Carbon and hydrogen metabolism of xylitol and various sugars in human erythrocytes.

1) Erythrocytes are able to metabolize D-ribose, D-xylitol, D-xylulose, D-fructose and D-glucose; the rates of metabolism increase in that order from 2430 to 26200 ng atom C/ml packed cells per 120 min of incubation. 2) The utilization of the carbon of these substrates and its recovery in the products were found to be in balance, when the change in the 2,3-bisphosphoglycerate concentration was taken into account. 3) The metabolic rates strongly affected the 2,3-bisphosphoglycerate level. Without addition of substrate the decomposition rate of this intermediate was found to be 1030 nmol/ml packed cells per 120 min. 4) The net decrease of the 2,3-bisphosphoglycerate concentration and the conversion of this compound into lactate provides a NAD regeneration system which enables the red blood cells to utilize xylitol. 5) The rate of carbon metabolism via the pentose phosphate cycle is determined by the NADPH requirement of the erythrocytes which was found to be 160 nmol/ml packed cells per 120 min under the experimental conditions employed.     

Carbon balance studies with various substrates in human erythrocytes.

Carbon metabolism in erythrocytes has been found to be in balance with a variety of substrates studied. The contribution of the 2,3-bisphosphoglycerate pathway to carbon metabolism depends on the rates of carbon utilization and increases with increasing metabolic rates. Net decrease of the 2,3-bisphosphoglycerate level acts as a NAD regenerating system thus facilitating uptake of polyalcohols such as xylitol.     

Xylitol production from sugarcane bagasse hydrolyzate in fluidized bed reactor. Effect of air flowrate

Cells of Candida guilliermondii immobilized onto porous glass spheres were cultured batchwise in a fluidized bed bioreactor for xylitol production from sugarcane bagasse hemicellulose hydrolyzate. An aeration rate of only 25 mL/min ensured minimum yields of xylose consumption (0.60) and biomass production (0.14 g(DM)/g(Xyl)), as well as maximum xylitol yield (0.54 g(Xyt)/g(Xyl)) and ratio of immobilized to total cells (0.83). These results suggest that cell metabolism, although slow because of oxygen limitation, was mainly addressed to xylitol production. A progressive increase in the aeration rate up to 140 mL/min accelerated both xylose consumption (from 0.36 to 0.78 g(Xyl)/L.h) and xylitol formation (from 0.19 to 0.28 g(Xyt)/L.h) but caused the fraction of immobilized to total cells and the xylitol yield to decrease up to 0.22 and 0.36 g(Xyt)/g(Xyl), respectively. The highest xylitol concentration (17.0 g(Xyt)/L) was obtained at 70 mL/min, but the specific xylitol productivity and the xylitol yield were 43% and 22% lower than the corresponding values obtained at the lowest air flowrate, respectively. The concentrations of consumed substrates and formed products were used in material balances to evaluate the xylose fractions consumed by C. guilliermondii for xylitol production, complete oxidation through the hexose monophosphate shunt, and cell growth. The experimental data collected at variable oxygen level allowed estimating a P/O ratio of 1.35 mol(ATP)/mol(O) and overall ATP requirements for biomass growth and maintenance of 3.4 mol(ATP)/C-mol(DM).     

Impact of low-flow ischemia on substrate oxidation and glycolysis in the isolated perfused rat heart

Interventions that stimulate carbohydrate oxidation appear to be beneficial in the setting of myocardial ischemia or infarction. However, the mechanisms underlying this protective effect have not been defined, in part because of our limited understanding of substrate utilization under ischemic conditions. Therefore, we used (1)H and (13)C NMR spectroscopy to investigate substrate oxidation and glycolytic rates in a global low-flow model of myocardial ischemia. Isolated male Sprague-Dawley rat hearts were perfused for 30 min under conditions of normal flow (control) and low-flow ischemia (LFI, 0.3 ml/min) with insulin and (13)C-labeled lactate, pyruvate, palmitate, and glucose at concentrations representative of the physiological fed state. Despite a approximately 50-fold reduction in substrate delivery and oxygen consumption, oxidation of all exogenous substrates plus glycogen occurred during LFI. Oxidative metabolism accounted for 97% of total calculated ATP production in the control group and approximately 30% in the LFI group. For controls, lactate oxidation was the major source of ATP; however, in LFI, this shifted to a combination of oxidative and nonoxidative glycogen metabolism. Interestingly, in the LFI group, anaplerosis relative to citrate synthase increased sevenfold compared with controls. These results demonstrate the importance of oxidative energy metabolism for ATP production, even during very-low-flow ischemia. We believe that the approach described here will be valuable for future investigations into the underlying mechanisms related to the protective effect of increasing cardiac carbohydrate utilization and may ultimately lead to identification of new therapeutic targets for treatment of myocardial ischemia.     

