Abstract: To elucidate the specific molecular mechanisms by which α-ketoglutarate (AKG) regulates fat metabolism in zebrafish, this study demonstrated that oral administration of palm oil feed or ex vivo incubation with palmitic acid (PA) reduced the expression of AKG-metabolism-related genes in zebrafish livers or hepatocytes, thereby decreasing AKG levels. This study revealed that feeding palm oil in vivo or incubation with exogenous palmitic acid (PA) reduced the expression of AKG metabolism-related genes in zebrafish livers or hepatocytes, thereby decreasing AKG levels. Further experiments demonstrated that exogenous supplementation of AKG alleviates PA-induced AKG metabolic disturbances while modulating the expression of peroxisome proliferator-activated receptors (PPARs), thereby promoting fatty acid β-oxidation, inhibiting lipogenesis, and reducing lipid deposition. Conversely, inhibition of endogenous AKG synthesis using glutamate dehydrogenase inhibitor R162suppressed fatty acid β-oxidation, enhanced lipogenesis, and induced hepatic lipid deposition. Additionally, PA-induced abnormal lipid deposition was associated with aberrant activation of the mTORC1signaling pathway, and exogenous AKG supplementation effectively inhibited this activation. In conclusion, feeding palm oil may induce AKG metabolic disturbances, activate the mTORC1signaling pathway, alter PPARs expression, subsequently inhibit fatty acid β-oxidation, promote lipogenesis, and ultimately lead to hepatic lipid deposition in zebrafish. These findings provides a theoretical foundation for understanding how AKGregulateslipid metabolism in fish and identify potential regulatory targets for the efficient utilization of palm oil feeds.