Dale S. Edgerton

Insulin's direct effects on the liver dominate the control of hepatic glucose production

Insulin inhibits glucose production through both direct and indirect effects on the liver; however, considerable controversy exists regarding the relative importance of these effects. The first aim of this study was to determine which of these processes dominates the acute control of hepatic glucose production (HGP). Somatostatin and portal vein infusions of insulin and glucagon were used to clamp the pancreatic hormones at basal levels in the nondiabetic dog. After a basal sampling period, insulin infusion was switched from the portal vein to a peripheral vein. As a result, the arterial insulin level doubled and the hepatic sinusoidal insulin level was reduced by half. While the arterial plasma FFA level and net hepatic FFA uptake fell by 40-50%, net hepatic glucose output increased more than 2-fold and remained elevated compared with that in the control group. The second aim of this study was to determine the effect of a 4-fold rise in head insulin on HGP during peripheral hyperinsulinemia and hepatic insulin deficiency. Sensitivity of the liver was not enhanced by increased insulin delivery to the head. Thus, this study demonstrates that the direct effects of insulin dominate the acute regulation of HGP in the normal dog.

Physiologic action of glucagon on liver glucose metabolism

Glucagon is a primary regulator of hepatic glucose production (HGP) in vivo during fasting, exercise and hypoglycaemia. Glucagon also plays a role in limiting hepatic glucose uptake and producing the hyperglycaemic phenotype associated with insulin deficiency and insulin resistance. In response to a physiological rise in glucagon, HGP is rapidly stimulated. This increase in HGP is entirely attributable to an enhancement of glycogenolysis, with little to no acute effect on gluconeogenesis. This dramatic rise in glycogenolysis in response to hyperglucagonemia wanes with time. A component of this waning effect is known to be independent of hyperglycemia, though the molecular basis for this tachyphylaxis is not fully understood. In the overnight fasted state, the presence of basal glucagon secretion is essential in countering the suppressive effects of basal insulin, resulting in the maintenance of appropriate levels of glycogenolysis, fasting HGP and blood glucose. The enhancement of glycogenolysis in response to elevated glucagon is critical in the life‐preserving counterregulatory response to hypoglycaemia, as well as a key factor in providing adequate circulating glucose for working muscle during exercise. Finally, glucagon has a key role in promoting the catabolic consequences associated with states of deficient insulin action, which supports the therapeutic potential in developing glucagon receptor antagonists or inhibitors of glucagon secretion.

The direct and indirect effects of insulin on hepatic glucose production in vivo

IntroductionGlucose production by the liver is controlled on aminute to minute basis by the plasma insulin and glu-cagon concentrations. Glucagon increases glycogenbreakdown and stimulates the gluconeogenic path-way and insulin inhibits both glycogenolysis and glu-coneogenesis [1]. Until ten years ago it was generallyaccepted that the insulin level within the hepatic sinu-soids was responsible for the hormone’s inhibitory ef-fect on the liver. Recently, however, that concept hasbeen challenged.In 1987 this concept was first called into question[2]. It was noted that in obese non-diabetic humanssuppression of glucose production could occur in re-sponse to insulin infusion even when the estimatedportal insulin concentrations did not rise. Insulin wasinfused into a peripheral vein and euglycaemia wasmaintained with a glucose infusion. Endogenous in-sulin secretion, estimated from C-peptide concentra-tions, decreased by about 50% over the course ofthe experiment. Based on the fall in endogenous insu-lin release and the prevailing peripheral insulin con-centration, the authors estimated that the portal veininsulin concentration probably remained unchanged.Nevertheless glucose production was suppressed byabout 80% at the end of their study. It should be re-membered, however, that about 20% of liver bloodflow is derived from the hepatic artery, so a rise in ar-terial insulin would have increased the liver sinusoi-dal insulin concentration somewhat, even though theportal insulin level did not change. Likewise, al-though the authors assumed that their tracer datayielded an estimate of hepatic glucose production, itis now clear that it reflects both hepatic and renal glu-cose output [3]. After an overnight fast renal glucoseproduction is not great (5–25% of total glucose pro-duction) but it is insulin sensitive [4]. It is likely there-fore that a small portion of the insulin induced fall inglucose production which they observed was due toa decrease in renal glucose production. Despite theabove caveats, the study [2] was of importance be-cause it was the first to focus attention on the indirecteffect of insulin to inhibit glucose production by theliver.To further investigate the effect of peripherally de-livered insulin on glucose production, experimentswere carried out [5] in which conscious dogs were giv-en insulin intraportally (at 0.3 to 10 pmol × kg

Brain insulin action augments hepatic glycogen synthesis without suppressing glucose production or gluconeogenesis in dogs

