Studing of biosynthesis and catabolism of glycogen. Regulation of glycogen metabolism. Biosynthesis of glucose – gluconeogenesis. Hormonal regulation and pathologies of carbohydrate metabolism.

Synthesis of glycogen. Regulation of glycogen synthese activity

Glycogen Synthesis

Synthesis of glycogen from glucose is carried out the enzyme glycogen synthase. This enzyme utilizes UDP-glucose as one substrate and the non-reducing end of glycogen as another. The activation of glucose to be used for glycogen synthesis is carried out by the enzyme UDP-glucose pyrophosphorylase. This enzyme exchanges the phosphate on C-1 of glucose-1-phosphate for UDP. The energy of the phospho-glycosyl bond of UDP-glucose is utilized by glycogen synthase to catalyze the incorporation of glucose into glycogen. UDP is subsequently released from the enzyme. The α-1,6 branches in glucose are produced by amylo-(1,4 - 1,6)-transglycosylase, also termed the branching enzyme. This enzyme transfers a terminal fragment of 6-7 glucose residues (from a polymer at least 11 glucose residues long) to an internal glucose residue at the C-6 hydroxyl position.

Addition of Glucose to Glycogen

For de novo glycogen synthesis to proceed the first glucose residue is attached to a protein known as glycogenin. Glycogenin has the unusual property of catalyzing its own glycosylation, attaching C-1 of a UDP-glucose to a tyrosine residue on the enzyme. The attached glucose then serves as the primer required by glycogen synthase to attach additional glucose molecules via the mechanism described above.

Glycogen Branching Activity

Hormonal regulation of glycogen metabolism

Regulation of Glycogen Synthesis

Glycogen synthase ia a tetrameric enzyme consisting of 4 identical subunits. The activity of glycogen synthase is regulated by phosphorylation of serine residues in the subunit proteins. Phosphorylation of glycogen synthase reduces its activity towards UDP-glucose. When in the non-phosphorylated state, glycogen synthase does not require glucose-6-phosphate as an allosteric activator, when phosphorylated it does. The two forms of glycogen synthase are identifed by the same nomenclature as used for glycogen phosphorylase. The unphosphorylated and most active form is synthase-a and the phosphorylated glucose-6-phosphate-dependent form is synthase-b shift.

Pathways involved in the regulation of glycogen synthase. See the text for details of the regulatory mechanisms. PKA is cAMP-dependent protein kinase. PPI-1 is phosphoprotein phosphatase-1 inhibitor. Whether a factor has positive (+ve) or negative (-ve) effects on any enzyme is indicated. Briefly, glycogen synthase a is phosphorylated, and rendered much less active and requires glucose-6-phosphate to have any activity at all. Phosphorylation of glycogen synthase is accomplished by several different enzymes. The most important is synthase-phosphorylase kinase the same enzyme responsible for phosphorylation (and activation) of glycogen phosphorylase. PKA (itself activated through receptor mediated mechanisms) also phosphorylates glycogen synthase directly. The effects of PKA on PPI-1 are the same as those described above for the regulation of glycogen phosphorylase. The other enzymes shown to directly phosphorylate glycogen synthase are protein kinase C (PKC), calmodulin-dependent protein kinase, glycogen synthase kinase-3 (GSK-3) and two forms of casein kinase (CK-I and CK-II). The enzyme PKC is activated by Ca²⁺ ions and phospholipids, primarily diacylglycerol, DAG. DAG is formed by receptor-mediated hydrolysis of membrane phosphatidylinositol bisphosphate (PIP₂).

Phosphorylation of synthase occurs primarily in response to hormonal activation of PKA. One of the major kinases active on synthase is synthase-phosphorylase kinase; the same enzyme that phosphorylates glycogen phosphorylase. However, at least 5 additional enzymes have been identified that phosphorylate glycogen synthase directly. One of of these glycogen synthase phosphorylating enzymes is PKA itself. One important glycogen synthase phosphorylating enzyme is active independently of increases in cAMP levels. This enzyme is glycogen synthase kinase 3 (GSK-3). Each phosphorylation event occurs at distinct serine residues which can result in a progressively increased state of synthase phosphorylation.

Glycogen synthase activity can also be affected by epinephrine binding to α-adrenergic receptors through a pathway like that described above for regulation of glycogen phosphorylase.

