Targeted Deletion of the Mouse α2 Nicotinic Acetylcholine Receptor Subunit Gene (Chrna2) Potentiates Nicotine-Modulated Behaviors

Chrna2 mutant mice exhibit reduced motor coordination on the accelerating rotarod. Mice were observed for their latency to fall during a 3 min test period on each of five consecutive days. Using a Dunnett's test, Chrna2^−/− and Chrna2^+/− mice demonstrated significant reductions versus Chrna2^+/+ mice in the latency to fall over the 5 d of testing. *p < 0.05 versus Chrna2^−/− mice. ⁺⁺p < 0.01 versus Chrna2^+/− mice (n = 10–17/group). ⁺p < 0.05 versus Chrna2^+/− mice (n = 10–17/group).

Chrna2^−/− mice exhibit enhanced nicotine-induced analgesia on the hotplate assay. Mice were assessed for nicotine (1.25 mg/kg, i.p.) induced analgesia 5 min after injection. Relative to Chrna2^+/+ mice, post hoc t test analysis revealed a significant enhancement of nicotine-induced analgesia on the hotplate assay in Chrna2^−/− mice: **p < 0.01. A trend (p < 0.06) for reduced baseline values (before nicotine injection) was observed in Chrna2^−/− versus wild-type control mice (data not shown); n = 10–11/group.

An enhancement of nicotine-facilitated cued fear conditioning is seen in female Chrna2^−/− mice. Female (A, B) and male (C, D) mice were trained on both cued and contextual fear conditioning after pretraining and testing administrations of saline or nicotine (0.09 mg/kg, i.p.). Post hoc t test comparisons demonstrate a significant enhancement of nicotine-facilitated cued fear conditioning specifically in female mice (A). **p < 0.01, Bonferroni corrected for multiple comparisons (n = 5–12/group).

Enhanced nicotine self-administration by Chrna2^−/− mice is observed during the first 2 d of acquisition and at the highest dose of nicotine examined compared with saline responding. A, Male mice were food trained up to a fixed ratio 5 (FR5), time out 20 s (TO20 s) schedule of reinforcement. Data are presented as the mean number of food rewards earned (± SEM). B, Mice were then transitioned to 0.03 mg/infusion/kg of intravenous nicotine (acquisition period at an FR5, TO20 s schedule of reinforcement). Data are presented as the mean number of nicotine infusions earned (± SEM) with the active lever (solid line). Values for the inactive lever represent the mean number of lever presses divided by 5 for comparison purposes with the FR5 schedule of the active lever (dotted line). C, Both Chrna2^+/+ and Chrna2^−/− mice were then given access to a range of nicotine doses, which resulted in an inverted U-shaped dose–response curve. Data for the active lever are presented as the mean number of nicotine infusions earned (± SEM, solid line), and for the inactive lever as mean number of lever presses/5 (± SEM, dotted line). D, Post hoc comparisons demonstrate significantly enhanced nicotine self-administration during the first 2 d of nicotine self-administration. **p < 0.01, Bonferroni corrected for multiple comparisons. ***p < 0.001, Bonferroni corrected for multiple comparisons. E, Matched pair t test comparisons between saline- and nicotine-reinforced responses demonstrated that Chrna2^−/− mice had significantly enhanced responses at the highest dose of nicotine tested (0.4 mg/kg), which was not observed in Chrna2^+/+ mice. **p < 0.001, Bonferroni corrected for multiple comparisons. F, At the highest dose of nicotine tested, Chrna2^−/− mice demonstrated significant preference for the reinforced lever versus the nonreinforced lever, which was not observed in Chrna2^+/+ mice. *p < 0.05, Bonferroni corrected for multiple comparisons (n = 4–10/group).

When assayed in a novel environment, a significant increase in mecamylamine-precipitated withdrawal scores was observed in nicotine-treated Chrna2^+/+ and Chrna2^−/− mice. Mice were administered nicotine (24 mg/kg per day) using an Alzet osmotic minipump. On the 13th day of nicotine exposure, mice were placed into a novel environment, habituated for 40 min, administered mecamylamine (3 mg/kg, i.p.), and evaluated for somatic symptoms of withdrawal for an additional 20 min. A, Quantifying paw tremors, head shakes, backing, and curls, t test post hoc analysis revealed a significant enhancement of behavioral signs during nicotine withdrawal. **p < 0.01 (Bonferroni corrected for multiple comparisons). *p < 0.05 (Bonferroni corrected for multiple comparisons). Using a Dunnett's test (post hoc comparisons), Chrna2^−/− versus Chrna2^+/+ wild-type mice demonstrated significant enhancement of withdrawal signs after nicotine, but not saline, treatment. *p < 0.0575. B, Quantifying grooming, scratching, chewing, cage scratching, head nodding, and jumping, t test post hoc analysis revealed a significant reduction of behavioral signs during nicotine withdrawal. Using a Dunnett's test (post hoc comparisons), Chrna2^−/− versus wild-type mice demonstrated significant enhancement of withdrawal signs in nicotine-, but not saline-treated, mice. *p < 0.05, Bonferroni corrected for multiple comparisons (n = 11–20/group).