Red cells of newborn rats have low bisphosphoglyceromutase and high pyruvate kinase activities in association with low 2,3-bisphosphoglycerate

2,3-Bisphosphoglycerate is a physiologically important regulator of red cell oxygen affinity during mammalian development. The rat has no fetal hemoglobin, but the newborn red cell has low 2,3-bisphosphoglycerate and high ATP concentrations, and high oxygen affinity. This report shows that red cell bisphosphoglyceromutase activity increases from near zero in the newborn rat to very high levels by four weeks of age. This increase roughly parallels the increase in red cell 2,3-bisphosphoglycerate concentration. Red cell pyruvate kinase activity declines ten-fold from birth to four weeks of age. This decrease is associated with a changeover in red cell populations from larger to smaller cells. The glycolytic rate is at least 50% higher in newborn than adult rat red cells. The data suggest that high pyruvate kinase activity and glycolytic rate contribute to the high ATP concentration in newborn rat red cells, but that their low 2,3-bisphosphoglycerate concentration is due primarily to low bisphosphoglyceromutase activity.     

2,3-Bisphosphoglycerate inhibits ATP-stimulated proteolysis

Intracellular protein breakdown could be regulated at the substrate level by changes in the environment. Under in vitro conditions, ATP increases the proteolytic susceptibility of several mitochondrial and cytosolic proteins, while 2,3-bisphosphoglycerate not only has the opposite effect but also prevents the ATP-stimulated proteolysis. ATP and 2,3-bisphosphoglycerate, present at relatively high levels in many tissues, provide a good model of environmental components that may influence intracellular proteolysis.     

Metabolism of pyruvate and carnitine esters in bovine epididymal sperm mitochondria

Filipin-treated bovine epididymal spermatozoa have been used to study mitochondrial l-acetylcarnitine, l-palmitoylcarnitine, and pyruvate metabolism. The cells were supplemented with malate to allow rapid rates of substrate oxidation. The rate of l-palmitoylcarnitine-supported state 3 respiration was slow. In contrast, pyruvate, acetylcarnitine, or lactate supported rapid and approximately equal respiratory rates. l-Palmitoylcarnitine was a weak inhibitor of pyruvate-supported respiration and pyruvate use and a more potent inhibitor of l-acetylcarnitine. l-Carnitine was an effective inhibitor of l-acetylcarnitine oxidation; however, it did not influence l-palmitoylcarnitine oxidation or inhibit pyruvate utilization. Pyruvate (1.4 mm) disappearance was rapid and was complete within 6–7 min; the lactate produced during pyruvate metabolism was then oxidized. ATP synthesis was constant throughout the 20-min incubation. With pyruvate plus l-acetylcarnitine as substrate, the l-acetylcarnitine concentration initially dropped and then recovered to a level that was dependent on free carnitine addition. Data obtained from experiments using [2-¹⁴C]pyruvate indicated that the ¹⁴C label from pyruvate and lactate entered the l-acetylcarnitine pool and labeling was maximal when free l-carnitine was added. The rate of citrate synthesis was maximal when pyruvate was being metabolized; the largest total accumulation occurred when all three substrates were included in the incubation. The data suggest that the high NAD⁺/ NADH maintained during pyruvate metabolism may restrict flux through the citric acid cycle. The relationships of l-carnitine and the l-carnitine esters to pyruvate metabolism are discussed.     

Inhibitory effect of mitoxantrone on activity of protein kinase C and growth of HL60 cells.