Pathways involved in the regulation of glycogen synthase by epinephrine activation of α-adrenergic receptors. See the text for details of the regulatory mechanisms. PKC is protein kinase C. PLC-γ is phospholipase C-γ. The substrate for PLC-γ is phosphatidylinositol-4,5-bisphosphate (PIP₂) and the products are IP₃ and DAG.

When α-adrenergic receptors are stimulated there is an increase in the activity of PLC-γ with a resultant increase in PIP₂ hydrolysis. The products of PIP₂ hydrolysis are DAG and IP₃. As described above for glycogen phoshorylase, DAG and the Ca²⁺ ions released by IP₃ activate PKC which phosphorylates and inactivates glycogen synthase. Additional responses of calcium are the activation of calmodulin-dependent protein kinase (calmodulin is a component of many enzymes that are responsive to Ca²⁺) which also phosphorytes glycogen synthase.

The effects of these phosphorylations leads to:

1. Decreased affinity of synthase for UDP-glucose.

2. Decreased affinity of synthase for glucose-6-phosphate.

3. Increased affinity of synthase for ATP and P_i.

Reconversion of synthase-b to synthase-a requires dephosphorylation. This is carried out predominately by protein phosphatase-1 (PP-1) the same phosphatase involved in dephosphorylation of phosphorylase.

The activity of PP-1 is also affected by insulin. The pancreatic hormone exerts an opposing effect to that of glucagon and epinephrine. This should appear obvious since the role of insulin is to increase the uptake of glucose from the blood.

Genetic desorders of glycogen metabolism.

Since glycogen molecules can become enormously large, an inability to degrade glycogen can cause cells to become pathologically engorged; it can also lead to the functional loss of glycogen as a source of cell energy and as a blood glucose buffer. Although glycogen storage diseases are quite rare, their effects can be most dramatic. The debilitating effect of many glycogen storage diseases depends on the severity of the mutation causing the deficiency. In addition, although the glycogen storage diseases are attributed to specific enzyme deficiencies, other events can cause the same characteristic symptoms. For example, Type I glycogen storage disease (von Gierke disease) is attributed to lack of glucose-6-phosphatase. However, this enzyme is localized on the cisternal surface of the endoplasmic reticulum (ER); in order to gain access to the phosphatase, glucose-6-phosphate must pass through a specific translocase in the ER membrane. Mutation of either the phosphatase or the translocase makes transfer of liver glycogen to the blood a very limited process. Thus, mutation of either gene leads to symptoms associated with von Gierke disease, which occurs at a rate of about 1 in 200,000 people.

Several glycogenoses are the result of deficiencies in enzymes of glycolysis whose symptoms and signs are similar to those seen in McArdle disease (type V GSD). These include deficiencies in muscle phosphoglycerate kinase and muscle pyruvate kinase as well as deficiencies in fructose 1,6-bisphosphatase, lactate dehydrogenase and phosphoglycerate mutase.

Table of Glycogen Storage Diseases

Type: Name	Enzyme Affected	Primary Organ	Manifestations
GSD0a	glycogen synthase-2	liver	hypoglycemia, early death, hyperketonia
von Gierke GSD1a	glucose-6-phosphatase	liver	hepatomegaly, kidney failure, thrombocyte dysfunction
GSD1b	microsomal glucose-6-phosphate translocase	liver	like Ia, also neutropenia, bacterial infections
GSD1c	microsomal Pi transporter	liver	like Ia
Pompe GSD2	lysosomal acid α-glucosidase also called acid maltase	skeletal and cardiac muscle	infantile form = death by 2 juvenile form = myopathy adult form = muscular dystrophy-like
Cori or Forbes GSD3	liver and muscle debranching enzyme	liver, skeletal and cardiac muscle	infant hepatomegaly, myopathy
Andersen GSD4	branching enzyme	liver, muscle	hepatosplenomegaly, cirrhosis
McArdle GSD5	muscle phosphorylase	skeletal muscle	excercise-induced cramps and pain, myoglobinuria
Hers GSD6	liver phosphorylase	liver	hepatomegaly, mild hypoglycemia, hyperlipidemia and ketosis, improvement with age
Tarui GSD7	muscle PFK-1	muscle, RBC's	like V, also hemolytic anemia
GSD9a GSD9b	phosphorylase kinase β-subunit of PK	liver, leukocytes, muscle	like VI
Fanconi-Bickel hepatorenal glycogenosis	glucose transporter-2 (GLUT-2)	liver	failure to thrive, hepatomegaly, rickets, proximal renal tubular dysfunction

Enzymes of Type I Glycogen Storage Diseases

The mechanism by which free glucose is released from glucose-6-phosphate involves several different steps. Glucose-6-phosphate must first be transported from the cytosol where it is generated either through phosphorylation of free glucose or from gluconeogenesis, into the lumen of the endoplasmic reticulum, ER. Inside the ER the phosphate is removed through the action of ER localized glucose 6-phosphatase. The free glucose must then be transported back to the cytosol as well as the released inorganic phosphate, P_i. Defects in the process of glucose release from glucose-6-phosphate result in elevations in cytosolic glucose-6-phosphate which then leads to increases in incorporation into glycogen and subsequent excessive storage.

