Background: Metastasis remains one of the greatest challenges involved in treating gastric cancer (GC). Ropivacaine (Rop) is not only a well-documented local anesthetic medicament but also has been reported to exert an antitumor role in cancer development. This study explored the effects of ropivacaine on the growth, migration and invasion of gastric cancer and the underlying mechanisms. Methods: Cell Counting Kit-8 (CCK8) assay was conducted to test the effect of Rop on the proliferation of AGS and BGC-823 GC cells. Moreover, cell apoptosis, migration and invasion were examined by flow cytometry and transwell assay, respectively. The expression of miR-520a-3p was determined by qRT-PCR. miRNA targeting sites were analyzed using bioinformatics analysis and dual-luciferase reporter assay. Protein levels of WEE1 and PI3K/ AKT were detected by Western blot. Furthermore, the tumor-forming experiment of nude mice was used to detect the growth of cells in vivo. Results: Rop inhibited proliferation but promoted apoptosis of GC cells. Besides, the migration and invasion of GC cells were also inhibited by Rop. Moreover, miR-520a-3p expression was enhanced by Rop, and transfection with miR-520a-3p mimic decreased cell proliferation, migration and invasion. The upregulation of miR-520a-3p was partly contributed to the inhibitory effect of ropivacaine on GC cell lines. Finally, Rop inactivated WEE1 and PI3K/AKT pathway via upregulation of miR-520a-3p. Conclusion: Our results suggested that Rop decreased growth, migration and invasion of GC cells via regulating miR-520a-3p expression and further inactivated WEE1 and PI3K/ AKT signaling pathways.