Xenopus laevis oocytes expressing various nAChR subtypes were subjected to TEVC electrophysiology. Oocytes were stimulated with 1 s pulses of ACh in the absence or presence of ascending concentrations of α-Ctx VnIB. (A) α-Ctx VnIB-mediated block of β2-containing rat nAChRs and the α7, α9α10 and human muscle-type α1β1δε nAChRs. VnIB blocks the ACh-evoked responses through α6/α3β2β3 with a low micromolar IC50 (3.2 μM) but has no significant effect on the other receptor subtypes (IC50 > 10 μM). (B) α-Ctx VnIB-mediated block of β4-containing rat nAChRs. VnIB blocks the ACh-evoked currents through α6β4 and α6/α3β4 nAChRs with similar antagonist potencies (IC50 12 nM and 18 nM, respectively) and the currents through the α3β4 receptor with a ~20-fold higher IC50-value (0.32 μM), but the toxin has no significant effect on α2β4 and α4β4 (IC50 > 10 μM). (C) α-Ctx VnIB-mediated block of selected human neuronal receptor subtypes. VnIB potently blocks α6/α3β4 (IC50 = 5.3 nM), but has no or low antagonistic effect at α3β4, α4β2 and α6/α3β2β3. Each data point represents mean values ± SD based on at least 3 replicates. r, rat; h, human.