Introduction to Pompe Disease

Glycogen storage disease type II (GSDII) is also known as Pompe disease or acid maltase deficiency (AMD). This disease was originally referred to as Pompe disease since J-C Pompe (published in 1932) made the important observation of a massive accumulation of glycogen within the vacuoles of all tissues in a 7-month-old female who died suddenly from idiopathic hypertrophy of the heart. Additional clinical manifestations associated with idiopathic cardiomegaly associated with storage of glycogen were defined over the next several decades. These symptoms included hepatomegaly, marked hypotonia, muscular weakness and death before 1 year of age. Through the investigations carried out by the Cori's (G. Cori and C. Cori) this disease was classified as glycogen storage disease type II. GSDII is the most severe of all of the glycogen storage diseases. The excess storage of glycogen in the vacuoles is the consequence of defects in the lysosomal hydrolase, acid α-glucosidase which removes glucose residues from glycogen in the lysosomes.

The acid α-glucosidase gene (designated GAA) resides on chromosome 17q25 spanning 20 kb and composed of 20 exons. GSDII has been shown to be caused by missense, nonsense and splice-site mutations, partial deletions and insertions. Some mutations are specific to certain ethnic groups. There are three common allelic forms of acid α-glucosidase that segregate in the general population. These forms are designated GAA1, GAA2 and GAA4. The normal function of acid α-glucosidase is to hydrolyze both α-1,4- and α-1,6-glucosidic linkages at acid pH. The activity of the enzyme leads to the complete hydrolysis of glycogen which is its natural substrate. As would be expected from this activity, deficiency in acid α-glucosidase leads to the accumulation of structurally normal glycogen in numerous tissues, most notably in cardiac and skeletal muscle.

Clinical Features of Pompe Disease

The clinical presentation of GSDII encompasses a wide range of phenotypes but all include various degrees of cardiomegaly. The different phenotypes can be classified dependent upon age of onset, extent of organ involvement and the rate of progression to death. The infantile-onset form is the most severe and was the phenotype described by Pompe. The other extreme of this disorder is a slowly progressing adult onset proximal myopathic disease. The late onset disease usually presents as late as the second to sixth decade of life and usually only involves the skeletal muscles. There is also a heterogeneous group of GSDII disorders that are classified generally by onset after early infancy and called the juvenile or childhood form. In addition to classical symptoms that can lead to a diagnosis of GSDII, analysis for the level and activity of acid α-glucosidase in muscle biopsies is used for confirmation.

The infantile onset form of GSDII presents in the first few months of life. Symptoms include marked cardiomegaly, striking hypotonia (leading to the designation of "floppy baby syndrome") and rapid progressive muscle weakness. Patients will usually exhibit difficulty with feeding and have respiratory problems that are frequently complicated by pulmonary infection. The prominent cardiomegaly that can be seen on chest X-ray is normally the first indication leading to a preliminary diagnosis of GSDII. There is currently no cure for GSDII. In 2006 the US FDA approved the use of Myozyme® (alglucosidase alpha) as an enzyme replacement therapy (ERT) for treatment of  infantile-onset Pompe disease. Supportive therapy with attention to treatment of respiratory function can impact the course of the disease in the late-onset form.

Introduction to Andersen Disease

Glycogen storage disease type IV (GSDIV) is also known as Andersen disease or amylopectinosis. This disease was originally described by D. Andersen in 1956, hence the association of his name with the disease. The disease was seen in a patient exhibiting progressive hepatosplenomegaly along with the storage of an abnormal glycogen that had poor solubility in the liver. The abnormal glycogen had few branch points with long outer chains containing more α-1,4-linked glucose than the normal polysaccharide which contains both α-1,4- and α-1,6-glycosidic linkages. This resultant structure was similar to that of amylopectin (the structure of starch in plants), thus the associated name of amylopecintosis. Because of the structure of the glycogen it was suspected that there was a deficiency in glycogen branching enzyme activity. This was indeed found to be true in 1966.

