Tobacco use is the leading preventable cause of mortality in the world. The limited number of smoking cessation aids currently available are minimally effective, highlighting the need for novel therapeutic interventions. We describe a genome-wide approach to identify potential candidates for such interventions. Next-generation sequencing was performed using RNA isolated from the habenulo-interpeduncular circuit of male mice withdrawn from chronic nicotine treatment. This circuit plays a central role in the nicotine withdrawal response. Differentially expressed miRNAs and mRNAs were validated using RT-qPCR. Many of the differentially expressed mRNAs are predicted targets of reciprocally expressed miRNAs. We illustrate the utility of the dataset by demonstrating that knockdown in the interpeduncular nucleus of a differentially expressed mRNA, that encoding profilin 2, is sufficient to induce anxiety-related behavior. Importantly, profilin 2 knockdown in the ventral tegmental area did not affect anxiety behavior. Our data reveal wide-spread changes in gene expression within the habenulo-interpeduncular circuit during nicotine withdrawal. This dataset should prove to be a valuable resource leading to the identification of substrates for the design of innovative smoking cessation aids.