Continuous exposure to brain-derived neurotrophic factor is required for persistent activation of TrkB receptor, the ERK signaling pathway, and the induction of neuropeptide Y production in cortical cultures

We have previously demonstrated that brain-derived neurotrophic factor (BDNF) induces persistent neuropeptide Y (NPY) production in cortical cultures in an ERK1/2 -dependent manner. In some studies, it was shown that BDNF leads to the downregulation of TrkB receptor and some of its downstream responses, whereas in others it does not. We examined whether the BDNF requirement for induction of persistent NPY production correlates with that for induction of phosphorylation of TrkB and ERK 1/2. Continuous 24-h exposure to BDNF led to a 2- to 3-fold increase in NPY production (maximal level). While I h of BDNF exposure induced NPY production at a half maximal level, 8 h was required for induction of a maximal level. BDNF-induced NPY production was completely inhibited by co-exposure to TrkB-Fc fusion protein (TrkB extracellular domain fused to Fc) and partially inhibited by TrkB-Fc added 1 h after BDNF; TrkC-Fc did not do so. Activation of TrkB receptor was analyzed at two potential tyrosine phosphorylated sites, the activation loop and the Shc binding. BDNF led to coordinated phosphorylation of the two sites that persisted for 6 - 8 h, and this was not associated with changes in the content of TrkB protein. The presence of BDNF throughout the 6- to 8-h period was required for the persistent phosphorylation of TrkB and ERK1/2. Thus, continuous BDNF activation of TrkB is required for persistent activation of the ERK1/2 pathway and induction of NPY production. We propose that, within the time frame analyzed in this study, BDNF does not lead to the downregulation of TrkB receptor or of the biological responses leading to NPY production. (C) 2004 Elsevier B.V. All rights reserved.