Glycogen branching enzyme (gene symbol GBE1), also called amylo-(1,4 to 1,6) transglycosylase, is a monomeric protein. The α-1,6 branches in glucose are produced by this enzyme through a process involving the transfer of a terminal fragment of 6-7 glucose residues (from a polymer at least 11 glucose residues long) to an internal glucose residue at the C-6 hydroxyl position. The GBE1 gene is located on chromosome 3p12 and encodes an mRNA of 2,106 bp containing a coding region of 702 amino acids.

Reaction catalyzed by glycogen branching enzyme

Clinical Features of Andersen Disease

Analysis of mutants in the branching enzyme gene demonstrated that both hepatic and neuromuscular forms of GSDIV were the result of defects in the same gene. The clinical presentation of GSDIV symptoms usually occurs in the first few months of life and is characterized by hepatosplenomegaly and failure to thrive. The disease progresses liver cirrhosis, portal vein hypertension, esophageal varices and ascites. Death will usually ensue by 5 years of age. Because of the similarities in symptoms between GSDIV and other causes of cirrhosis in infancy it is necessary to carry out a biopsy of the liver and examine for the presence of the associated abnormal glycogen. In addition, assay for branching enzyme deficiency in muscle, leukocytes, erythrocytes, or fibroblasts can be carried out to determine the exact defect resulting in the hepatomegaly. Treatment of GSDIV normally involves maintenance of normal blood glucose along with adequate nutrient intake both of which will improve liver function and muscle strength. In cases of progressive liver failure, transplant may be the only effective option.

Introduction to McArdle Disease

Glycogen storage disease type V (GSDV) is also known as McArdle disease. This disease was originally described by B. McArdle in 1951, hence the association of his name with the disease. The disease was seen in a 30-year-old patient who was suffering from muscle weakness, pain and stiffness following slight exercise. It was observed that blood lactate fell in this patient during exercise instead of the normal rise that would be seen. This indicated the patient had a defect in the ability to convert muscle glycogen to glucose and ultimately lactate. The identification that the deficiency causing these symptoms was the result of a muscle phosphorylase defect was not made until 1959. Glycogen phosphorylase (most commonly just called phosphorylase) exists in multiple tissue-specific isoforms. The muscle, brain and liver forms are encoded by separate genes. The muscle form is the only isozyme found expressed in mature muscle.

The gene for muscle phosphorylase (symbol PYGM) is located on chromosome 11q13-qter and is composed of 20 exons encoding a 842 amino acid protein. The muscle isoform represents about 50% of total phosphorylase in cardiac muscle, 30% of the brain phosphorylase and is completely absent from the liver. At least 14 different mutations have been identified as causing GSDV The most commonly encountered mutation (75% of patients) is a nonsense mutation that changes a C to a T in exon 1 converting an arginine codon to a stop codon (R49X).

Reaction catalyzed by glycogen phosphorylase

Clinical Features of McArdle Disease

The clinical presentation of GSDV is usually seen in young adulthood and is characterized by exercise intolerance and muscle cramps following slight exercise. Attacks of myoglobinuria frequently accompany the muscle symptoms of GSDV. About half of GSDV patients will exhibit burgundy colored urine after exercise. Diagnosis of muscle glycogenoses such as GSDV can be made by the observation of a lack of increased blood lactate upon ischemic exercise testing. In addition, there will be an associated large increase in blood ammonia levels. In order to distinguish GSDV from other muscle defects along the pathway from glycogen to glucose to lactate, an enzymatic evaluation of muscle phosphorylase must be done. Also, molecular analysis for known mutations in the muscle phosphorylase gene can be accomplished using DNA extracted from leukocytes.

Glycogenolysis: difference from glycolysis.

