Varenicline, a partial agonist for α4β2* nicotinic acetylcholine receptors (nAChRs) and a full agonist for α3β4 and α7 nAChRs, is approved for smoking cessation treatment. Although, partial agonism at α4β2* nAChRs is believed to be the mechanism underlying the effects of varenicline on nicotine reward, the contribution of other nicotinic subtypes to varenicline's effects on nicotine reward is currently unknown. Therefore, we examined the role of α5 and α7 nAChR subunits in the effects of varenicline on nicotine reward using the conditioned place preference (CPP) test in mice. Moreover, the effects of varenicline on nicotine withdrawal-induced hyperalgesia and aversion are unknown. We also examined the reversal of nicotine withdrawal in mouse models of dependence by varenicline. Varenicline dose-dependently blocked the development and expression of nicotine reward in the CPP test. The blockade of nicotine reward by varenicline (0.1 mg/kg) was preserved in α7 knockout mice but reduced in α5 knockout mice. Administration of varenicline at high dose of 2.5 mg/kg resulted in a place aversion that was dependent on α5 nAChRs but not β2 nAChRs. Furthermore, varenicline (0.1 and 0.5 mg/kg) reversed nicotine withdrawal signs such as hyperalgesia and somatic signs and withdrawal-induced aversion in a dose-related manner. Our results indicate that the α5 nAChR subunit plays a role in the effects of varenicline on nicotine reward in mice. Moreover, the mediation of α5 nAChRs, but not β2 nAChRs are probably needed for aversive properties of varenicline at high dose. Varenicline was also shown to reduce several nicotine withdrawal signs.