increased Fos immunoreactivity (Fig. 4a,b). This effect of the high nicotine dose was almost completely abolished in the knockout mice. Wildtype and α5 knockout mice displayed similar Fos immunoreactivity in the ventromedial hypothalamus (Supplementary Fig. 12), a region in which Fos induction is highly stress responsive44, suggesting that altered stress responses in knockout mice did not account for this effect. Nicotine-induced increases in Fos immunoreactivity in the VTA, which controls the reinforcing effects of nicotine, were similar in wildtype and α5 knockout mice (Supplementary Fig. 13). Nevertheless, there was a non-statistically significant trend toward lower VTA Fos immunoreactivity in the knockout mice in response to the high nicotine dose. Considering that the VTA can also regulate aversive responses to nicotine45, it is possible that α5* nAChRs in VTA may differentially regulate activation of this structure in response to aversive but not rewarding doses of nicotine. Taken together, these findings are consistent with our behavioral data in which the reinforcing effects of nicotine, likely involving VTA activation, are substantially conserved in the knockout mice. However, recruitment of an aversive/satiety pathway by nicotine overconsumption, likely involving habenular-driven activation of IPN, is diminished in animals with deficient α5* nAChR signaling.
