J. Biol. Chem., Vol. 263, Issue 32, 16725-16730, Nov, 1988
Model to examine pathways of carbon flux from lactate to glucose at the first branch point in gluconeogenesis
WG Blackard and JN Clore
Department of Medicine, Medical College of Virginia, Richmond 23298.
The first branch point in gluconeogenesis occurs at the conversion of
pyruvate to oxaloacetate. To determine the amount of lactate carbon
reaching glucose via the direct pyruvate carboxylase pathway versus the
tricarboxylic acid cycle, adult rat hepatocytes in primary culture were
incubated for 2 h with one of the following isotopic substrates: [1-
14C]lactate, [U-14C]lactate, or [1,2-14C]acetate. Production of 14CO2 and
[14C]glucose from each substrate was assessed. The amount of lactate carbon
2 and 3 incorporated into glucose or oxidized to CO2 was determined by
subtracting values using [1-14C]lactate from those using [U-14C]lactate.
After quantitation of CO2 formed from carbons 2 and 3 of lactate, the
amount of these carbons incorporated into glucose via the tricarboxylic
acid cycle can be determined by simple proportionality from the ratio of
label incorporated into glucose or CO2 from [1,2-14C]acetate. The remaining
carbons 2 and 3 of lactate incorporated into glucose are derived from the
pyruvate carboxylase pathway directly. Ethanol which on oxidation provides
NADH and acetate decreased lactate oxidation and enhanced the pyruvate
carboxylase pathway. Glucagon increased carbon flux through both pathways
but primarily through the pyruvate carboxylase pathway. In summary, a
simple model is presented to examine carbon flux from lactate via the
pyruvate carboxylase and tricarboxylic acid pathways during
gluconeogenesis.
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