In rodents, acute brain insulin action reduces blood glucose levels by suppressing the expression of enzymes in the hepatic gluconeogenic pathway, thereby reducing gluconeogenesis and endogenous glucose production (EGP). Whether a similar mechanism is functional in large animals, including humans, is unknown. Here, we demonstrated that in canines, physiologic brain hyperinsulinemia brought about by infusion of insulin into the head arteries (during a pancreatic clamp to maintain basal hepatic insulin and glucagon levels) activated hypothalamic Akt, altered STAT3 signaling in the liver, and suppressed hepatic gluconeogenic gene expression without altering EGP or gluconeogenesis. Rather, brain hyperinsulinemia slowly caused a modest reduction in net hepatic glucose output (NHGO) that was attributable to increased net hepatic glucose uptake and glycogen synthesis. This was associated with decreased levels of glycogen synthase kinase 3β (GSK3β) protein and mRNA and with decreased glycogen synthase phosphorylation, changes that were blocked by hypothalamic PI3K inhibition. Therefore, we conclude that the canine brain senses physiologic elevations in plasma insulin, and that this in turn regulates genetic events in the liver. In the context of basal insulin and glucagon levels at the liver, this input augments hepatic glucose uptake and glycogen synthesis, reducing NHGO without altering EGP.

Molecular Characterization of Insulin-Mediated Suppression of Hepatic Glucose Production In Vivo

OBJECTIVE Insulin-mediated suppression of hepatic glucose production (HGP) is associated with sensitive intracellular signaling and molecular inhibition of gluconeogenic (GNG) enzyme mRNA expression. We determined, for the first time, the time course and relevance (to metabolic flux) of these molecular events during physiological hyperinsulinemia in vivo in a large animal model. RESEARCH DESIGN AND METHODS 24 h fasted dogs were infused with somatostatin, while insulin (basal or 8× basal) and glucagon (basal) were replaced intraportally. Euglycemia was maintained and glucose metabolism was assessed using tracer, 2H2O, and arterio-venous difference techniques. Studies were terminated at different time points to evaluate insulin signaling and enzyme regulation in the liver. RESULTS Hyperinsulinemia reduced HGP due to a rapid transition from net glycogen breakdown to synthesis, which was associated with an increase in glycogen synthase and a decrease in glycogen phosphorylase activity. Thirty minutes of hyperinsulinemia resulted in an increase in phospho-FOXO1, a decrease in GNG enzyme mRNA expression, an increase in F2,6P2, a decrease in fat oxidation, and a transient decrease in net GNG flux. Net GNG flux was restored to basal by 4 h, despite a substantial reduction in PEPCK protein, as gluconeogenically-derived carbon was redirected from lactate efflux to glycogen deposition. CONCLUSIONS In response to acute physiologic hyperinsulinemia, 1) HGP is suppressed primarily through modulation of glycogen metabolism; 2) a transient reduction in net GNG flux occurs and is explained by increased glycolysis resulting from increased F2,6P2 and decreased fat oxidation; and 3) net GNG flux is not ultimately inhibited by the rise in insulin, despite eventual reduction in PEPCK protein, supporting the concept that PEPCK has poor control strength over the gluconeogenic pathway in vivo.

Effects of Insulin on the Metabolic Control of Hepatic Gluconeogenesis In Vivo

OBJECTIVE Insulin represses the expression of gluconeogenic genes at the mRNA level, but the hormone appears to have only weak inhibitory effects in vivo. The aims of this study were 1) to determine the maximal physiologic effect of insulin, 2) to determine the relative importance of its effects on gluconeogenic regulatory sites, and 3) to correlate those changes with alterations at the cellular level. RESEARCH DESIGN AND METHODS Conscious 60-h fasted canines were studied at three insulin levels (near basal, 4×, or 16×) during a 5-h euglycemic clamp. Pancreatic hormones were controlled using somatostatin with portal insulin and glucagon infusions. Glucose metabolism was assessed using the arteriovenous difference technique, and molecular signals were assessed. RESULTS Insulin reduced gluconeogenic flux to glucose-6-phosphate (G6P) but only at the near-maximal physiological level (16× basal). The effect was modest compared with its inhibitory effect on net hepatic glycogenolysis, occurred within 30 min, and was associated with a marked decrease in hepatic fat oxidation, increased liver fructose 2,6-bisphosphate level, and reductions in lactate, glycerol, and amino acid extraction. No further diminution in gluconeogenic flux to G6P occurred over the remaining 4.5 h of the study, despite a marked decrease in PEPCK content, suggesting poor control strength for this enzyme in gluconeogenic regulation in canines. CONCLUSIONS Gluconeogenic flux can be rapidly inhibited by high insulin levels in canines. Initially decreased hepatic lactate extraction is important, and later reduced gluconeogenic precursor availability plays a role. Changes in PEPCK appear to have little or no acute effect on gluconeogenic flux.

Current strategies for the inhibition of hepatic glucose production in type 2 diabetes.

