Many venomous organisms produce toxins that disrupt neuromuscular communication to paralyze their prey. One common class of such toxins comprises nicotinic acetylcholine receptor antagonists (nAChRs). Thus, most toxins that act on nAChRs are targeted to the neuromuscular subtype. The toxin characterized in this report, alpha-conotoxin GIC, is a most striking exception. The 16-amino acid peptide was identified from a genomic DNA clone from Conus geographus. The predicted mature toxin was synthesized, and synthetic toxin was used in all studies described. alpha-Conotoxin GIC shows no paralytic activity in fish or mice. Furthermore, even at concentrations up to 100 microm, the peptide has no detectable effect on the human muscle nicotinic receptor subtype heterologously expressed in Xenopus oocytes. In contrast, the toxin has high affinity (IC(50) approximately 1.1 nm) for the human alpha3beta2 subunit combination, making it the most neuronally selective nicotinic antagonist characterized thus far. Although alpha-conotoxin GIC shares some sequence similarity with alpha-conotoxin MII, which is also a potent alpha3beta2 nicotinic antagonist, it is much less hydrophobic, and the kinetics of channel block are substantially different. It is noteworthy that the nicotinic ligands in C. geographus venom fit an emerging pattern in venomous predators, with one nicotinic antagonist targeted to the muscle subtype (thereby causing paralysis) and a second nicotinic antagonist targeted to the alpha3beta2 nAChR subtype (possibly inhibiting the fight-or-flight response).