Download Glycogen branches out: new perspectives on the role of glycogen

Am J Physiol Endocrinol Metab 291: E1–E8, 2006;
doi:10.1152/ajpendo.00652.2005.
Invited Review
Glycogen branches out: new perspectives on the role of glycogen metabolism
in the integration of metabolic pathways
Cynthia C. Greenberg, Michael J. Jurczak, Arpad M. Danos, and Matthew J. Brady
Department of Medicine, Committee on Molecular Metabolism and Nutrition, the University of Chicago, Chicago, Illinois
GLYCOGEN STRUCTURE AND KEY ENZYMES OF
ITS METABOLISM
GLYCOGEN SYNTHESIS follows a simple but strictly ordered
process, resulting in a complex structure (38, 54). The initiation of glycogen synthesis is provided by self-glucosylation of
glycogenin (55). To this oligosaccharide primer, glycogen
synthase utilizes UDP-glucose to add glucose molecules by
␣1– 4 linkages, which is the rate-controlling step of glycogen
synthesis. Glycogen synthase activity is controlled by covalent
modification, allosteric activation, and enzymatic translocation. The enzyme is phosphorylated on up to nine residues,
resulting in its progressive inactivation and decreased sensitivity to allosteric activators (31). Binding of glucose 6-phosphate
(G-6-P) to glycogen synthase causes unfolding of the enzyme,
resulting in allosteric activation that overrides inhibition by
phosphorylation. In addition, G-6-P binding promotes conformational changes that favor dephosphorylation of the enzyme.
Translocation of glycogen synthase to glycogen particles in
response to stimuli such as insulin represents a third mechanism by which enzymatic activity is regulated (13, 42, 49).
When the elongating glycogen chain consists of at least 11
residues, branching enzyme transfers a chain of seven molecules to another chain by an ␣1– 6 bond. Thus glycogen
synthase elongates the glycogen chain, and branching enzyme
produces new branches, creating a molecule with a helical
structure of 12 concentric tiers (38).
For glycogen degradation, the synchronous activities of
glycogen phosphorylase and debranching enzyme are required.
Address for reprint requests and other correspondence: M. J. Brady,
MC1027, 5841 S. Maryland Ave., Chicago, IL 60637 (e-mail:
[email protected]).
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Glycogen phosphorylase, which catalyzes the rate-limiting step
of glycogenolysis, cleaves ␣1– 4 linkages to remove glucose
molecules from the glycogen chain. When four glucose units
remain before a branch point, the transferase activity of debranching enzyme catalyzes the transfer of three glucose residues to an adjacent branch of the glycogen chain. Through a
second enzymatic glucosidase component located in the same
protein, debranching enzyme next cleaves the ␣1– 6 bond to
release the glucose moiety from the branch point. Glycogen
phosphorylase is then able to continue removal of glucose
residues from the glycogen chain (38).
Glycogen phosphorylase is regulated by phosphorylation of
a single serine residue at the NH2 terminus and by allosteric
binding of a number of molecules, including AMP, ATP,
G-6-P, glucose, and caffeine (25). The enzyme exists as a
dimer with each subunit linked to the essential cofactor pyridoxal phosphate, which provides the phosphate as an electron
donor for release of glucose 1-phosphate. The covalent modification of glycogen phosphorylase in response to changes in
intracellular cAMP and calcium levels is mediated by the
activation of phosphorylase kinase, the sole enzyme that phosphorylates and activates glycogen phosphorylase (5). Thus
both glycogen synthase and phosphorylase are regulated by
multiple mechanisms that enable glycogen metabolism to be
acutely responsive to the varying metabolic demands in both
liver and skeletal muscle cells.
SKELETAL MUSCLE GLYCOGEN
During rest, storage of glucose as glycogen is enhanced by
insulin stimulation of glucose uptake in muscle, which is
mediated by the translocation of GLUT4 transporters to the
plasma membrane (62). Entering glucose is phosphorylated to
0193-1849/06 $8.00 Copyright © 2006 the American Physiological Society
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Greenberg, Cynthia C., Michael J. Jurczak, Arpad M. Danos, and Matthew
J. Brady. Glycogen branches out: new perspectives on the role of glycogen
metabolism in the integration of metabolic pathways. Am J Physiol Endocrinol
Metab 291: E1–E8, 2006; doi:10.1152/ajpendo.00652.2005.—Glycogen is the
storage form of carbohydrate for virtually every organism from yeast to primates.
