The aim of this study was to further confirm that glial cell line-derived neurotrophic factor (GDNF) exerts a neuro-protective effect on dopaminergic neurons (DAs) and to investigate the protective mechanism. Cadherins are calcium-dependent adhesion proteins, and N-cadherins are found in neurons. Our study attempted to ascertain whether GDNF activates the PI3K/Akt signaling pathway through the mediation of N-cadherin to confer a protective effect on DAs. Flow cytometry and Hoechst 33258 staining results indicated that the apoptosis rate of damaged neurocytes increased following interference of N-cadherin expression. Immunoblotting results demonstrated that the amount of phosphorylated Akt (p-Akt) in the cytoplasm decreased, while the total Akt quantity remained unchanged following interference of N-cadherin expression. The immunohistochemical staining results demonstrated that the levels of total N-cadherin, phosphorylated N-cadherin (Tyr860) and p-Akt decreased; however, the amount of total Akt remained unchanged. In addition, we also demonstrated that Tyr860 and p-Akt levels were reduced in a GDNF dose-dependent manner with the phosphorylation level peaking at GDNF dose of 50 ng/ml (in vitro) and 50 ng/4 µl (in vivo), and also in a time-dependent manner with the phosphorylation level peaking at 15 min (in vitro) and 30 min (in vivo). Statistical analysis also showed that changes in the phosphorylation levels of Tyr860 and p-Akt demonstrated a positive correlation. Collectively, GDNF activates the PI3K/Akt pathway via N-cadherin to protect DAs.