Human genetic and pharmacological studies have demonstrated that voltage-gated sodium channels (VGSCs) are promising therapeutic targets for the treatment of pain.Spider venom contains many toxins that modulate the activity of VGSCs.To date,only 0.01％ of such spider toxins has been explored,and thus there is a great potential for discovery of novel VGSC modulators as useful pharmacological tools or potential therapeutics.In the current study,we identified a novel peptide,μ-TRTX-Ca1a (Ca1a),in the venom of the tarantula Cyriopagopus albostriatus.This peptide consisted of 38 residues,including 6 cysteines,i.e.IFECSISCEIEKEGNGKKCKPKKCKGGWKCKFNICVKV.In HEK293T or ND7/23 cells expressing mammalian VGSCs,this peptide exhibited the strongest inhibitory activity on Nav1.7 (IC50 378 nM),followed by Nav1.6 (IC50 547 nM),Nav1.2 (IC50 728 nM),Nav1.3 (IC50 2.2 μM) and Nav1.4 (IC50 3.2 μM),and produced negligible inhibitory effect on Nav1.5,Nav1.8,and Nav1.9,even at high concentrations of up to 10 uM.Furthermore,this peptide did not significantly affect the activation and inactivation of Nav1.7.Using site-directed mutagenesis of Nav1.7 and Nav1.4,we revealed that its binding site was localized to the DIIS3-S4 linker region involving the D816 and E818 residues.In three different mouse models of pain,pretreatment with Cala (100,200,500 μg/kg) dose-dependently suppressed the nociceptive responses induced by formalin,acetic acid or heat.These results suggest that Ca1a is a novel neurotoxin against VGSCs and has a potential to be developed as a novel analgesic.