Most mammalian tissues store glucose as glycogen, with the major depots located
in muscle and liver. The French physiologist Claude Bernard first identified a
starch-like substance in liver and muscle and coined the term glycogen, or “sugar
former,” in the 1850s. During the 150 years since its identification, researchers in
the field of glycogen metabolism have made numerous discoveries that are now
recognized as significant milestones in biochemistry and cell signaling. Even so,
more questions remain, and studies continue to demonstrate the complexity of the
regulation of glycogen metabolism. Under classical definitions, the functions of
glycogen seem clear: muscle glycogen is degraded to generate ATP during
increased energy demand, whereas hepatic glycogen is broken down for release of
glucose into the bloodstream to supply other tissues. However, recent findings
demonstrate that the roles of glycogen metabolism in energy sensing, integration of
metabolic pathways, and coordination of cellular responses to hormonal stimuli are
far more complex.
Invited Review
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NEW PERSPECTIVES ON THE ROLE OF GLYCOGEN METABOLISM
Glycogen Metabolism During Exercise
Immediate fuel utilization. At the beginning of a bout of
exercise, contraction of skeletal muscle results in the hydrolysis of ATP and a transient rise in ADP and Pi levels. Creatine
phosphate/creatine kinase acts as a cellular buffer for ATP
levels by providing a source of Pi that can be added to ADP to
form ATP by creatine kinase. However, this system is quickly
overwhelmed during sustained exercise, and increasing ADP
levels stimulate the activity of adenylate kinase, which catalyzes the conversion of two ADP molecules to one ATP and
one AMP molecule. As the ATP from this pathway is rapidly
consumed, the AMP-to-ATP ratio rises, initiating a number of
metabolic events geared toward maintaining energy required
for contraction.
Regulation of glycolysis by glycogen levels. As the aforementioned immediate energy sources are depleted, the muscle
cell must then rely on the glycolytic pathway to provide ATP
AJP-Endocrinol Metab • VOL
for continued contraction. Glycolysis is initially activated by
the increases in AMP and Pi, which stimulate the rate-limiting
enzyme in glycolysis, phosphofructokinase. Glycolysis is stimulated as well by an increase in provision of G-6-P to this
pathway, which occurs at the onset of exercise initially due to
augmented rates of glycogenolysis and, as exercise continues,
through an increase in glucose uptake via the contractioninduced, insulin-independent translocation of GLUT4 vesicles
to the plasma membrane (24). The metabolism of G-6-P by the
glycolytic pathway results in the generation of fructose 2,6bisphosphate, which is a potent allosteric activator of phosphofructokinase, and thus a key metabolic regulator of glycolytic
rates and ATP production (53).
A variety of stimuli are integrated in the regulation of
glycogen phosphorylase activity, allowing glycogen breakdown to change in parallel with the energy demands of the
exercising muscle. The onset of contraction results in the
release of calcium stores from the sarcoplasmic reticulum,
causing the activation of phosphorylase kinase and subsequent
phosphorylation and activation of glycogen phosphorylase.
The majority of glycogen phosphorylase and phosphorylase
kinase in the cell is found at the sarcoplasmic reticulum bound
to glycogen (32, 39, 63), allowing for the coupling of contraction, calcium release, and glycogen breakdown. Glycogen
phosphorylase is also allosterically activated by increased [Pi]
within the cell, which rises as ATP levels fall, reflecting an
energy deficit. As activity continues, glycogen phosphorylase
may also be activated in response to increased extracellular
epinephrine levels, which result in activation of PKA, which in
turn phosphorylates and activates phosphorylase kinase.
Tight metabolic coupling exists between glycogen mobilization and ATP production via glycolysis. In fact, studies
indicate that the G-6-P arising from muscle glycogen breakdown is more efficiently coupled to glycolysis than that arising
from GLUT4 and hexokinase, particularly during high-intensity exercise, when 80% of the glucose consumed by glycolysis
arises from glycogenolysis (56). Additionally, at moderateintensity exercise, the rate of glycogenolysis greatly exceeded
the rate of glucose transport, suggesting a greater contribution
of glycogen breakdown to maintenance of ATP levels. Thus, at
the onset of exercise, the provision of G-6-P to the glycolytic
pathway from glycogen breakdown would inhibit extracellular
glucose transport by both allosteric inhibition of hexokinase II
and diminishment of the glucose concentration gradient across
the plasma membrane.
