Rationale: Cyp2a5, the mouse homologue of human CYP2A6, encodes for the enzyme responsible for the primary metabolism of nicotine. Variation in human CYP2A6 activity can alter the amount smoked such as number of cigarettes smoked per day and smoking intensity. Different mouse strains self-administer different amounts of oral nicotine and quantitative trait loci analyses in mice suggested that Cyp2a5 may be involved in differential nicotine consumption behaviors. Objectives: The goal of this study was to examine whether in vivo nicotine consumption levels were associated with CYP2A5 protein levels and in vitro nicotine metabolism in mice. Method: F2 mice propagated from high (C57Bl/6) and low (St/bJ) nicotine consuming mice were analyzed for CYP2A5 hepatic protein levels and in vitro nicotine metabolizing activity. Results: We found that F2 male high-nicotine (n=8; 25.1 +/- 1.2 mu g nicotine/day) consumers had more CYP2A5 protein, compared to low (n=11; 3.8 +/- 1.4 mu g nicotine/day) consumers (10.2 +/- 1.0 vs 6.5 +/- 1.3 CYP2A5 units). High consumers also metabolized nicotine faster than the low consumers (6 mu M: 0.18 +/- 0.04 vs 0.14 +/- 0.07; 30 mu M: 0.36 +/- 0.06 vs 0.26 +/- 0.13; 60 mu M: 0.49 +/- 0.05 vs 0.32 +/- 0.17 nmol/min/mg). In contrast, female high- (25.1 +/- 2.1 mu g nicotine/day) and low-nicotine (4.7 +/- 1.4 mu g nicotine/day) consumers did not show pronounced differences in nicotine metabolism or CYP2A4/5 protein levels; this is consistent with other studies of sex differences in response to nicotine. Conclusions: These data suggested that among male F2 mice, increased nicotine self-administration is associated with increased rates of nicotine metabolism, most likely, as a result of greater CYP2A5 protein levels.