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Objective Melittin is the main peptide in bee venom.Previous studies demonstrated that this peptide could cause persistent spontaneous pain, primary heat and mechanical hyperalgesia, and enhance the excitability of spinal nociceptive neurons.However, the underlying mechanism of melittin-induced cutaneous hypersensitivity is still not clear.Methods In the present study, melittin was applied topically to acutely dissociated rat dorsal root ganglion (DRG) neurons to examine its effects on the excitability of sensory neurons and the underlying cellular mechanisms using whole-cell patch clamp and calcium imaging techniques.Results It was found that melittin induced intracellular calcium ([Ca2+]i) increases in 60% of small (<25 μm) and medium (<40 μm) diameter sensory neurons.Current clamp recording showed that topical application of melittin evoked long-lasting firing in 55% (56/102) of small and medium neurons tested.The sodium channel blocker tetrodotoxin (TTX) failed to eliminate firing evoked by melittin.Voltage clamp experiments showed that melittin evoked inward currents in sensory neurons in a concentration-dependent manner.Repeated application of melittin caused increased amplitude of the inward currents.Most of the melittin-sensitive neurons were capsaicin-sensitive, and 65% were isolectin B4 (IB4) positive.Further results showed that capsazepine, the TRPV1 receptor inhibitor, completely abolished the melittin-induced inward currents and intracellular calcium transients.The signaling pathways involved in the melittin-induced inward current were also examined.PLA2, but not PLC, was involved in producing the melittin-induced inward currents.Inhibitors of cyclooxygenase (COX) and lipoxygenase (LOX), two key components of the arachidonic acid metabolism pathway, each partially suppressed the inward current evoked by melittin.Inhibitors of protein kinase A (PKA), but not of PKC, also abolished the melittin-induced inward currents.Conclusion These results indicate that melittin can directly excite small and medium sensory neurons at least in part by activating TRPV1 receptors.