Chronic nicotine up-regulates α6-GFP* nAChRs in four brain regions. (A) Direct fluorescence imaging of SNc and VTA in mice expressing α6-GFP subunits after treatment with saline or nicotine (2 mg/kg/h, 10 d). (D₁ and E₁) Immunohistochemical imaging of medial habenula and superior colliculus in mice expressing α6-GFP subunits after treatment with saline or nicotine. For all images, montages were compiled from individual 60× images. (A and E₁) Bars, 100 µm. (D₁) Bars, 20 µm. (B₁ and C₁) Quantification of α6-GFP intensities in SNc and VTA of saline- and nicotine-treated mice (n = 6 mice). (D₂ and E₂) Quantification of α6-GFP intensities in medial habenula and superior colliculus of saline- and nictotine-treated mice (n = 6 mice). (D₃ and E₃) Quantification of α6-GFP immunofluorescence of medial habenula and superior colliculus neurons in saline- and nicotine-treated mice (n = 4–5 mice). (B₂, C₂, D₃, and E₃) Cumulative percentage plots show changes in α6-GFP fluorescence in all brain regions with chronic nicotine. ***, P < 0.005; *, P < 0.05.