Glucose transport during exercise. Due to the finite nature
of glycogen stores, as contraction continues glycogen phosphorylase activity decreases to prevent complete depletion of
cellular glycogen levels (7). The release of glycogen phosphorylase from glycogen partially accounts for the observed reversal in glycogenolysis (21). As glycogen breakdown diminishes
during continued contraction, the unfavorable conditions for
extracellular glucose uptake and utilization are dissipated.
Glucose transport into the muscle is increased, correlated
acutely with enhanced GLUT4 translocation to the plasma
membrane (24, 52), and, in trained individuals, upregulation of
GLUT4 mRNA and protein levels (20, 30). Several studies
indicate that muscular glycogen content prior to exercise also
affects the regulation of glucose transport (2, 12), although
others report no effect (1, 18). The differential effects on
glucose uptake seen in these studies could be due to differences
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G-6-P by hexokinase II and may enter glycolysis to produce
ATP or be converted by phosphoglucomutase to glucose
1-phosphate, which is further metabolized for glycogen synthesis. Additionally, insulin induces the dephosphorylation of
glycogen synthase via inhibition of upstream kinases and
activation of phosphatases (4). Thus insulin synergistically
stimulates glycogen synthesis in muscle by coordinately increasing intracellular concentrations of G-6-P and by dephosphorylation and activation of glycogen synthase.
In humans, skeletal muscle is the primary site for glucose
disposal, where up to 90% of a glucose load is converted to
glycogen (26). Skeletal muscle fibers differ in the amount of
glycogen they store and are classified as type I (fast switch) or
type II (slow twitch), based on their contractile speed and
metabolic properties. Type I fibers are responsible for slowspeed contractions and utilize oxidative pathways for ATP
production, whereas type II fibers are capable of rapid contractions and rely primarily on glycolytic pathways for energy
production. Type II fibers are further classified as type IIa and
type IIb; IIa fibers make use of both oxidative and glycolytic
pathways for energy production, whereas IIb rely primarily on
glycolysis. Thus type I fibers rely more on free fatty acid (FFA)
oxidation during contraction, whereas type II fibers are more
dependent on glucose uptake and glycolysis.
Muscle glycogen metabolism can be viewed simply as the
major site of insulin-stimulated glucose storage during times of
energetic abundance, and conversely, glycogen is broken down
to provide ATP to the working muscle during exercise. However, recent work in transgenic animals has challenged the
notion that muscle glycogen levels are essential for exercise, as
modulation of muscle glycogen stores did not impact on
exercise performance (45, 46). Furthermore, because glycogen
stores are finite, their depletion results in the switching of
energy utilization to extracellular glucose and FFA as exercise
continues (21). A number of factors may influence when and to
what extent glycogen and other fuels are utilized to maintain
ATP homeostasis, including the intensity of the exercise, the
dietary supply of carbohydrate or lipid, and the training status
of the individual (21). In this section, we consider energy
utilization in muscle before, during, and after a single bout of
exercise in order to demonstrate that glycogen metabolism is a
key regulator that integrates multiple energetic pathways.
Invited Review
NEW PERSPECTIVES ON THE ROLE OF GLYCOGEN METABOLISM
AJP-Endocrinol Metab • VOL
cluding increased oxidation of lipids, enhanced glucose transport, and several of the long-term adaptations associated with
exercise, such as upregulation of GLUT4 expression and increased mitochondrial content (6, 67). AMPK inactivates glycogen synthase (68), inhibiting glycogen synthesis in an effort
to shunt glucose delivery from storage to glycolysis.
Several lines of evidence suggest that changes in intracellular glycogen stores may regulate AMPK activation. First, an
inverse relation between high glycogen levels and AMPK
activity is logical and has been observed in rat and human
skeletal muscle (11, 68, 69). Conversely, low muscular glycogen levels at the onset of exercise enhanced AMPK activation
compared with high muscle glycogen controls (11, 68, 69).
Finally, the loss of glycogen stores in the muscle glycogen
synthase knockout mouse model correlated with a robust increase in AMPK phosphorylation state (44). AMPK has been
shown to directly bind glycogen (48), but because glycogen
addition did not affect AMPK activity in vitro (48), the significance of this interaction may be at the level of subcellular
localization of the kinase rather than a direct modulation of
enzymatic activity. Thus, during sustained exercise, as glycogen stores are exhausted, activation of AMPK enables muscle
fibers to switch over to energy utilization from extracellular
glucose (via increased GLUT4 translocation) and FFA oxidation (via inhibition of acetyl-CoA carboxylase, the rate-limiting enzyme for fatty acid synthesis).
