Smoking is the leading cause of preventable death and disease in the U.S., and women disproportionately suffer some health consequences of smoking, such as heart disease and cervical cancer. Compared to men, women experience stronger withdrawal symptoms and have more difficulty maintaining long-term abstinence from smoking, and smoking-cessation treatments are not equally helpful to men and women. The underlying differences in brain function that drive these male-female differences remain unknown, and understanding them can guide development of personalized smoking-cessation therapies. Glutamatergic neurotransmission is a promising target for smoking cessation therapy, but male-female differences that can impact the efficacy of glutamatergic treatments have not been investigated. Reducing glutamate levels within the Salience Network, especially in the dorsal anterior cingulate cortex (dACC) may facilitate smoking cessation. Smokers have higher glutamate (Glu) in the dACC than nonsmokers and exhibit a positive correlation of dACC Glu with reactivity within the default-mode network to smoking-related cues. Given these findings and sex differences in brain glutamate levels in healthy individuals, it is important to test for male-female differences in the influence of dACC Glu on craving and negative states during withdrawal and their relief after smoking, as well as the neural substrates that underlie these states. We will combine self-report measures with magnetic resonance spectroscopy (MRS) and functional magnetic resonance imaging (fMRI) to address three specific aims: Aim 1. Determine male-female differences in the association of Glu signal in the dACC with cigarette craving and negative states during abstinence (negative affect, psychological withdrawal, and anxiety). Aim 2. Determine male-female differences in the association of Glu in the dACC with smoking-induced relief of craving and negative states (i.e., negative reinforcement); and Aim 3. Determine male-female differences in the association of Glu in the dACC with resting-state functional connectivity (rsFC) within and between large-scale neural networks. The aims of this project focus on pre-smoking assessments of Glu in the dACC and rsFC of neural networks; however, pre- and post-smoking assessments (self-report, fMRI, and MRS) will be completed in 60 smokers (30 men and 30 women) after overnight (~12h) abstinence. Women will complete these assessments on 2 separate days, during the follicular and luteal phases, to allow evaluation of menstrual-cycle and ovarian hormone effects. The findings can help advance development of personalized, sex-specific smoking cessation therapies. RELEVANCE (See instructions): Glutamatergic neurotransmission is a promising target for smoking cessation therapy, but male-female differences, which can impact the efficacy of glutamatergic treatments, have not been investigated. We will combine self-report measures with magnetic resonance spectroscopy and ...