TCA Cycle Models: Implications for Tracer Estimates of Gluconeogenesis

Abstract

A fundamental lueasurement in the assessment of the metabolic status of man and experimental animal is the rate of gluconeogenesis. Yet, reliable methods for measaring this important indicator are not available. A common experimental procedure is to expose gluconeogenic tissue to a labeled carbon molecule that is metabolized to glucose (acetate or pyruvate, for example) and measure the rate of appearance of labeled carbon in glucose. This method requires a correction factor for isotope dilution as the labeled carbon intersects with the tricarboxylic acid (TCA) cycle en route to glucose. Correction for isotope dilution in this instance requires a modeling approach to evaluate quantitatively the intersection of the two pathways. An important feature of TCA cycle models in current use for estimating isotope dilution is the assumption that no carbon enters the TCA cycle except the gluconeogenic flux from pyruvate to oxaloacetate and the flux of acetyl CoA to citrate that resupplies two carbon units to the cycle. We have evaluated the impact of additional fluxes exchanging carbon with TCA cycle pools in the pathway between citrate and oxaloacetate (example, exchange of glutamate with 2-oxyglutarate). Our major findings are: (1) Estimates of the flux of pyruvate to oxaloacetate (y) derived from the glucose labeling ratio are affected by this assumption. Thus, values of y obtained from the classic model are incorrect if additional exchange of carbon occurs. (2). Estimates of gluconeogenesis from tracer pyruvate are strongly dependent on this assumption that carbon enters the TCA cycle only from pyruvate and acetyl CoA. (3). Estimates of glucoeogenesis from carbon tracers yielding labeled acetyl CoA are not affected by this assumption and no error is introduced due to additional fluxes into and out of the TCA cycle. This work illustrates the value of algebraic models of the TCA cycle and related pathways as an aid in developing improved practical tracer methods for estimating gluconeogenesis.