NT3 inhibits FGF2-induced neural progenitor cell proliferation via the PI3K/GSK3 pathway

Neurotrophin 3 (NT3), a member of the neurotrophin family, antagonizes the proliferative effect of fibroblast growth factor 2 (FGF2) on cortical precursors. However, the mechanism by which NT3 inhibits FGF2-induced neural progenitor (NP) cell proliferation is unclear. Here, using an FGF2-dependent rat neurosphere culture system, we found that NT3 inhibits both FGF2-induced neurosphere growth and bromodeoxyuridine (BrdU) incorporation in a dose-dependent manner. U0126, a mitogen-activated protein kinase kinase 1/2 (MEK1/2) inhibitor, and LY294002, a phosphatidylinositol 3-kinase (PI3K) inhibitor, both inhibited FGF2-induced BrdU incorporation, suggesting that the extracellular signal-regulated kinase1/2 (ERK1/2) and PI3K pathways are required for FGF2-induced NP cell proliferation. NT3 significantly inhibited FGF2-induced phosphorylation of Akt and glycogen synthase kinase 3beta (GSK3beta), a downstream kinase of Akt, whereas phosphorylation of ERK1/2 was unaffected. The inhibitory effect of NT3 on FGF2-induced NP cell proliferation was abolished by LY294002, and treatment with SB216763, a specific GSK3 inhibitor, antagonized the NT3 effect, rescuing both neurosphere growth and BrdU incorporation. Moreover, experiments with anti-NT3 antibody revealed that endogenous NT3 also plays a role in inhibiting FGF2-induced NP cell proliferation, and that anti-NT3 antibody enhanced phospho-Akt and phospho-GSK3beta levels in the presence of FGF2. These findings indicate that FGF2-induced NP cell proliferation is inhibited by NT3 via the PI3K/GSK3 pathway.