Mitoxantrone, a new anthraquinone, showed inhibitory an effect on protein kinase C (PKC) activity. Its IC50 value was 4.4 micrograms/ml (8.5 microM), which is much lower than those of the well-known anthracyclines daunorubicin and doxorubicin, the IC50 values of which are more than 100 micrograms/ml (> 170 microM). Kinetic studies demonstrated that mitoxantrone inhibited PKC in a competitive manner with respect to histone H1, and its Ki value was 6.3 microM (Ki values of daunorubicin and doxorubicin were 0.89 and 0.15 mM, respectively), and in a non-competitive manner with respect to phosphatidylserine and ATP. Inhibition of phosphorylation by mitoxantrone was observed with various substrates including S6 peptide, myelin basic protein and its peptide substrate derived from the amino-terminal region. Their IC50 values were 0.49 microgram/ml (0.95 microM), 1.8 micrograms/ml (3.5 microM), and 0.82 microgram/ml (1.6 microM), respectively. Mitoxantrone did not markedly inhibit the activity of cyclic AMP-dependent protein kinase, casein kinase I or casein kinase II, at concentrations of less than 10 micrograms/ml. On the other hand, brief exposure (5 min) of HL60 cells to mitoxantrone caused the inhibition of cell growth with an IC50 value of 52 ng/ml (0.1 microM). In HL60 cells, most of the PKC activity (about 90%) was detected in the cytosolic fraction. When HL60 cells exposed to 10 micrograms/ml mitoxantrone for 5 min were observed with fluorescence microscopy, the fluorescence elicited from mitoxantrone was detected in the extranuclear area. These results indicated that mitoxantrone is a potent inhibitor of PKC, and this inhibition may be one of the mechanisms of antitumor activity of mitoxantrone.     

D-xylulose-induced depletion of ATP and Pi in isolated rat hepatocytes

Xylitol is known to cause hepatic ATP catabolism by inducing the trapping of Pi in the form of glycerol 3-P as a consequence of an increase in the NADH:NAD+ ratio, resulting from the oxidation of xylitol to D-xylulose. The question was therefore raised whether D-xylulose also depletes hepatic ATP. In isolated rat hepatocytes, 5 mM D-xylulose decreased ATP by 80% within 5 min compared to 40% with 5 mM xylitol. Intracellular Pi decreased by 70% within the same time interval with both compounds, but was restored three-fold faster with D-xylulose. The rate of utilization of D-xylulose reached 5 mumol.min-1.g-1 of cells, as compared with 1.5 for xylitol, indicating that reduction of xylitol into D-xylulose is a rate-limiting step in the metabolism of the polyol. D-Xylulose barely modified the concentration of glycerol 3-P but increased xylulose 5-P from 0.02 to 0.5 mumol/g within 5 min. The main cause of the ATP- and Pi-depleting effects of D-xylulose was found to be an accumulation of sedoheptulose 7-P from a basal value of 0.1 to 5 mumol/g of cells after 10 min. Ribose 5-P increased from 0.03 to 0.5 mumol/g at 5 min. Ribose 1-P also accumulated, albeit outside of the cells. This extracellular accumulation can be explained by the release of intracellular purine nucleoside phosphorylase from damaged hepatocytes acting on inosine that had diffused out of the cells. Smaller increases in the concentrations of sedoheptulose 7-P and pentose phosphates were recorded after incubations of the cells with xylitol.     

Metabolism of 3-phosphoglyceroyl phosphate in phosphoenolpyruvate-enriched human erythrocytes

The concentration of 3-phosphoglyceroyl phosphate in erythrocytes was increased by more than 100-fold when red cells were incubated with extracellular phosphoenolpyruvate at 37 degrees C. Since these elevated levels were maintained for 60 min, the metabolism of 3-phosphoglyceroyl phosphate and related compounds could be investigated in phosphoenolpyruvate-treated erythrocytes. 2,3-Bisphosphoglycerate synthesis was not affected by intracellular pH when the 3-phosphoglyceroyl phosphate level was constant but did vary with 3-phosphoglyceroyl phosphate concentration. On the other hand, the relationship between the rate of 2,3-bisphosphoglycerate synthesis and 3-phosphoglyceroyl phosphate concentration was not straightforward. At relatively low concentrations of 3-phosphoglyceroyl phosphate, the observed rate of 2,3-bisphosphoglycerate synthesis agreed with a rate calculated from a formula incorporating kinetic parameters of purified 2,3-bisphosphoglycerate synthase (Rose, Z.B. (1973) Arch. Biochem. Biophys. 158, 903-910). However, at high concentrations of 3-phosphoglyceroyl phosphate, the observed rate of 2,3-bisphosphoglycerate synthesis was lower than the calculated value. The concentration of glucose 1,6-bisphosphate did not increase even when 3-phosphoglyceroyl phosphate was elevated to 200 microM. Elevated levels of intracellular 2,3-bisphosphoglycerate did not inhibit glycolytic activity in these erythrocytes. These results suggest that incubation of erythrocytes with phosphoenolpyruvate is a useful technique to investigate the effect of metabolic perturbations at the intermediate stages of glycolysis.     