Glycogen Metabolism During Recovery
Regulation of cellular responses to hormonal input. After a
strenuous bout of exercise, skeletal muscle is geared toward
increased energy uptake and metabolism to produce ATP and,
subsequently, repletion of glycogen stores. Two of the hallmark features of recovery from glycogen-depleting exercise
are the increased response and sensitivity to insulin, resulting
in enhanced GLUT4 translocation and glucose uptake (29), and
activation of glycogen synthase following refeeding. In exercised rats, glucose uptake was significantly greater in muscle
with low glycogen compared with high-glycogen muscle when
perfused at the same insulin concentration (12). Furthermore, it
was found that the rate of glycogen synthesis limited the rate of
glucose transport, as opposed to the availability of intracellular
substrate, indicating that transport itself was not limiting.
The elevation in glycogen synthase activity after glycogen
depletion may be regulated at the level of protein translocation.
As is the case with glycogen phosphorylase, glycogen synthase
binds directly to glycogen (39); yet basal and insulin-stimulated glycogen synthase activity is negatively correlated to
glycogen levels (3, 10, 42). Using a subcellular fractionation
technique, Nielsen et al. (42) demonstrated that, in glycogenreplete muscle from rat, glycogen synthase is primarily located
in a glycogen-rich membrane fraction, whereas in glycogendepleted muscle, glycogen synthase is found in a cytoskeleton
fraction. Furthermore, enzymatic activity was enhanced to a
greater extent in the low-glycogen than in the high-glycogen
rats in response to both insulin and muscle contraction, suggesting that glycogen stores dictated the subcellular localization of glycogen synthase, which in turn correlated with alterations in modulation of glycogen synthase activity (42). In a
recent study, Prats et al. (49) demonstrated the redistribution of
glycogen synthase in a phosphorylation-dependent manner to
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in the exercise intensity of the protocols. In the studies where
moderate-intensity exercise without fatigue is used, the observed increases in fat oxidation may compensate for the
reduction in muscle glycogenolysis in the low-starting-glycogen groups, such that glucose transport into the muscle was not
elevated. However, at higher intensities, when glycogen stores
are significantly depleted and lipid metabolism becomes limiting, an increase in glucose uptake would become necessary to
fuel continued muscular contraction.
Regulation of fuel partitioning by glycogen metabolism. In
addition to increasing glucose transport as glycogen stores
become depleted, skeletal muscle can also oxidize FFA to
produce ATP to preserve glycogen levels. Support for the role
of glycogen levels in regulating the switch in fuel utilization by
muscle during exercise comes from studies where glycogen
content was manipulated prior to exercise (1, 2, 9, 18, 65). In
each of these studies, increased muscle glycogen storage prior
to exercise led to increased carbohydrate oxidation during
exercise compared with controls. Conversely, decreasing muscle glycogen stores prior to exercise decreased carbohydrate
oxidation and increased lipid oxidation. Studies also found that
subjects with low glycogen prior to exercise displayed increased norepinephrine and FFA levels during exercise (65).
These findings are similar to observations made in McArdle’s
patients, who lack functional muscle glycogen phosphorylase
and cannot break down glycogen stores (33). Substantial evidence suggests that the inability to mobilize muscle glycogen
in McArdle’s patients can be directly communicated to the
central nervous system, in turn promoting lipid utilization (64,
65). Further evidence supporting a role for glycogen in the
regulation of substrate utilization during exercise comes from
a recently described transgenic mouse model that lacks the
muscle isoform of glycogen synthase and thus muscle glycogen stores (44). The authors observed no change in exercise
capacity compared with wild-type animals (46), but there was
an increase in type I fiber content in the gastrocnemius muscle,
indicative of a compensatory enhancement in lipid oxidative
capacity. So, although the exercise capacity of the transgenic
mice was unaffected by the loss of muscle glycogen stores, the
adaptive differentiation of oxidative type I fibers indicates that,
physiologically, muscle glycogen stores serve as an important
energy source during exercise.
Cumulatively, these observations reveal that the metabolic
changes resulting from glycogen depletion during exercise
promote the utilization of FFA for ATP generation. Furthermore, repeated or sustained loss of skeletal muscle glycogen,
through exercise training or in transgenic animals, results in
adaptive responses at the levels of gene transcription and
muscle fiber type differentiation to increase lipid oxidative
capacity. These adaptive changes in substrate utilization are
geared toward the preservation of finite muscle glycogen stores
(47).
