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Figure 3


Fig. 3. The recombinant pepE is expressed in Purkinje cells and is targeted to nerve terminals. (A) Tg-2 PC double immunostained for calbindin (a) and pepE (b). PepE immunoreactivity is absent from the cell body (arrow) and concentrated in presynaptic boutons. Bar, 100 µm. (B) Primary cerebellar (CB) and neocortical (CTX) cultures from Tg-2 mice were solubilized at 8-10 DIV and subjected to Tris-tricine SDS-PAGE. Expression of synapsin I (10.22 antibody; Syn I), actin, calbindin, VAMP2 and recombinant pepE (G304 antibody) is shown. The first lane on the left contained synthetic pepE (50 ng). (C) Primary wild-type hippocampal neurons were transfected to overexpress either GFP-synapsin Ia (a), GFP-domain E (b) or GFP alone (c). Bar, 200 µm. (D) Expression and targeting of GFP-synapsin Ia (a) or GFP-domain E (b) were followed by counterstaining neurons with VAMP2 antibodies (c,d). Both synapsin Ia and domain E GFP chimeras are transported along the axon and localized in en-passant varicosities and VAMP2-positive axon terminals (see arrows in e,f). Bar, 100 µm.





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