Fig. 1

tPA is localized to presynaptic terminals associated with DA-neurons. (A) Western blot of tPA, TH, and actin in the SN and hippocampus of WT mice (n = 3–4). Actin was used as a loading control. (B) In-gel zymography showing tPA proteolytic activity in the SN and hippocampus of WT mice. Human tPA was used as a positive control (n = 3). (C) Coronal sections of the SN showing tPA (red) and TH (green) staining in WT mice. White squares indicate 60x closeup (right panels) and white dashed lines depict SNpc (n = 3). (D) Western blot of crude synaptosome protein extracts from SN of WT mice. Synaptosome and cytosol fractions were run and developed in the same gel and membrane (n = 3). (E) In-gel zymography showing tPA proteolytic activity, and (F) quantification of band intensity of supernatants from crude synaptosome preparations from the SN after 5 mM or 50 mM KCl treatment (n = 3–4). tPA-KO mice were used as a negative control in (A), (B), and (D). (G) Confocal images of the SN showing tPA (red), TH (white), and VAMP-2 (green). Merged image shows colocalization of tPA and VAMP-2 (yellow and white arrows). Panels next to merged image (xy) are orthogonal view indicating xz (bottom panel) and yz (right panel) colocalization (n = 3; MOC = 0.4 ± 0.04). Data are shown as mean ± SEM, *p < 0.05; two-tailed t-test. Scale bar = 500 μm (C), 20 μm (close up, C), 10 μm (G)