Figure 3: Double immunofluorescence staining demonstrating the location of nicotine (NIC), and nicotine pyrrolidine methiodide (NIC-PM) induced c-Fos immunoreactive (IR) neurons with respect to tyrosine hydroxylase (TH) IR cells of ventral tegmental area (VTA). Panels A–C: Control data showing physiological saline (PS)-induced c-Fos IR cells (A), tyrosine hydroxylase TH-IR cells (B) and merge images of c-Fos with TH IR cells (C) in VTA. Panels D-F and G-I: Low and high power magnification showing NICinduced c-Fos IR cells (D, G), TH IR cells (E, H) and merge images of c-Fos with TH IR cells (F, I) in VTA. Panels J-L and M-O: Low and high power magnification showing NIC-PM induced c-Fos IR cells (J, M), TH IR cells (K, N) and merge images of c-Fos with TH IR (L, O) cells in VTA. Squares indicate areas that are magnified in photomicrographs. Abbreviations: MM=medial mammillary nucleus, RM=retromamillary nucleus, VTAR=ventral tegmental area rostral.