Brain-derived neurotrophic factor (BDNF) plays an important role in neuronal growth; however, the downstream regulatory mechanisms remain unclear. In this study, we investigated whether BDNF exerts its neurotrophic effects through the Wnt/beta-catenin signaling pathway in human embryonic spinal cord neurons in vitro. We found that neuronal growth (soma size and average neurite length) was increased by transfection with a BDNF overexpression plasmid. Western blotting and real-time quantitative PCR showed that expression of the BDNF pathway components TrkB, PI3K, Akt and PLC-gamma was increased by BDNF overexpression. Furthermore, the Wnt signaling factors Wnt, Frizzled and Dsh and the downstream target beta-catenin were upregulated, whereas GSM-3 beta was downregulated. In contrast, when BDNF signaling was downregulated with BDNF siRNA, the growth of neurons was decreased. Furthermore, BDNF signaling factors, Wnt pathway components and beta-catenin were all downregulated, whereas GSM-3 beta was upregulated, This suggests that BDNF affects the growth of neurons in vitro through crosstalk with Wnt signaling, and that GSM-3 beta may be a critical factor linking these two pathways. To evaluate this possibility, we treated neurons with 6-bromoindirubin-3'-oxime (BIO), a small molecule GSM-3 beta inhibitor. BIO reduced the effects of BDNF upregulation/downregulation on soma size and average neurite length, and suppressed the impact of BDNF modulation on the Wnt signaling pathway. Taken together, our findings suggest that BDNF promotes the growth of neurons in vitro through crosstalk with the Wnt/beta-catenin signaling pathway, and that this interaction may be mediated by GSK-3 beta. (C) 2015 Elsevier Ltd. All rights reserved.