Energetic value; regulation of glycogen phosphorylase activity

GLYCOGENOLYSIS IS NOT THE REVERSE OF GLYCOGENESIS BUT IS A SEPARATE PATHWAY

Glycogen phosphorylase catalyzes the rate-limiting step in glycogenolysis by promoting the phosphorylytic cleavage by inorganic phosphate (phosphorylysis; cf hychdrolysis) of the 1→4 linkages of glycogen to yield glucose 1-phosphate. The terminal glucosyl residues from the outermost chains of the glycogen molecule are removed sequentially until approximately four glucose residues remain on either side of a 1→6 branch

Another enzyme (_-[1v4]v_-[1v4] glucan transferase) transfers a trisaccharide unit from one branch to the other, exposing the 1→6 branch point. Hydrolysis of the 1→6 linkages requires the debranching

enzyme. Further phosphorylase action can then proceed. The combined action of phosphorylase and these other enzymes leads to the complete breakdown

of glycogen. The reaction catalyzed by phosphoglucomutase is reversible, so that glucose 6-phosphate can be formed from glucose 1-phosphate. In liver (and kidney), but not in muscle, there is a specific enzyme, glucose-6-phosphatase, that hydrolyzes glucose 6-phosphate, yielding glucose that is exported, leading to an increase in the blood glucose concentration.

CYCLIC AMP INTEGRATES THE REGULATION OF GLYCOGENOLYSIS and GLYCOGENESIS

The principal enzymes controlling glycogen metabolism— glycogen phosphorylase and glycogen synthase— are regulated by allosteric mechanisms and covalent modifications due to reversible phosphorylation and dephosphorylation of enzyme protein in response to hormone action.

Cyclic AMP (cAMP) is formed from ATP by adenylyl cyclase at the inner surface of cell membranes and acts as an intracellular second messenger

in response to hormones such as epinephrine, norepinephrine, and glucagon. cAMP is hydrolyzed by phosphodiesterase, so terminating hormone action. In

liver, insulin increases the activity of phosphodiesterase.

Phosphorylase Differs Between Liver and Muscle

In liver, one of the serine hydroxyl groups of active phosphorylase a is phosphorylated. It is inactivated by hydrolytic removal of the phosphate by protein phosphatase- 1 to form phosphorylase b. Reactivation requires

rephosphorylation catalyzed by phosphorylase kinase.

Muscle phosphorylase is distinct from that of liver. It is a dimer, each monomer containing 1 mol of pyridoxal phosphate (vitamin B6). It is present in two forms: phosphorylase a, which is phosphorylated and active in either the presence or absence of 5′-AMP (its allosteric modifier); and phosphorylase b, which is dephosphorylated and active only in the presence of 5′-AMP. This occurs

during exercise when the level of 5′-AMP rises, providing, by this mechanism, fuel for the muscle. Phosphorylase a is the normal physiologically active form of the enzyme. cAMP Activates Muscle Phosphorylase

Phosphorylase in muscle is activated in response to epinephrine acting via cAMP. Increasing the concentration of cAMP activates cAMP-dependent protein kinase, which catalyzes the phosphorylation by ATP of inactive phosphorylase kinase b to active phosphorylase kinase a, which in turn, by means of a further phosphorylation, activates phosphorylase b to phosphorylase a.

Ca²⁺ Synchronizes the Activation of Phosphorylase With Muscle Contraction

Glycogenolysis increases in muscle several hundred-fold immediately after the onset of contraction. This involves the rapid activation of phosphorylase by activation of phosphorylase kinase by Ca^2+, the same signal as that which initiates contraction in response to nerve stimulation. Muscle phosphorylase kinase has four

types of subunits—α, β, γ, and δ—in a structure represented as (αβγδ)4. The α and β subunits contain serine residues that are phosphorylated by cAMP-dependent

protein kinase. The δ subunit binds four Ca²⁺ and is identical to the Ca^2+-binding protein calmodulin. The binding of Ca²⁺ activates the catalytic site of the γ subunit while the molecule remains in the dephosphorylated b configuration. However, the phosphorylated a form is only fully activated in the presence of Ca^2+. A second molecule of calmodulin, or TpC (the structurally similar Ca^2+-binding protein in muscle), can interact with phosphorylase kinase, causing further activation. Thus, activation of muscle contraction and glycogenolysis are carried out by the same Ca^2+-binding protein, ensuring their synchronization.

Glycogenolysis in Liver Can Be cAMP-Independent

In addition to the action of glucagon in causing formation of cAMP and activation of phosphorylase in liver 1-adrenergic receptors mediate stimulation of glycogenolysis by epinephrine and norepinephrine. This involves a cAMP-independent mobilization of Ca²⁺ from mitochondria into the cytosol, followed by the stimulation of a Ca^2+/calmodulin-sensitive phosphorylase kinase. cAMP-independent glycogenolysis is also caused by vasopressin, oxytocin, and angiotensin II acting through calcium or the phosphatidylinositol bisphosphate

pathway.