Role of AMPK in fuel utilization. Recent work has suggested
that the enzyme AMP-activated protein kinase (AMPK) may
be the molecular link between alterations in glycogen levels
and the timing of the shift from carbohydrate to lipid metabolism during exercise (66). AMPK is a key regulator of cellular
energy metabolism due to its activation by increases in the
AMP/ATP ratio (17) and its effects on increasing energy
production and inhibition of energy storage, particularly as
glycogen. AMPK regulates numerous cellular processes, in-
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NEW PERSPECTIVES ON THE ROLE OF GLYCOGEN METABOLISM
LIVER GLYCOGEN
Overview
Although the majority of postprandial plasma glucose is
stored in skeletal muscle, hepatic glycogen makes up 10% of
total liver weight when fully replete, reflecting the importance
of glycogen metabolism in liver function. When plasma glucose levels rise after a meal, the liver clears glucose and stores
it as glycogen. Conversely, when blood glucose concentrations
fall during fasting or exercise, hepatic glycolytic rates drop due
to decreased levels of the key glycolytic regulator fructose-2,6biphosphatase. The liver produces glucose in response to an
increase in the circulating glucagon-to-insulin ratio via mobilization of hepatic glycogen stores and increased expression of
key gluconeogenic enzymes resulting in de novo synthesis of
glucose from 3-carbon precursors. Thus hepatic glycogen metabolism plays a central role in maintaining circulating blood
glucose levels around the physiological set point of 5 mM.
AJP-Endocrinol Metab • VOL
The differential functions for liver vs. muscle glycogen
metabolism are highlighted by the tissue-specific enzymatic
isoforms regulating glucose and glycogen metabolism. Muscle
and liver express different isoforms of glycogen synthase and
phosphorylase. Although their regulation by covalent modification is similar, each enzyme is differently sensitive to allosteric regulators, reflecting the metabolic demands of liver and
muscle. Additionally, the major glucose transporter in liver is
GLUT2, which, unlike GLUT4, is constitutively localized in
the plasma membrane and possesses a relatively low affinity
for glucose. Hepatocytes also express a specialized hexokinase,
termed glucokinase, which is not subject to feedback inhibition
by G-6-P. Similar to their function in the pancreatic ␤-cell, the
kinetic properties of GLUT2 and glucokinase together act as a
“glucose sensor” in which glucose uptake fluctuates in synchrony with changes in plasma glucose concentrations, enabling the regulated uptake and production of glucose as
needed.
Hepatic Glycogen Metabolism in the Fasted State: Two
Models of Hepatic Glucose Output
The liver provides glucose to the bloodstream during times
of fast or increased energy demand through gluconeogenesis
and the mobilization of intracellular glycogen stores. These
pathways have been considered separate entities, each providing the G-6-P that is dephosphorylated and secreted from
hepatocytes (Fig. 1A). However, recent work in several areas
has suggested that gluconeogenesis and glycogen metabolism
comprise a single interconnected metabolic circuit rather than
two discrete arms (Fig. 1B). These results suggest that, as in
muscle, hepatic glycogen metabolism occupies a key role in
the coordination of cellular responses with nutrient status under
a variety of physiological conditions.
Evidence for the interrelation of gluconeogenesis and glycogenolysis came from studies where human volunteers were
infused with gluconeogenic precursors under insulin and glucagon clamps. It was hypothesized that an increased supply of
gluconeogenic precursors would result in enhanced hepatic
glucose output. Instead, researchers found that hepatic glucose
production and plasma glucose concentrations were not significantly altered, suggesting that glycogen breakdown was
downregulated when gluconeogenic precursor supply was increased (23). Similarly, inhibition of gluconeogenesis did not
correct hyperglycemia in type 2 diabetic patients, again because glycogen breakdown was increased to maintain hepatic
glucose output (50). The results of these studies suggested the
hypothesis of “hepatic interregulation,” in which modulation of
one arm of glucose production would be compensated for by
changes in the other, thus maintaining hepatic glucose output at
the same level. However, in 1998, Martin et al. (34) demonstrated that glycogenolysis could be reduced in vivo in a mouse
model of diabetes by administration of a glycogen phosphorylase inhibitor, CP-91149. Unlike studies in which gluconeogenesis was inhibited, reduction of glycogenolysis resulted in
decreased hepatic glucose output and improvement of hyperglycemia (34). Fosgerau et al. (14) reported similar results
using another glycogen phosphorylase inhibitor, diaminobenzidine, in a canine model. These studies demonstrated that
changes in gluconeogenesis may result in alterations in glycogenolysis, but that the converse does not occur.