Diabetes is a complex disease involving multiple organs with dysregulation in glucose and lipid metabolism. Hepatic insulin insensitivity can contribute to elevated fasting glucose levels and impaired glucose tolerance in individuals with diabetes. Several currently available therapeutics address defects at the liver. Metformin inhibits glucose production, potentially through effects on AMPK. Thiazolidinediones activate PPAR-gamma and improve hepatic insulin sensitivity, primarily through indirect effects on lipid metabolism. Insulin analogs and secretagogues suppress glucose production and increase liver glucose utilization by both direct and indirect hepatic actions. Incretins, incretin mimetics, and dipeptidyl peptidase-4 inhibitors reduce postprandial hepatic glucose production by increasing insulin secretion and limiting glucagon release, as well as through possible direct effects on the liver. Pramlintide reduces the increase in plasma glucagon that occurs following a meal in individuals with diabetes, and may thereby suppress inappropriate stimulation of liver glucose production. Many other hepatic targets are being considered which may lead to alternative strategies for the treatment of diabetes. This review focuses on currently available therapeutics which target insulin resistance in the liver.

A Novel Glucagon Receptor Antagonist, NNC 25-0926, Blunts Hepatic Glucose Production in the Conscious Dog

Elevated glucagon is associated with fasting hyperglycemia in type 2 diabetes. We assessed the effects of the glucagon receptor antagonist (2R)-N-[4-({4-(1-cyclohexen-1-yl)[(3,5-dichloroanilino)carbonyl]anilino}methyl)benzoyl]-2-hydroxy-b-alanine (NNC 25-0926) on hepatic glucose production (HPG) in vivo, using arteriovenous difference and tracer techniques in conscious dogs. The experiments consisted of equilibration (–140 to –40 min), control (40–0 min), and experimental [0–180 min, divided into P1 (0–60 min) and P2 (60–180 min)] periods. In P1, NNC 25-0926 was given intragastrically at 0 (veh), 10, 20, 40, or 100 mg/kg, and euglycemia was maintained. In P2, somatostatin, basal intraportal insulin, and 5-fold basal intraportal glucagon (2.5 ng/kg/min) were infused. Arterial plasma insulin levels remained basal throughout the study in all groups. Arterial plasma glucagon levels remained basal during the control period and P1 and then increased to ∼70 pg/ml in P2 in all groups. Arterial plasma glucose levels were basal in the control period and P1 in all groups. In P2, the arterial glucose level increased to 245 ± 22 and 172 ± 15 mg/dl in the veh and 10 mg/kg groups, respectively, whereas in the 20, 40, and 100 mg/kg groups, there was no rise in glucose. Net hepatic glucose output was ∼2 mg/kg/min in all groups during the control period. In P2, it increased by 9.4 ± 2 mg/kg/min in the veh group. In the 10, 20, 40, and 100 mg/kg groups, the rise was only 4.1 ± 0.9, 1.6 ± 0.6, 2.4 ± 0.7, and 1.5 ± 0.3 mg/kg/min, respectively, due to inhibition of glycogenolysis. In conclusion, NNC 25-0926 effectively blocked the ability of glucagon to increase HGP in the dog.

Evidence against a Physiologic Role for Acute Changes in CNS Insulin Action in the Rapid Regulation of Hepatic Glucose Production

This Perspective will discuss the physiologic relevance of data that suggest CNS insulin action is required for the rapid suppression of hepatic glucose production. It will also review data from experiments on the conscious dog, which show that although the canine brain can sense insulin and, thereby, regulate hepatic glucoregulatory enzyme expression, CNS insulin action is not essential for the rapid suppression of glucose production caused by the hormone. Insulins direct hepatic effects are dominant, thus it appears that insulins central effects are redundant in the acute regulation of hepatic glucose metabolism.

Glucagon as a Critical Factor in the Pathology of Diabetes

Studies from the laboratory of Roger Unger presented in the current issue of Diabetes highlight the potential benefit of reducing glucagon action by examining the effects of glucagon receptor knockout (Gcgr−/−) on the phenotype of type 1 diabetes in the mouse (1). The aim of the study was to determine if glucagon action, by itself, causes the lethal consequences of insulin deficiency. Because treatment of Gcgr−/− mice with the β-cell toxin streptozotocin (STZ) previously had no effect on circulating insulin levels or pancreatic islet architecture (2), Lee et al. (1) administered a double dose of STZ to maximize β-cell destruction. Unlike STZ treated wild-type Gcgr+/+ mice, which became severely hyperglycemic, STZ-treated mice lacking glucagon signaling appeared to be in a normal state of health and were completely protected from the manifestations of diabetes (1), as shown previously by the same group in alloxan treated Gcgr−/− mice (3) and by Hancock et al. (4) in STZ-treated mice lacking glucagon because of α-cell deletion. Fasting hyperglycemia did not occur in STZ-treated Gcgr−/− mice, and astonishingly, the animals demonstrated normal or even improved glucose disposal in response to a glucose tolerance test, despite the absence of a rise in plasma insulin. These results led the authors to speculate that insulin action during glucose absorption is largely directed toward overcoming the hepatic actions of glucagon. They theorized that insulin would have little or no role in a liver not exposed to the action of glucagon because it would be in a permanent glucose storage mode. Glucagon antagonistic peptides, neutralizing antibodies, receptor antisense oligonucleotides, and/or receptor nonpeptidyl antagonists have previously been shown to lower plasma glucose in several rodent models of diabetes (5,6). Likewise, reversal of diabetes by leptin therapy in the rodent has been attributed to a …