Evidence for reverse flux through pyruvate kinase in skeletal muscle

Jin ES, Sherry AD, Malloy CR. Evidence for reverse flux through pyruvate kinase in skeletal muscle. Am J Physiol Endocrinol Metab 296: E748-E757, 2009. First published February 3, 2009; doi:10.1152/ajpendo.90935.2008.-Conversion of lactate to glucose was examined in myotubes, minced muscle tissue, and rats exposed to (H2O)-H-2 or C-13-enriched substrates. Myotubes or minced skeletal muscle incubated with [U-C-13(3)] lactate released small amounts of [1,2,3-C-13(3)]- or [4,5,6-C-13(3)] glucose. This labeling pattern is consistent with direct transfer from lactate to glucose without randomization in the tricarboxylic acid (TCA) cycle. After exposure of incubated muscle to (H2O)-H-2, [U-C-13(3)] lactate, glucose, and glutamine, there was minimal release of synthesized glucose to the medium based on a low level of H-2 enrichment in medium glucose but 50- to 100-fold greater H-2 enrichment in glucosyl units from glycogen. The C-13 enrichment pattern in glycogen from incubated skeletal muscle was consistent only with direct transfer of lactate to glucose without exchange in TCA cycle intermediates. C-13 nuclear magnetic resonance (NMR) spectra of glutamate from the same tissue showed flux from lactate through pyruvate dehydrogenase but not flux through pyruvate carboxylase into the TCA cycle. Carbon from an alternative substrate for glucose production that requires metabolism through the TCA cycle, propionate, did not enter glycogen, suggesting that TCA cycle intermediates do not exchange with phosphoenolpyruvate. In vivo, the C-13 labeling patterns in hepatic glycogen and plasma glucose after administration of [U-C-13(3)] lactate did not differ significantly. However, skeletal muscle glycogen was substantially enriched in [1,2,3-C-13(3)]- and [4,5,6-C-13(3)] glucose units that could only occur through skeletal muscle glyconeogenesis rather than glycogenesis. Lactate serves as a substrate for glyconeogenesis in vivo without exchange into symmetric intermediates of the TCA cycle.