Nerve growth factor (NGF) binds to two neurotrophin receptors: p75NTR and p140Trk (TrkA). Both receptors dimerize in response to NGF binding. TrkA homodimers and heteromers of TrkA and p75NTR promote cell survival whereas homodimers of p75NTR mediate apoptosis upon binding of NGF. The interaction between receptor and NGF can be inhibited either on the level of the ligand by altering NGF conformation so that NGF is no longer recognized by the receptor or on the level of the receptor by blocking the binding site of p75NTR or TrkA. The effect of altering NGF conformation on NGF signaling was investigated in two neuron‐like cell lines: in human SK‐N‐MC cells that express only p75NTR and in rat PC12 cells that express both p75NTR and TrkA. In the present study we demonstrate that Ro 08‐2750 binds to the NGF dimer thereby probably inducing a change in its conformation such that NGF cannot bind to p75NTR anymore. In SK‐N‐MC cells this leads to inhibition of NGF‐induced programmed cell death. In PC12 cells enhanced signaling through TrkA was observed. J. Neurosci. Res. 61:263–272, 2000. © 2000 Wiley‐Liss, Inc.

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The effects of the protein kinase inhibitor H-7 on early and delayed responses to nerve growth factor (NGF) were investigated in PC12 cells . H-7 reduced the NGF-induced expression of c-Fos in a dose-dependent manner without affecting the time course of c-Fos appearance . Conversely, H-7 potentiated delayed NGF effects, i .e ., neurite outgrowth and Ca t+/phospholipid-dependent protein kinase (PKC) induction, but not choline acetyltransferase induction . Long-term treatment with NGF resulted in an increase of at least four tyrosine-phosphorylated protein bands with molecular masses between 39 and 48 kDa, which was also potentiated by H-7 . In the absence of NGF, H-7 had no significant effect on c-Fos expression, tyrosine phosphorylation of the 45 kDa protein, or choline acetyltransferase activity . However, 4 days of exposure to H-7 alone induced PKC activity and tyrosine phosphorylation of the 39-kDa protein . The action of H-7 derivatives on neurite outgrowth did not correlate with their inhibition profile of cyclic nucleotide-dependent protein kinases . Down-regulation of PKC activity by prolonged exposure to phorbol ester did not completely abolish the effects of NGF and H-7 on induction of c-Fos, choline acetyltransferase activity, and neurite outgrowth, indicating that PKC-independent pathways contribute to these actions . These results suggest that additional pathways) sensitive to H-7 may exist, which induce immediate early gene expression and suppress neuronal differentiation of PC12 cells .