The aim of this study was to identify neurochemical pathways and candidate genes involved in adaptation to nicotine treatment and withdrawal. Locomotor sensitization was assessed in a nicotine challenge test after exposure to intermittent nicotine treatment and withdrawal. About 24 h after the challenge test the ventral tegmentum of the mesencephalon was dissected and processed using oligonucleotide microarrays with 22,690 probe sets (Affymetrix 430A 2.0). Quasi-congenic RQI, and donor BALB/cJ mice developed significant locomotor sensitization, while sensitization was not significant in the background partner, C57BL/6By. Comparing saline treated controls of C57BL/6ByJ and BALB/cJ by a rigorous statistical microarray analysis method we identified 238 differentially expressed transcripts. Quasi-congenic strains B6.Cb(4)i(5)-alpha 4/Vad and B6.Ib(5)i(7)-beta 25A/Vad significantly differed from the background strain in 11 and 11 transcripts, respectively. Identification of several cis- and trans-regulated genes indicates that further work with quasi-congenic strains can quickly lead to mapping of Quantitative Trait Loci for nicotine susceptibility because donor chromosome regions have been mapped in quasi-congenic strains. Nicotine treatment significantly altered the abundance of 41, 29, 54, and 14 ventral tegmental transcripts in strains C57BL/6ByJ, BALB/cJ, B6.Cb(4)i(5)-alpha 4/Vad, and B6.Ib(5)i(7)-beta 25A/Vad, respectively. Although transcript sets overlapped to some extent, each strain showed a distinct profile of nicotine sensitive genes, indicating genetic effects on nicotine-induced gene expression. Nicotine-responsive genes were related to processes including regulation of signal transduction, intracellular protein transport, proteasomal ubiquitin-dependent protein catabolism, and neuropeptide signaling pathway. Our results suggest that while there are common regulatory mechanisms across inbred strains, even relatively small differences in genetic constitution can significantly affect transcriptome response to nicotine.