Background: Gastric cancer (GC) is a considerable health burden around the world. Circular RNA Nance-Horan syndrome-like 1 (circNHSL1) is reported to be highly expressed in GC. Nevertheless, the function and molecule mechanism of circNHSL1 are still unclear. Methods: The expression levels of circNHSL1, microRNA-149-5p (miR-149-5p) and YWHAZ were detected by real-time quantitative polymerase chain reaction (RT-qPCR). The subcellular fractionation identified the remarkable cytoplasmic localization of circNHSL1. Cell migration and invasion were measured by transwell assays. The levels of glutamine, glutamate and α-ketoglutarate (α-KG) were assessed by the corresponding kit. The protein levels of CD63, CD9, CD81, alanine, serine, cysteine-preferring transporter 2 (ASCT2), glutaminase 1 (GLS1), and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta (YWHAZ) were detected by Western blot assay. The binding relationship between miR-149-5p and circNHSL1 or YWHAZ was predicted by starBase 3.0 and then verified by RNA pull-down and dual-luciferase reporter assays. Xenograft tumor model examined the biological role of circNHSL1 in vivo. Exosomes were examined by a transmission electron microscope and nanoparticle tracking analysis (NTA). Results: CircNHSL1 was highly expressed in GC cell-derived exosomes, GC tissues, and cells. Its knockdown impeded GC cell migration, invasion, and glutaminolysis. Mechanism analysis showed that circNHSL1 could affect YWHAZ expression by sponging miR-149-5p, thereby regulating GC progression. CircNHSL1 downregulation blocked GC tumor growth in vivo. Conclusion: Our studies disclosed that circNHSL1 knockdown repressed migration, invasion, and glutaminolysis in vitro and inhibited tumor growth in vivo by miR-149-5p/ YWHAZ axis in GC, implying an underlying circRNA-targeted therapy for GC treatment.