In carnivorous fish, conversion of a glucose load to hepatic glycogen is widely used to assess their metabolic flexibility towards carbohydrate utilization, but the activities of direct and indirect pathways in this setting are unclear. We assessed the conversion of an intraperitoneal glucose load (2 g.kg−1) enriched with [U-13C6]glucose to hepatic glycogen in juvenile seabass and seabream. 13C-NMR analysis of glycogen was used to determine the contribution of the load to glycogen synthesis via direct and indirect pathways at 48-hr post-injection. For seabass, [U-13C6]glucose was accompanied by deuterated water and 2H-NMR analysis of glycogen 2H-enrichment, allowing endogenous substrate contributions to be assessed as well. For fasted seabass and seabream, 47 ± 5% and 64 ± 10% of glycogen was synthesized from the load, respectively. Direct and indirect pathways contributed equally (25 ± 3% direct, 21 ± 1% indirect for seabass; 35 ± 7% direct, 29 ± 4% indirect for seabream). In fasted seabass, integration of 2H- and 13C-NMR analysis indicated that endogenous glycerol and anaplerotic substrates contributed an additional 7 ± 2% and 7 ± 1%, respectively. In fed seabass, glucose load contributions were residual and endogenous contributions were negligible. Concluding, direct and indirect pathways contributed equally and substantially to fasting hepatic glycogen repletion from a glucose load in juvenile seabream and seabass.