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an intracellular compartment where glycogen resynthesis occurred following depletion of glycogen stores (49). However,
the enzymatic mechanisms underlying alterations in glycogen
synthase subcellular distribution and degree of enzymatic activation remain poorly understood.
Glycogen supercompensation. The phenomenon of glycogen
supercompensation provides further evidence for the role of
glycogen levels in dictating muscle energy metabolism and
hormonal sensitivity. Glycogen supercompensation occurs following exercise and carbohydrate refeeding, when muscle
glycogen stores are replenished to levels above that of the
normal fed store. Glycogen supercompensation results in a
transient insulin resistance that is corrected upon return of
cellular glycogen levels to their physiological set point. Studies
have shown that both insulin-stimulated and contraction-induced translocation of GLUT4 to the plasma membrane were
reduced in glycogen-supercompensated muscle of rats, resulting in decreased glucose uptake (28). Furthermore, the reduction masked the adaptive increases in GLUT4 expression
resulting from exercise training (22, 27, 28).
Conversely, Garcia-Roves et al. (15) examined the effects of
preventing glycogen repletion on the postexercise GLUT4
increase. Rats were subjected to bouts of swimming exercise
and then either fasted or fed normal or carbohydrate-free chow.
In rats fed normal chow following exercise, elevated GLUT4
levels and insulin sensitivity returned to baseline levels within
42 h postexercise, after glycogen levels were restored. In
contrast, in the rats fed carbohydrate-free food to prevent
glycogen restoration, the increases in GLUT4 expression and
insulin sensitivity were retained for at least 66 h. At this point,
when fed carbohydrate-containing chow, the rats accumulated
glycogen to a similar extent as rats fed normal chow immediately following exercise. In parallel, GLUT4 protein levels
returned to baseline concentrations (15). Thus these data show
that the adaptations in order to increase glucose transport after
exercise will persist until glycogen levels are restored, underscoring the importance of glycogen stores in the physiological
responses of skeletal muscle. However, the molecular mechanisms by which alteration of glycogen levels, and presumably
modulation of metabolic fluxes into and out of glycogen stores,
are translated into adaptive changes in muscle gene expression
are not currently understood.
Invited Review
NEW PERSPECTIVES ON THE ROLE OF GLYCOGEN METABOLISM
Under the classical model (Fig. 1A), where gluconeogenesis
and glycogenolysis are considered two separate entities, the
mechanism for interregulation is unclear. The view of two
discrete pathways also makes it difficult to hypothesize why
alterations of gluconeogenesis affect glycogenolysis but not the
converse. Instead, the results of these studies suggest a model
in which gluconeogenesis and glycogen metabolism are a
continuous pathway (Fig. 1B). In this model, gluconeogenic
precursors are metabolized to G-6-P, which would then be
used for glycogen synthesis. Glycogenolysis would proceed,
and the pool of G-6-P arising from glycogen breakdown would
be dephosphorylated and extruded to the bloodstream. This
model helps predict multiple layers of regulation and explains
how inhibition of glycogen phosphorylase would result in
reduction of hepatic glucose production and plasma glucose
levels.
Hepatic Glycogen Metabolism in the Fed State: Glucose and
Gluconeogenic Precursor Disposal in the Liver
Further evidence in support of a continuous pathway for
gluconeogenesis and glycogen metabolism comes from studies
of nutrient disposal in the fed state. Glycogen may be synthesized from extracellular glucose (direct pathway) or from
glucose derived from gluconeogenic substrates (indirect pathway). Although studies dating back to the 1930s showed that
AJP-Endocrinol Metab • VOL
administration of labeled glucose through a single bolus resulted in the majority of hepatic glycogen being synthesized
via the direct pathway, a growing body of evidence in the
1970s indicated that G-6-P from glucose was inefficiently used
as the immediate precursor of liver glycogen. This apparent
inconsistency was termed the “glucose paradox” (reviewed in
Ref. 36).
To address the different outcomes of these studies, Newgard
et al. (41) compared results from different experimental designs and evaluated which more closely approximated physiological conditions. In the older experiments, administration of
the large intragastric bolus resulted in a nonphysiological
hyperglycemia that presumably allowed glucose to enter the
hepatocyte via GLUT2 and glucokinase at higher than normal
rates. In the later studies, labeled glucose was infused intragastrically at a constant rate to achieve circulating glucose
levels similar to those in rats refed following a fast. Under
these conditions, less than one-third of liver glycogen was
synthesized via the direct pathway. Additional evidence came
from studies in humans by Shulman and colleagues (8, 58, 60),
which indicated that gluconeogenic compounds are the major
carbon source for hepatic glycogen. Finally, O’Doherty et al.