Synthesis and evaluation of a classical 2,4-diamino-5-substituted-furo[2,3-d]pyrimidine and a 2-amino-4-oxo-6-substituted-pyrrolo[2,3-d]pyrimidine as antifolates

Two classical antifolates, a 2,4-diamino-5-substituted furo[2,3-d]pyrimidine and a 2-amino-4-oxo-6-substituted pyrrolo[2,3-d]pyrimidine, were synthesized as potential inhibitors of dihydrofolate reductase (DHFR) and thymidylate synthase (TS). The syntheses were accomplished by condensation of 2,6-diamino-3(H)-4-oxo-pyrimidine with alpha-chloro-ketone 21 to afford two key intermediates 23 and 24, followed by hydrolysis, coupling with l-glutamate diethyl ester and saponification of the diethyl ester to afford the classical antifolates 13 and 14. Compounds 13 and 14 with a single carbon atom bridge are both substrates for folylpoly-gamma-glutamate synthetase (FPGS), the enzyme responsible for forming critical poly-gamma-glutamate antifolate metabolites with increased potency and/or increased cell retention. Compound 14 is a highly efficient FPGS substrate demonstrating that 2,4-diamino-5-substituted furo[2,3-d]pyrimidines are important lead structures for the design of antifolates with FPGS substrate activity. It retains inhibitory potency for DHFR and TS compared to the two atom bridged analog 5. Compound 13 is a poor inhibitor of purified DHFR and TS, and both 13 and 14 are poor inhibitors of the growth of CCRF-CEM human leukemia cells in culture, indicating that single carbon bridged compounds in these series though conducive to FPGS substrate activity were not potent inhibitors.     

14C-labeled substrate catabolism by human diploid fibroblasts derived from infants and adults

Untransformed diploid skin fibroblasts from eight normal adults, aged 24 to 74 years, catabolized several 14C-labeled substrates less effectively than cells from ten normal male infants. 14C-labeled substrate metabolism was quantitated either by measuring the evolution of 14CO2 from the 14C-labeled compounds or the incorporation of 14C into cellular protein via transamination of tricarboxylic acid cycle intermediates derived from the 14C-labeled substrates. With these methods, adult cells catabolized [1-14C]butyrate, [1-14C]octanoate, and 1-[2-14C]leucine at rates 44 to 64% of those found in infant cells. The oxidation of [1,4-14C]succinate and [U-14C]malate was identical in both infant and adult cells, while [2,3-14C]succinate catabolism was mildly decreased in adult cells (65-80% of control). These observations parallel those made in rat tissues and confirm that the same phenomenon occurs in cultured human fibroblasts.     

The relationship between glucose concentration and rate of lactate production by human erythrocytes in an open perfusion system

A thermodynamically open system, based on an assembly of capillaries with semi-permeable walls was constructed in order to study glycolysis in human erythrocytes in high haematocrit suspensions. A phenomenological expression for the rate of lactate production as a function of glucose concentration was obtained. The rate was measured under steady-state conditions with low substrate concentrations (approx. 50 μmol/l). In a corresponding closed system, this concentration of glucose would be exhausted within a few minutes. A mathematical model of the whole system consisted of five differential equations, and involved parameters relating to flow rates, volumes of reaction chambers, the rates of lactate efflux from erythrocytes and the expression for the rate of lactate production by red cells. The binding of [¹⁴C]pyruvate to haemoglobin and the rate of efflux of [¹⁴C]lactate from red cells were measured to yield additional information for the model. The concentrations of ATP and 2,3-bisphosphoglycerate were measured during the perfusion experiments, and a detailed analysis of a model of red cell hexokinase was carried out; the former two compounds inhibit hexokinase and alter the apparent K_m and V_max for glucose in vivo. These steady-state parameters were similar to the glucose concentration at the half-maximal rate of lactate production and the maximal rate, respectively. These findings are consistent with the known high control-strength for hexokinase in glycolysis in human red cells. The practical and theoretical validation of this perfusion system indicates that it will be valuable for NMR-based studies of red cell metabolism using a flow-cell in the spectrometer.     

