Gardenia (Gardenia jasminoides Ellis) is a food homologous Chinese herbal medicine, often indicated for its hypolipidemic and anti-inflammatory effects. Geniposide is the main active ingredient in gardenia, which has been shown to improve lipid metabolism disorders. This work aims to debate the hypolipidemic effect of geniposide and resolve whether its hypolipidemic mechanism is mediated by the glucocorticoid receptor (GR). In vitro experiments demonstrated that geniposide was able to alleviate oleic acid (OA) + palmitic acid (PA) - induced lipid accumulation in AML12 cells and down-regulate glucose 6-phosphatase (G6Pase), phosphoenolpyruvate carboxykinase 1 (PCK1), fructose-1,6-bisphosphatase 1 (FBP1), stearoyl-CoA desaturase (SCD1), pyruvate carboxylase (PC), sterol regulatory element binding protein-1c (SREBP1-C), CCAAT/enhancer binding protein β (C/EBPβ) gluconeogenesis and lipid metabolism-related genes and proteins expression level. Furthermore, we constructed a hyperlipidaemic zebrafish model to further investigated the alleviating effect of geniposide on hyperlipidaemia in vivo. To explain the mechanism of action of geniposide, a series of experiments revealed that geniposide could bind to GR, inhibit dexamethasone-induced GR nuclear translocation and display GR antagonistic activity. To confirm whether the therapeutic effect of geniposide was mediated by GR, we performed GR knockdown experiments in AML12 cells, and then discovered that GR knockdown abolished the inhibition of G6pase, PCK1, FBP1, SCD1, SREBP1-C expression by geniposide. Since geniposide can ameliorate hyperlipidaemia in zebrafish, it might be used as a potential source in the development of dietary supplements.