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Fig. 3. Slice culture model of SVZ-olfactory bulb neurogenesis. (A) DiI labeling of
cells migrating from SVZ to olfactory bulb (OB) in a P10 explant 3 days after
DiI crystal placement in SVZ. The inset shows the bipolar appearance of
migrating cells (arrows) at higher magnification. (B) BrdU labeling of cells
in the rostral migratory stream (RMS) of a P10 slice culture. Arrows in B show
scattered, putative glia that incorporated BrdU. (B') Higher
magnification of the boxed area in B. BrdU was given in vivo 6 hours before
slice preparation and culture for 3 days. (C) Doublecortin-expressing
neuroblasts in the SVZ and RMS of a P10 explant after 3 DIV. (C') Higher
magnification of the boxed area in C. (D) Schematic of the electroporation
technique. Plasmid DNA was placed in the open lateral ventricle of a
hemisphere, electroporated, and parasagittal slices prepared. (E) GFP labeling
of cells in the SVZ (arrows) and RMS (arrowheads) of a P2 slice 3 days after
electroporation of a GFP reporter. Propidium iodide counterstain (PI). (F)
Four days after electroporation of a GFP reporter, labeled cells appear in the
SVZ-olfactory bulb pathway. The inset is a higher magnification view of the
boxed area showing a cell with migratory morphology. LV, lateral ventricle;
Cx, frontal cortex. (G,H) To examine dual plasmid electroporation efficiency,
P2 slices were co-electroporated with GFP and dsRed reporters, cultured for 4
DIV, fixed and resectioned at 50 µm. Nearly 100% co-electroporation
efficiency was found in the SVZ. All explant images in this and subsequent
figures display parasagittal sections with anterior to the right and dorsal at
top. Scale bars: in A, 150 µm for A,B',C, 250 µm for B,F and 500
µm for E; in inset in A, 75 µm for C' and insets in A and F; and
100 µm for G,H.
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