Metabolic interactions between glucose, glycerol, alanine and acetate in Leishmania braziliensis panamensis promastigotes

13C-nuclear magnetic resonance (NMR) spectroscopy was used to investigate the products of glycerol and acetate metabolism released by Leishmania braziliensis panamensis promastigotes and also to examine the interaction of each of these substrates with glucose or alanine. The NMR data were supplemented by measurements of the rates of oxygen consumption and substrate utilization, and of 14CO2 production from 14C-labeled substrate. Cells incubated with [2-13C]glycerol released acetate, succinate and D-lactate in addition to CO2. Cells incubated with acetate released only CO2. More succinate C-2/C-3 than C-1/C-4 was released from both [2-13C]glycerol and [2-13C]glucose, indicating that succinate was formed predominantly by CO2 fixation followed by reverse flux through part of the Krebs cycle. Some redistribution of the position of labeling was also seen in alanine and pyruvate, suggesting cycling through pyruvate/oxaloacetate/phosphoenolpyruvate. Cells incubated with combinations of 2 substrates consumed oxygen at the same rate as cells incubated with 1 or no substrate, even though the total substrate utilization had increased. When promastigotes were incubated with both glycerol and glucose, the rate of glucose consumption was unchanged but glycerol consumption decreased about 50%, and the rate of 14CO2 production from [1,(3)-14C]glycerol decreased about 60%. Alanine did not affect the rates of consumption of glucose or glycerol, but decreased 14CO2 production from these substrates by increasing flow of label into alanine. Although glucose decreased alanine consumption by 70%, it increased the rate of 14CO2 production from [U-14C]- and [l-14C]alanine by about 20%.(ABSTRACT TRUNCATED AT 250 WORDS)     

2,3-Bisphosphoglycerate,fructose, 2,6-bisphosphate and glucose 1,6-bisphosphate during maturation of reticulocytes with low 2,3-bisphosphoglycerate content

In contrast to the species with erythrocytes of high 2,3-bisphosphoglycerate content, in the sheep the concentration of 2,3-bisphosphoglycerate decreases during maturation of reticulocytes. The decrease can be explained by the drop of the phosphofructokinase/pyruvate kinase and 2,3-bisphosphoglycerate synthase/2,3-bisphosphoglycerate phosphatase activity ratios that result from the decline of phosphofructokinase, pyruvate kinase, phosphoglycerate mutase and the bifunctional enzyme 2,3-bisphosphoglycerate synthase/phosphatase. The concentrations of fructose 2,6-bisphosphate and aldohexose 1,6-bisphosphates also decrease during sheep reticulocyte maturation in parallel to the 6-phosphofructo 2-kinase and the glucose 1,6-bisphosphate synthase activities.     

Theoretical phosphorylation rates after addition of a small amount of glucose to intact ascites tumor cells

Changes were measured in glycolytic and respiratory rates during the entire period of glycolysis and respiratory inhibition after addition of 0.08 or 0.15 mM glucose to Ehrlich ascites carcinoma cells in 54 mM phosphate buffer (pH 7.3) at 37°. Glycolytic products fully accounted for the glucose utilized.

Theoretical rates of glycolytic ATP synthesis were calculated from the rates of accumulation of glycolytic products, and rates of oxidative phosphorylation were calculated from respiratory rates, assuming a P:O ratio of 3.0. The maximum in the glycolytic phosphorylation rate curve preceded the minimum in the respiratory phosphorylation rate curve. As a consequence, the total phosphorylation rate curve was biphasic, first rising above, then falling below, and finally returning to the initial, pre-glucose rate. The area under the early rise approximately equalled the area above the later dip and corresponded to between 1 and 2 μmoles of ATP/ml cells. The low rate of change in the ATP content of the cells indicated that most of the change in phosphorylation rate represented changes in both ATP synthesis and ATP utilization.

It is hypothesized that ATP synthesized by glycolysis is more readily available to the ATP-utilizing systems. On addition of glucose, ATP is shifted from a respiratory to a glycolytic reservoir and a period of more rapid ATP utilization associated with a decrease in the level of endogenous substrates involved in the ATP-utilizing reactions ensues; after cessation of glycolysis, the process is reversed, and ATP utilization is slowed for a period while the endogenous substrates increase again.     

