Regulation of L-pyruvate kinase activity by insulin and glycolytic intermediates.

The presence of insulin and various carbohydrates in long term cultures of hepatocytes was studied to examine the mechanism by which insulin and glycolytic metabolites regulate L-pyruvate kinase activity. When hepatocytes were isolated from a control rat and cultured in the presence of insulin, a constant level of enzyme activity (16 EU/mg DNA) was maintained for 12 days. The L-pyruvate kinase activity in hepatocytes from refed rats initially was elevated (45 EU/mg DNA) and decreased to control values by the 5th day in culture. Cells isolated from a fasted rat initially contained a low level of L-pyruvate kinase activity (5 EU/mg DNA) which increased to control values by the 8th day in culture. The enzyme activity was 5 EU/mg DNA when control cells were cultured for 4 days in medium containing either glucose, glycerol or fructose without insulin; 10 EU/mg DNA in medium containing galactose and insulin but without glucose, glycerol or fructose; and 20 EU/mg DNA in medium containing both insulin and either glucose, glycerol or fructose. It is suggested that insulin is essential for the induction and maintainance of L-pyruvate kinase activity, that carbohydrates in the absence of insulin are unable to maintain the enzyme activity in cultures of hepatocytes, and that insulin and glycolytic metabolites may act synergistically to increase the activity of L-pyruvate kinase by increasing the synthesis of the enzyme.