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Fig. 4. Modulation of granule cell precursor proliferation in the EGL by Shh and
BMPs. Organotypic slice cultures of cerebellum were maintained for 72 hours
and analysed for granule cell precursor proliferation by BrdU incorporation.
(A) Cerebellar slices were obtained (300 µm), cultured on polycarbonate
filters. Beads soaked in either PBS, Shh, Bmp2, Bmp4 or Bmp7 were implanted
close to the EGL. (B) Confocal images of cultured slices that were in toto
immunostained with anti-calbindin antibody (green cells) and anti-BrdU (red
nuclei). White dots indicate the location of the bead. Analysis of each
culture was performed by counting total numbers of BrdU-positive cells in a
fixed area surrounding the beads. (C) Quantification of proliferation in the
EGL of organotypic slice cultures with different beads implanted. Values are
expressed as mean±s.e.m. A significant increase in the total number of
BrdU-positive cells is observed in slices with Shh beads
(**P<0.001 Turkey’s Test) when compared with
PBS-soaked beads. A significant decrease on the total number of BrdU-positive
cells is observed in slices treated with Bmp2
(**P<0.001) and Bmp4 (*P<0.01)
when compared with PBS beads. Although Bmp4 seams less active in these
experiments, differences between Bmp2 and Bmp4 activities have no statistical
significance (Bmp2 versus Bmp4 P<0.05). Bmp7-loaded beads induced
a slight but not statistically significant increase on the total number of
BrdU-positive cells (P<0.05) when compared with PBS beads.
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