The purpose of this study was to explore the role and mechanism of the PI3K/AKT/mTOR signaling pathway in painful diabetic neuropathy (PDN). The diabetes mellitus (DM) model was established by intraperitoneal injection of streptozocin into SD rats. After 3 weeks of modeling, the DM + LY group was treated with PI3K inhibitor, the DM + vehicle group was treated with DMSO, and the DM group was untreated. The paw mechanical withdrawal thresholds (MWT) was measured by Von Frey filaments, and the expression of PI3K/AKT/mTOR pathway-related proteins and autophagy marker proteins were analyzed by Western blotting. We found that 3 weeks after modeling, the MWT values of diabetic rats were significantly reduced, p-PI3K, p-AKT and p-mTOR proteins expression in the spinal cord was increased, and Beclin1 and LC3-II expressions were reduced (P < 0.05). After administration of PI3K inhibitor, the MWT values in DM + LY group were improved, and the expressions of p-PI3K, p-AKT and p-mTOR proteins in the spinal cord were decreased significantly, and the expressions of Beclin1 and LC3-II were increased (P < 0.05). However, there were no significant changes in the DM + vehicle group compared with the DM group (P > 0.05). Therefore, we conclude that activation of the PI3K/AKT/mTOR pathway and impaired autophagy may be key factors that cause PDN. Inhibition of the PI3K/AKT/mTOR pathway could promote autophagy activity and alleviate PDN.