(43) demonstrated that hepatic glycogen synthesis was increased in rats overexpressing PTG, a glycogen-targeting subunit of protein phosphatase-1. Although glucose incorporation
into glycogen (direct pathway) was increased, the major mechanism for enhancement of glycogen synthesis was through
increased incorporation of gluconeogenic precursors into glycogen (indirect pathway).
It is now accepted that the major source of glucose for
hepatic glycogen stores is derived from 3-carbon compounds such as lactate, glycerol, and alanine and other
gluconeogenic amino acids rather than directly from extracellular glucose. The synthesis of these compounds into
glycogen comprises a system where gluconeogenesis and
glycogen metabolism form a highly interconnected pathway,
even in the fed state (Fig. 1B). Although these data represent
a fairly recent paradigm shift, studies dating from the 1940s
demonstrated that labeled carbon from gluconeogenic compounds could be tracked to glycogen (61, 70). To confirm
these initial observations, the laboratory of Radziuk administered [14C]- and [3H]glucose tracers orally or intravenously to human subjects to quantitate glycogen synthesis
from glucose compared with gluconeogenic precursors.
Data demonstrated 150% more deposition of labeled carbon
from gluconeogenic precursors over that from glucose (51).
Studies also suggest that, in addition to providing substrate
for glycogen synthesis, gluconeogenic compounds help regulate glycogen metabolism (reviewed in Ref. 71). For example, several groups demonstrated that incubation of isolated hepatocytes with gluconeogenic amino acids resulted
in increased activation of glycogen synthase (37, 40). Studies demonstrating feed-forward regulation of gluconeogenic
compounds on glycogen synthesis provide additional evidence for the proposed model of a continuous metabolic
pathway.
Glycogen Cycling as a Key Regulator of Hepatic Function
Under the proposed model, the funneling of gluconeogenic
precursors and glucose through glycogen appears to generate a
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Fig. 1. Schematic models of hepatic glucose output (HGO). A: classical model
of de novo glucose synthesis (gluconeogenesis) and glycogen breakdown
(glycogenolysis) independently contributing to HGO. B: diagram of cycling of
newly synthesized glucose through glycogen prior to secretion from hepatocytes. G6Pase, glucose-6-phosphatase; G-6-P, glucose 6-phosphate; G-1-P,
glucose 1-phosphate; UDPG, UDP-glucose; GP, glycogen phosphorylase; GS,
glycogen synthase; PEPCK, phosphoenolpyruvate carboxykinase.
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through glycogen is not constitutive but adapts to the metabolic
requirements of hepatocytes.
A Model To Predict Glucose Flux Through Glycogen
The work on the interplay among hepatic glucose output,
gluconeogenesis, and glycogen metabolism leads us to propose
the model in Fig. 2, where rates of glucose cycling through
glycogen are dependent on intracellular glycogen stores and
extracellular signals. Three to four hours after a meal, as blood
glucose levels start to fall, insulin secretion drops, leading to a
decrease in the insulin/glucagon ratio and an increase in cytosolic cAMP concentration ([cAMP]c; Fig. 2, x-axis, left). An
initial burst of glycogen mobilization occurs in order to provide
glucose for release into the bloodstream (Fig. 2, top left), and
glucose cycling would be low due to high glycogen stores and
low hepatic glucose production. As glycogen stores fall, the
gluconeogenic component of hepatic glucose output would
increase (Fig. 2, bottom left) as lactate and pyruvate are
synthesized into G-6-P. Glucose cycling through glycogen
would commence, in part to preserve glycogen stores. Potentially, the linkage of these pathways into one circuit would also
provide a metabolic mechanism for the liver to keep track of
total glucose production and enable the liver to balance energy
demand by the body with its capacity to provide glucose from
gluconeogenesis and glycogenolysis. When the fast is broken
by a meal, insulin and glucose levels would rise, [cAMP]c
would fall (Fig. 2, x-axis, right), and glucose storage as
glycogen would be favored. Initially, resynthesis of glycogen
chains would be strongly favored, preventing glucose cycling
(Fig. 2, bottom right). However, as glycogen levels grew,
glycogen degradation and glucose cycling would commence to
prevent overaccumulation of glycogen (Fig. 2, top right).
