The relationship between gluconeogenic precursor supply and glucose production has been investigated in 14-h and 86-h fasted humans. In protocols 1 and 2 [6,6-2H]glucose and [15N2]urea were infused to measure glucose and urea production rates (Ra) in response to infusions of glycerol and alanine. In protocol 3 first [15N]alanine, [3-13C]lactate, and [6,6-2H]glucose were infused before and during administration of dichloroacetate (DCA) to determine the response of glucose Ra to decreased fluxes of pyruvate, alanine, and lactate, then alanine was infused with DCA and glucose Ra measured. After a 14-h fast, neither alanine nor glycerol increased glucose Ra. Basal glucose Ra decreased by one-third after 86 h of fasting, yet glycerol and alanine infusions had no effect on glucose Ra. Glycerol always reduced urea Ra (P less than 0.05), suggesting that glycerol competitively inhibited gluconeogenesis from amino acids. DCA decreased the fluxes of pyruvate, alanine (P less than 0.01), and glucose Ra (P less than 0.01), which was prevented by alanine infusion. These findings suggest that 1) the reduction in glucose Ra after an 86-h fast is not because of a shortage of gluconeogenic substrate; 2) nonetheless, the importance of precursor supply to maintain basal glucose Ra is confirmed by the response to DCA; 3) an excess of one gluconeogenic substrate inhibits gluconeogenesis from others.

When an isotopic tracer is infused for the purpose of determining the rate of turnover or oxidation of a substrate, it is assumed that the resulting isotopic enrichment by the tracer will reflect the kinetics of only the pool of interest. However, this may not be the case when carbon-labeled lactate is infused, since rapid isotopic exchange with the intracellular pyruvate and alanine pools could potentially occur. Therefore we have determined the extent of isotopic exchange occurring during the infusion of [3-13C]lactate into six anesthetized dogs. In the steady state, pyruvate enrichment was 91 +/- 2.2% (means +/- SE) of the lactate enrichment, and alanine enrichment was 81 +/- 3.3% of the pyruvate enrichment and 72 +/- 2.6% of the lactate enrichment. In contrast, when [3-13C]alanine was infused (n = 2), pyruvate (and lactate) enrichment was 9.9% of the alanine enrichment. We therefore conclude that there is rapid isotopic equilibration between lactate and pyruvate but that interaction with alanine reflects the true metabolic flux rates, rather than isotopic exchange. Consequently, lactate kinetics, as traditionally determined, more accurately reflect whole body pyruvate kinetics.

The validity of the primed constant-infusion tracer technique to make short-term measurements of urea production rates (Ra) in humans in a physiological steady state and during disruption of steady state was evaluated. Four subjects received a primed constant infusion (P/I = 560 min) of [13C]urea for 8 h. A plateau in urea enrichment was reached after 2 h and maintained throughout. When [13C]- and [18O]urea were simultaneously infused into four subjects at P/I ratios of 560:1 and 360:1, respectively, both tracers reached plateau enrichment at the same time (2-4 h). The enrichment at plateau was a function of the infusion rate rather than the priming dose, and calculated urea Ra was the same with either prime. In five additional experiments the technique responded acutely to a physiological perturbation (alanine infusion) in a dose-dependent manner. The results confirm that this technique is appropriate for short-term measurements of urea Ra, and the requirement for accuracy in estimating the priming dose is not impractically stringent.

The relationship between the rate of lipolysis and rate of glucose production (Ra) was investigated in 14- and 86-h fasted humans. [6,6-2H]glucose and [2H]5glycerol were infused to measure glucose and glycerol Ra in response to infusions of nicotinic acid in 14- and 86-h fasted subjects (protocol 1). The response of glucose Ra to nicotinic acid alone and nicotinic acid plus unlabeled glycerol was also measured in 86-h fasted subjects (protocol 2). After a 14-h fast, nicotinic acid caused a 30% decrease in plasma insulin levels and a marked (66%) decrease in plasma free fatty acid levels but did not have any significant effect on glucose Ra and concentration. After 86 h of fasting, nicotinic acid decreased glycerol Ra and hence lipolytic rate by approximately 60%. This caused a significant decrease (P less than 0.05) of 16-20% in glucose Ra and uptake. This decrease in glucose Ra was abolished when unlabeled glycerol was also infused with nicotinic acid to maintain glycerol Ra. These findings suggest that, in normal humans, a decrease in the rate of lipolysis regulates glucose Ra via its effect on the availability of glycerol for gluconeogenesis.

The importance of the location of isotope infusion and blood sampling on calculating glucose kinetics was studied in five mongrel dogs in the basal state and when glucose turnover was increased during a hyperinsulinemic euglycemic clamp. [U-14C]glucose was infused into the pulmonary artery, unlabeled glucose was infused into a femoral vein to maintain euglycemia, and blood was sampled from the right atrium (AV mode) and the femoral artery (VA mode). In the basal state there was no difference between the AV or VA mode in plateau specific activity; hence, the calculated rate of appearance of glucose was the same with either mode. During the euglycemic clamp procedure, plateau specific activity of the AV mode was significantly lower than that of the VA mode (P less than 0.05). The rate of appearance of glucose calculated from the VA mode was almost identical to the rate of infusion of unlabeled glucose (13.0 +/- 1.4 vs. 12.6 +/- 1.4 mg.kg-1.min-1, respectively), but the rate of appearance of glucose calculated from the AV mode was 12% greater. This study demonstrates that the calculation of glucose kinetics is sensitive to differences in sampling site when the turnover rate is high relative to the mass flow rate (cardiac output times substrate concentration).