Measurement of metabolic fluxes through pyruvate kinase, phosphoenolpyruvate carboxykinase, pyruvate dehydrogenase, and pyruvate carboxylase in hepatocytes of different acinar origin

Isolated hepatocytes were prepared from the periportal and perivenous regions of the liver of 18-h-starved rats, These showed characteristic enzyme patterns and an enhanced rate of ureagenesis in the periportal cells; however, total cellular ATP content was unchanged in the two cell types, Measurements of pyruvate kinase flux showed no significant difference in the overall rate in the two cell types; however, the flux through phosphoenolpyruvate (PEP) carboxykinase was significantly higher in the periportal cells, such that the percentage of PEP being metabolized by pyruvate kinase was enhanced in the perivenous cells, The increase in partitioning of PEP through pyruvate kinase could account for only a small percentage of the difference in gluconeogenic flux in the two cell types, suggesting that the rate of provision of PEP was the principal limiting factor for glucose synthesis, The flux through pyruvate dehydrogenase showed no significant metabolic zonation, whereas pyruvate carboxylase flux was enhanced in the periportal zone, The partitioning of pyruvate between pyruvate carboxylase and pyruvate dehydrogenase was increased 2.8-fold in the periportal cells compared to that in the perivenous cells and it is suggested that this, together with possible alterations in phosphoenolpyruvate carboxykinase, is primarily responsible for the different gluconeogenic rates in the two zones of the liver. (C) 1996 Academic Press, Inc.