Exposure to cigarette smoking cues can trigger physiological arousal and desire to smoke. The brain substrates of smoking cue-induced craving (CIC) are beginning to be elucidated; however, it has been difficult to study this state independent of the potential contributions of pharmacological withdrawal from nicotine. Pharmacological withdrawal itself may have substantial effects on brain activation to cues, either by obscuring or enhancing it, and as CIC is not reduced by nicotine replacement strategies, its neuro-anatomical substrates may differ. Thus, characterizing CIC is critical for developing effective interventions. This study used arterial spin-labeled (ASL) perfusion fMRI, and newly developed and highly appetitive, explicit smoking stimuli, to examine neural activity to cigarette CIC in an original experimental design that strongly minimizes contributions from pharmacological withdrawal. Twenty-one smokers (12 females) completed smoking and nonsmoking cue fMRI sessions. Craving self-reports were collected before and after each session. SPM2 software was employed to analyze data. Blood flow (perfusion) in a priori-selected regions was greater during exposure to smoking stimuli compared to nonsmoking stimuli (po0.01; corrected) in ventral striatum, amygdala, orbitofrontal cortex, hippocampus, medial thalamus, and left insula. Perfusion positively correlated with intensity of cigarette CIC in both the dorsolateral prefrontal cortex (r 2 ¼ 0.54) and posterior cingulate (r 2 ¼ 0.53). This pattern of activation that includes the ventral striatum, a critical reward substrate, and the interconnected amygdala, cingulate and OFC, is consistent with decades of animal research on the neural correlates of conditioned drug reward. Keywords: cigarette smoking; craving; neuroimaging; ventral striatum; amygdala; DLPFC INTRODUCTION RationaleBoth nicotine and conditioned cues (reminders) maintain cigarette smoking and lead to relapse (Henningfield and Goldberg, 1983;Rose, 2006). Thus, effective smoking cessation treatments should address both factors. Nicotine replacement therapy (NRT) and bupropion are both effective at ameliorating nicotine withdrawal-induced craving, but neither block the craving elicited by learned associations formed between environmental cues and nicotine (Teneggi and Tiffany, 2002;Robinson and Berridge, 1993;Wise, 1988;Hughes et al, 1999;Hurt et al, 1997;Jorenby et al, 1999). Currently, medications that unequivocally prevent or reduce cue-induced craving (CIC) are not available. Further understanding of the underlying neurobiology of CIC would facilitate the development of effective therapies and improve the currently low (10-20% at 6 months to a year) success rates associated with current smoking interventions West et al, 2001). Thus, a major goal of this work is to examine the element of neurobiology underlying CIC distinct from that observed in CIC studies associated with nicotine withdrawal.Craving generated by drug cues may accrue slowly over time, recruiting supplementary neural substrates as...