Voltage-gated Na+ (NaV) channels are significant therapeutic targets for the treatment of cardiac and neurological disorders, thus promoting the search for novel NaV channel ligands. With the objective of discovering new blockers of NaV channel ligands, we screened an In-House vegetal alkaloid library using fluorescence cell-based assays. We screened 62 isoquinoline alkaloids (IA) for their ability to decrease the FRET signal of voltage sensor probes (VSP), which were induced by the activation of NaV channels with batrachotoxin (BTX) in GH3b6 cells. This led to the selection of five IA: liriodenine, oxostephanine, thalmiculine, protopine, and bebeerine, inhibiting the BTX-induced VSP signal with micromolar IC50. These five alkaloids were then assayed using the Na+ fluorescent probe ANG-2 and the patch-clamp technique. Only oxostephanine and liriodenine were able to inhibit the BTX-induced ANG-2 signal in HEK293-hNaV1.3 cells. Indeed, liriodenine and oxostephanine decreased the effects of BTX on Na+ currents elicited by the hNaV1.3 channel, suggesting that conformation change induced by BTX binding could induce a bias in fluorescent assays. However, among the five IA selected in the VSP assay, only bebeerine exhibited strong inhibitory effects against Na+ currents elicited by the hNav1.2 and hNav1.6 channels, with IC50 values below 10 µM. So far, bebeerine is the first BBIQ to have been reported to block NaV channels, with promising therapeutical applications.

CNRS ICR Aix Marseille Univ 13397 Marseille France

Department of Pathophysiology 1st Faculty of Medicine Charles University 160 00 Prague Czech Republic

Institute of Biology and Medical Genetics 1st Faculty of Medicine Charles University 128 00 Prague Czech Republic

Institute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences 166 10 Prague Czech Republic

Laboratory of Applied Biotechnology AZM Centre for Research in Biotechnology and Its Application Doctoral School for Sciences and Technology Tripoli 1352 Lebanon

Latoxan Laboratory 26800 Portes lès Valence France

Nantes Université CHU Nantes CNRS INSERM L'institut du Thorax 44000 Nantes France

Univ Angers INSERM CNRS MITOVASC Equipe CarME SFR ICAT 49000 Angers France

Univ Angers SONAS SFR QUASAV 49100 Angers France

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