Smoking cessation strategies are of prime medical importance. Despite availability of various pharmacological agents in combating addiction to nicotine, more effective medications are needed. Based on recent findings, the glutamatergic system in the brain may provide novel targets. Here, we evaluated the effects of acute administration of sub-anesthetic doses of ketamine, an NMDA receptor antagonist, in both male and female Sprague-Dawley rats trained to self-administer nicotine. Animals were injected subcutaneously with 5, 7.5 and 10 mg/kg ketamine or saline and the effects on the number of intravenous nicotine infusions during a 45 min session was measured. Ketamine treatment significantly reduced nicotine self-administration in a dose-dependent manner. Moreover, a differential sensitivity between the sexes was observed as male rats responded to a lower dose of ketamine and with higher magnitude of effect than female rats. It is concluded that glutamatergic receptor manipulations may offer a novel and potentially sex-dependent intervention in nicotine addiction.