Neuronal nicotinic acetylcholine receptors (nAChRs) are pentameric ligand gated ion channels composed of either heteromeric (non-α7) or homomeric (α7) subunits. To date, 12 neuronal subunit genes have been identified and the multiple combinations of subunits create a diverse family of nAChRs. α7 nAChRs have high relative permeability to Ca²⁺ and are implicated in disease states including schizophrenia and Alzheimer’s disease (Cassels et al., 2005). Subtype-selective tools are critical for dissecting the roles of particular nAChR subtypes. The α-conotoxins are neurotoxic peptides first identified in the venom of poisonous marine cone snails. They are small disulphide-rich peptides that target nAChRs to antagonise agonist activation. Subtype-selective α-conotoxins have proved to be valuable tools for investigating the structure, function and the diversity of nAChRs (McIntosh et al., 1999). In this study, we examined the selectivity of a novel α-conotoxin, an analogue of a clone obtained from Conus arenatus, for its activity at native α7 nAChRs compared with non-α7 nAChRs. The α-conotoxin ArIB competed for [¹²⁵I]-α-bungarotoxin binding to rat brain P2 membranes with IC₅₀ = 50nM. This indicates a competitive interaction with α7 nAChRs. Functional antagonism by ArIB was investigated with respect to agonist-evoked increases in intracellular Ca²⁺ in PC12 cells. Cells were loaded with Fluo-3 and changes in the intracellular Ca²⁺ levels were monitored as described previously (Dickinson et al., 2007). The α7 nAChR-selective agonist, choline (1-10mM), elicited small increases in fluorescence consistent with an increase in intracellular Ca²⁺. This response was amplified by preincubation (2min) with PNU 120596 (10μM; an α7 nAChR-selective positive allosteric modulator; Hurst et al., 2005) and could be blocked with α-bungarotoxin (100nM, 20min), confirming that it is mediated by α7 nAChRs. The response to choline and PNU 120596 was concentration-dependently blocked by ArIB, with complete blockade achieved with 300nM ArIB. The specificity of ArIB for α7 nAChRs was supported by its failure to inhibit Ca²⁺ increases mediated by non-α7 nAChRs (induced by nicotine or 5-iodo-Α-85380) or KCl. Thus ArIB is a potentially useful selective antagonist for α7 nAChRs.