Rates of utilization and fates of glucose, glutamine, pyruvate, fatty acids and ketone bodies by mouse macrophages.

The concentrations of ATP and the ATP/AMP concentration ratios were maintained in thioglycollate-elicited mouse peritoneal macrophages incubated in vitro for 90 min in the presence or absence of added substrate: rates of glycolysis, lactate formation and glutamine utilization were approximately linear with time for at least 60 min of incubation. The rate of oxygen consumption by macrophages was only increased above the basal rate (i.e. that in the absence of added substrate) by addition of succinate or pyruvate, or by addition of the uncoupling agent carboxyl cyanide m-chlorophenylhydrazone ('CCCP'); it was decreased by 75% by the addition of KCN. These findings suggest that metabolism of endogenous substrate can provide most, if not all, of the energy requirement of these cells, at least for a short period. The rates of glucose and glutamine utilization by incubated macrophages were approx. 300 and 100 nmol/min per mg of protein respectively. A large proportion of the glutamine that is utilized is converted into glutamate and aspartate, and very little (perhaps less than 10%) is oxidized. Similarly almost all of the glucose that is utilized is converted into lactate and very little is oxidized. This characteristic is similar to that of resting lymphocytes and rapidly dividing cells; in non-proliferating macrophages it may be a mechanism to provide precision in control of the rate of biosynthetic processes that utilize intermediates of these pathways, e.g. purines and pyrimidines for mRNA for the synthesis of secretory proteins and glycerol 3-phosphate for phospholipid synthesis for membrane recycling. No utilization of acetoacetate or 3-hydroxybutyrate by macrophages was detected. In contrast, both butyrate and oleate were oxidized. The rate of [14C]oleate conversion into 14CO2 (1.3 nmol/h per mg of protein) could account for most of the oxygen consumption by incubated macrophages, suggesting that long-chain fatty acids might provide an important fuel in situ. This may be one explanation for the secretion of lipoprotein lipase by these cells, to provide fatty acids for oxidation from the degradation of local triacylglycerol.     

Carbohydrate metabolism of hepatocytes from starved Japanese quail

Hepatocytes were isolated from livers of mature male and female starved Japanese quail (Coturnix coturnix japonica). The hepatocytes take up lactate and dihydroxyacetone extensively, and have a very high rate of glucose synthesis from these substrates. Fructose uptake and incorporation into glucose is much less. Pyruvate and alanine are taken up extensively, but form little glucose. There is negligible lipogenesis in cells of starved quail. Alanine increases up to 10-fold incorporation of ³HOH and ¹⁴C from several substrates into fatty acids, but it remains insignificant as compared to lipogenesis by cells of fed quail. There is little utilization of glucose, even in the presence of alanine, in marked contrast to hepatocytes from fed quail. However, glucose is phosphorylated at high rates, but most of the glucose 6-phosphate is recycled to glucose. There is a marked difference in the metabolism of polyols between the sexes. Glycerol, xylitol, and sorbitol are converted nearly quantitatively into glucose by hepatocytes of starved female quail. In cells of starved males, the uptake of polyols is higher, but conversion to glucose less efficient. In cells of starved male quail, alanine markedly stimulates the uptake of glycerol and xylitol and their conversion to glucose, but has no effect on sorbitol metabolism. In cells of female quail, alanine is without a significant effect on polyol metabolism.     

Regulation of carbohydrate metabolism in mouse liver. Effect of glucagon on gluconeogenic/glycolytic flux in isolated perfused livers.

1. Glucagon stimulated gluconeogenesis from both [U-14C]lactate and [14C]xylitol in isolated perfused mouse liver. 2. Addition of cyclic AMP also stimulated gluconeogenesis from [U-14C]lactate. 3. Glucagon caused a rapid (2.5 min) 12-fold increase in hepatic cyclic AMP but not cyclic GMP concentration. 4. Glucagon caused a rapid and stable decrease in hepatic fructose 1,6-diphosphatase activity measured in vitro. 5. The results are interpreted to indicate that glucagon stimulates hepatic gluconeogenesis in mice via cyclic AMP by two different mechanisms: (a) increased substrate uptake (i.e. utilization) and (b) increased gluconeogenic efficiency (i.e. inhibition of alternate substrate fates).