Figure 4.
NGF inactivates GSK-3β via p75NTR. (A) GSK-3 activity was determined in extracts from cultured hippocampal neurons that were incubated for 1 h in the presence or absence of 100 ng/ml NGF, 100 ng/ml NGF plus anti-p75NTR blocking antibody (1:100 diluted), or 100 ng/ml NGF plus K252a, a Trk inhibitor. Observe that the blockage of p75NTR prevented the NGF-induced inhibition of the kinase, whereas the inhibition of Trk receptors by K252a did not affect the inhibition of GSK-3 by NGF. (B) Western blot analysis of the same samples with anti-phospho-GSK-3α/β (Ser21/9) (top) and anti-GSK-3β (bottom) antibodies. The asterisk indicates the band corresponding to the heavy chain of the anti-p75 antibody. Note that in the presence of the p75-antibody, NGF did not induce Ser9 phosphorylation of the enzyme, whereas inhibition of the Trk receptor by K252a did not change the NGF-induced phosphorylation pattern. (C) Densitometric quantification of Western blots from three different experiments.