Mechanism of liver glycogen repletion in vivo by nuclear magnetic resonance spectroscopy.

Abstract

In order to quantitate the pathways by which liver glycogen is repleted, we administered 11-'3Cjglucose by gavage into awake 24-h fasted rats and examined the labeling pattern of 'IC in hepatic glycogen. Two doses of i1-'3Cjglucose, 1 and 6 mg/g body wt, were given to examine whether differences in the plasma glucose concentration altered the metabolic pathways via which liver glycogen was replenished. After 1 and 3 h (high-dose group) and after 1 and 2 h (low-dose group), the animals were anesthetized and the liver was quickly freeze-clamped. Liver glycogen was extracted and the purified glycogen hydrolyzed to glucose with amyloglucosidase. The distribution of the "3C-label was subsequently determined by "C-nuclear magnetic resonance spectroscopy. The percent 'IC enrichment of the glucosyl units in glycogen was: 15.1±0.8%(C-1), 1.5±0.1%(C-2), 1.2±0.1%(C-3), 1.1±0.1%(C-4), 1.6±0.1%(C-5), and 2.2±0.1%(C-6) for the high-dose study (n = 4, at 3 h); 16.5±0.5%(C-1), 2.0±0.1%(C-2), 1.3±0.1%(C-3), 1.1±0.1%(C4), 2.2±0.1%(C-5), and 2.4±0.1%(C-6) in the low-dose study (n = 4, at 2 h). The average "3C-enrichment of C-1 glucose in the portal vein was found to be 43±1 and 40±2% in the high-and low-dose groups, respectively. Therefore, the amount of glycogen that was synthesized from the direct pathway (i.e., glucose -glucose-6-phosphate -glucose-l-phosphate -UDP-glucose -glycogen) was calculated to be 31 and 36% in the high-and low-dose groups, respectively. The "C-enrichments of portal vein lactate and alanine were 14 and 14%, respectively, in the high-dose group and 11 and 8%, respectively, in the low-dose group. From these enrichments, the minimum contribution of these gluconeogenic precursors to glycogen repletion can be calculated to be 7 and 20% in the high-and low-dose groups, respectively. The maximum contribution of glucose recycling at the triose isomerase step to glycogen synthesis (i.e., glucose -triose-phosphatesglycogen) was estimated to be 3 and 1% in the high-and lowdose groups, respectively.In conclusion, our results demonstrate that (a) only one-third of liver glycogen repletion occurs via the direct conversion of glucose to glycogen, and that (b) only a very small amount of glycogen synthesis can be accounted for by the conversion of glucose to triose phosphates and back to glycogen; this suggests that futile cycling between fructose-6-phosphate and fructose-1,6-diphosphate under these conditions is minimal. Our results also show that (c) alanine and lactate account for a minimum of Receivedfor publication 22 February 1985. between 7 and 20% of the glycogen synthesized, and that (d) the three pathways through which the labeled flux is measured account for a total of only 50% of the total glycogen synthesized. These results suggest that either there is a sizeable amount of glycogen synthesis via pathway(s) that were not examined in the present experiment or that there is a much greater dilution of labeled alanine/lactate in the oxaloacetate pool than previously appreciated, or some combination of...

Inclusion of Low Amounts of Fructose With an Intraduodenal Glucose Load Markedly Reduces Postprandial Hyperglycemia and Hyperinsulinemia in the Conscious Dog

“…The frozen samples were stored at Ϫ70°C for subsequent analysis. On the day of the assay, samples were powdered and the glycogen was extracted and purified as previously described (42). The cold glycogen concentrations were determined by acid hydrolysis and enzyme degradation using ␣-1,4,6-amyloglucosidase (43).…”

Quantitative analysis of glycogen repletion by nuclear magnetic resonance spectroscopy in the conscious rat.

“…There is increasing evidence that the majority ofhepatic glycogen repletion after an oral glucose load in rat and man occurs via a gluconeogenic process (1)(2)(3)(4)(5)(6)(7)(8). It has been hypothesized that lactate and alanine are the primary gluconeogenic precursors for glycogen formation (2,(4)(5)(6).…”

“…It has been hypothesized that lactate and alanine are the primary gluconeogenic precursors for glycogen formation (2,(4)(5)(6). In a previous study using nuclear magnetic resonance spectroscopy (NMR),' we estimated that a minimum of 10-28% of newly formed glycogen was derived from alanine/lactate (6).…”

“…It has been hypothesized that lactate and alanine are the primary gluconeogenic precursors for glycogen formation (2,(4)(5)(6). In a previous study using nuclear magnetic resonance spectroscopy (NMR),' we estimated that a minimum of 10-28% of newly formed glycogen was derived from alanine/lactate (6). Quantitative contribution ofthese precursors to glycogenesis has been difficult to determine due to the exchange of carbon label in the oxaloacetate (OAA) pool, however, which serves as a common intermediate both in the gluconeogenic pathway and in the tricarboxylic (TCA) cycle (9-1 1).…”

Contributions of Hepatic Gluconeogenesis Suppression and Compensative Glycogenolysis on the Glucose-Lowering Effect of CS-917, a Fructose 1,6-Bisphosphatase Inhibitor, in Non-obese Type 2 Diabetes Goto-Kakizaki Rats

“…3: a and c. Shulman et al reported that glycogen is directly synthesized from a gluconeogenesis substrate such as lactate and alanine as well as from glucose (26). The mechanism by which CP-91149 decreased plasma glucose with increasing hepatic glycogen content was not fully understood, but these results suggested that suppressing breakdown of glycogen synthesized from gluconeogenesis substrate might be one of the mechanisms.…”