Conclusion
Classical studies of glycogen metabolism have figured
prominently in the progress of biochemistry and signal transduction due to seminal discoveries in the areas of protein
phosphorylation, intracellular second messengers, and phosphatase-targeting subunits. Following in that tradition, current
Fig. 2. Model predicting glycogen cycling in hepatocytes. Rate of glucose flux
through glycogen is dependent on intracellular levels of glycogen and extracellular levels of hormones and glucose. See text for details. [cAMP]c,
cytosolic cAMP concentration levels.
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futile cycle, in which glycogen synthesis and breakdown occur
simultaneously. However, numerous studies demonstrate that
glycogen cycling is a sensitively regulated circuit that allows
the hepatocyte to be highly responsive to multiple stimuli. In a
study of humans fasted for 60 h, as expected, hepatic glycogen
levels were low, and gluconeogenesis accounted for the vast
majority of hepatic glucose output (19). Subjects were then
infused for 10 h with [13C]glycerol, followed by a pulse of
glucagon, and glycogen stores were allowed to recover. Seventy-eight percent of the postglucagon glycogen was derived
from the gluconeogenic precursor (19). These data indicate that
glucose cycles through glycogen even when cellular stores are
extremely low, which may prevent the complete depletion of
cellular glycogen stores during prolonged fast.
Pharmacological inhibitors of glycogen phosphorylase have
provided another critical tool to study the contribution of
aberrant hepatic glucose output to hyperglycemia in diabetic
patients, but also glucose cycling through glycogen (34). In
primary hepatocytes from fasted rat, no net glycogen synthesis
was detectable (35). However, both glycogen synthase and
phosphorylase were in an activated state, and addition of a
phosphorylase inhibitor promoted glycogen synthesis. Additionally, in a study of fasted dogs, addition of phosphorylase
inhibitor resulted in accumulation of glucose derived from
gluconeogenic precursors into glycogen (57). These studies
provided further support to the notion that newly synthesized
glucose cycles through glycogen routinely in the fasted state.
Finally, treatment of primary hepatocytes with a phosphorylase
inhibitor blocked conversion of [14C]lactate to [14C]glucose by
over 50% and resulted in a threefold increase in [14C]lactate
incorporation into glycogen (34). This unexpected result suggests that, not only does glucose derived from gluconeogenesis
cycle through glycogen, this step may be required for efficient
dephosphorylation and secretion of newly synthesized glucose
into the bloodstream (Fig. 1B).
Glucose cycling through glycogen also occurs in the fed
state. By use of perfused rat livers under conditions favoring
glycogen synthesis, tissues were pulsed with [13C]glucose and
chased with [12C]glucose or [13C]glucose as a control (59). In
the continued presence of [13C]glucose, there was an increase
in nuclear magnetic resonance peak height in glycogen over
time. However, during a [12C]glucose chase perfusion, there
was a rapid decline in 13C label content in glycogen, implying
continual turnover in liver glycogen even under conditions
favoring glycogen accumulation, since the 13C label was
briskly being lost from glycogen during the chase with
[12C]glucose (59). These data also indicate a role for glycogen
turnover to prevent overaccumulation of hepatic glycogen
stores, which may be physiologically important due to expression of GLUT2 and glucokinase isoforms resulting in a continual flux of G-6-P into hepatocytes during prolonged hyperglycemia.
Finally, several studies have provided evidence that under
certain experimental conditions glucose cycling through glycogen is negligible. In cells that had high glycogen stores and
then were treated with a high ratio of glucagon to insulin,
glycogen turnover was undetectable due to low synthetic rates
(16). Additionally, in glycogen-depleted cells that were then
stimulated with high glucose and insulin, glycogen degradation
was markedly suppressed, enabling maximal rates of glycogen
repletion (14). These results indicate that glucose cycling
Invited Review
NEW PERSPECTIVES ON THE ROLE OF GLYCOGEN METABOLISM
research has highlighted additional roles for glycogen in cellular physiology beyond its simple function as an energy depot.
In muscle, where a variety of energy sources are utilized during
contraction, rates of glycogen metabolism help orchestrate the
changing integration of cellular fuel utilization pathways during exercise. In liver, glycogen turnover links gluconeogenesis
into a single circuit to coordinately modulate hepatic glucose
output. These findings also generate new questions regarding
glycogen metabolic function and regulation, ensuring that the
study of glycogen will continue to be a dynamic, expanding
field.
GRANTS
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