currents


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