Rainbow trout (Oncorhynchus mykiss) fasted for 4 months had lower plasma glucose concentration and hepatocyte glycogen content than the fed fish. Gluconeogenesis from alanine in the fasted fish was not significantly different from that of the fed fish, whereas both alanine glyconeogenesis and oxidation increased with fasting by 400% and 300%' respectively. Glyconeogenesis from glucose was not significantly different with fasting, whereas glucose oxidation decreased significantly with fasting. The presence of glucose (10 mM) in the medium did not modify alanine glyconeogenesis or gluconeogenesis in either the fed or the fasted groups. However, glucose did increase alanine oxidation in the fed group but not in the fasted group, this response being further stimulated in the presence of insulin. Alanine (2 mM) in the medium had no effect on glyconeogenesis from glucose in either fed or fasted trout hepatocytes. Insulin significantly decreased glucose oxidation in the fed fish, whereas the insulin effect on glucose oxidation in the fasted fish occurred only in the presence of alanine. The results indicate that nutritional state affects hepatocyte responsiveness to substrate utilization and insulin stimulation in trout. Specifically, the hepatocyte potential for glyconeogenesis is enhanced in the fasted fish, although this increased glycogen production is not due to an increased glucose utilization. o 1995 Wiley-Liss, Inc.