Extensive evidence indicates that varenicline reduces nicotine craving and withdrawal symptoms by modulating dopaminergic function at α4β2* nicotinic acetylcholine receptors (nAChRs) (the asterisk indicates the possible presence of other nicotinic subunits in the receptor complex). More recent data suggest that α6β2* nAChRs also regulate dopamine release and mediate nicotine reinforcement. The present experiments were therefore done to test the effect of varenicline on α6β2* nAChRs and their function, because its interaction with this subtype is currently unclear. Receptor competition studies showed that varenicline inhibited α6β2* nAChR binding ( K i = 0.12 nM) as potently as α4β2* nAChR binding ( K i = 0.14 nM) in rat striatal sections and with ∼20-fold greater affinity than nicotine. Functionally, varenicline was more potent in stimulating α6β2* versus α4β2* nAChR-mediated [ 3 H]dopamine release from rat striatal synaptosomes with EC 50 values of 0.007 and 0.086 μM, respectively. However, it acted as a partial agonist on α6β2* and α4β2* nAChR-mediated [ 3 H]dopamine release with maximal efficacies of 49 and 24%, respectively, compared with nicotine. We also evaluated varenicline’s action in striatum of monkeys, a useful animal model for comparison with humans. Varenicline again potently inhibited monkey striatal α6β2* ( K i = 0.13 nM) and α4β2* ( K i = 0.19 nM) nAChRs in competition studies. Functionally, it potently stimulated both α6β2* (EC 50 = 0.014 μM) and α4β2* (EC 50 = 0.029 μM) nAChR-mediated [ 3 H]dopamine release from monkey striatal synaptosomes, again acting as a partial agonist relative to nicotine at both subtypes. These data suggest that the ability of varenicline to interact at α6β2* nAChRs may contribute to its efficacy as a smoking cessation aid.