Protein Phosphatase-1 Inactivates Phosphorylase

Both phosphorylase a and phosphorylase kinase a are dephosphorylated and inactivated by protein phosphatase- 1. Protein phosphatase-1 is inhibited by a

protein, inhibitor-1, which is active only after it has been phosphorylated by cAMP-dependent protein kinase. Thus, cAMP controls both the activation and inactivation of phosphorylase. Insulin reinforces this effect by inhibiting the activation of phosphorylase b. It does this indirectly by increasing

uptake of glucose, leading to increased formation of glucose 6-phosphate, which is an inhibitor of phosphorylase kinase.

Glycogen Synthase and Phosphorylase Activity Are Reciprocally Regulated

Like phosphorylase, glycogen synthase exists in either a phosphorylated or nonphosphorylated state. However, unlike phosphorylase, the active form is dephosphorylated (glycogen synthase a) and may be inactivated to residues by no fewer than six different protein kinases. Two of the protein kinases are Ca²⁺calmodulindependent (one of these is phosphorylase kinase). Another kinase is cAMP-dependent protein kinase, which allows cAMP-mediated hormonal action to inhibit glycogen synthesis synchronously with the activation of glycogenolysis. Insulin also promotes glycogenesis in muscle at the same time as inhibiting glycogenolysis by raising glucose 6-phosphate concentrations, which stimulates the dephosphorylation and activation of glycogen synthase. Dephosphorylation of glycogen synthase b is carried out by protein phosphatase-1, which is under the control of cAMP-dependent protein kinase.

Diagnostic significance of the glucose measurement in urine: types of glucosouria

The laboratory diagnosis of diabetes depends on finding glucose in the urine (glucosuria), however the appearance of glucosuria may result from a variety of causes. In diabetes mellitus, glucose appears in the urine because a hyperglycemia condition exists in the blood. Therefore, the glucose concentration should also be measured in the blood. Glucosuria can occur temporarily from emotional stress or pain, hyperthyroidism, alimentary hyperglycemia, and meningitis.

Apparently, the kidney acts as a safety valve against the excessive accumulation of glucose in the blood. If the glucose level becomes too high, then the renal threshold in the kidneys may be exceeded. The renal threshold is a concentration level above which all glucose is not reabsorbed in the blood, but the excess above the threshold concentration remains in the urine. A threshold is analogous to a dam and only when the water level becomes too high, does the water spill over the dam. Many other substances in the blood have their own threshold levels and when the threshold is exceeded, the substance appears in the urine.

In renal diabetes, the threshold is abnormally low and glucose appears in the urine at a much lower concentration than normal. The relationships between glucosuria and the renal threshold are illustrated in the diagram on the left.

Another relatively easy laboratory test can be made for ketone bodies in the urine. The condition is known as acetonuria from the acetone present. Ketones bodies result in diabetes mellitus for the very same reasons as given for starvation. Ketone bodies are not normally found in urine nor are they present with the other types of diabetes listed. Ketone bodies are present in various amounts depending upon the severity of the diabetes mellitus.

ІІ Biosynthesis of glucose – gluconeogenesis.

Hormonal regulation and pathologies of carbohydrate metabolism.

Gluconeogenesis. Biological role of this process.

Gluconeogenesis is the biosynthesis of new glucose, (i.e. not glucose from glycogen). The production of glucose from other metabolites is necessary for use as a fuel source by the brain, testes, erythrocytes and kidney medulla since glucose is the sole energy source for these organs. During starvation, however, the brain can derive energy from ketone bodies which are converted to acetyl-CoA

Synthesis of glucose from three and four carbon precursors is essentially a reversal of glycolysis. The relevant features of the pathway of gluconeogenesis are diagrammed below.

The relevant reactions of gluconeogenesis are depicted. The enzymes of the 3 bypass steps are indicated in green along with phosphoglycerate kinase. This latter enzyme is included since when functioning in the gluconeogenic direction the reaction consumes energy. Gluconeogenesis from 2 moles of pyruvate to 2 moles of 1,3-bisphosphoglycerate consumes 6 moles of ATP. This makes the process of gluconeogenesis very costly from an energy standpoint considering that glycolysis to 2 moles of pyruvate only yields 2 moles of ATP. Note that several steps are required in going from 2 moles of 1,3-bisphosphoglycerate to 1 mole of fructose-1,6-bisphosphate. First there is a reversal of the glyceraldehyde-3-phosphate dehydrogenase reaction which requires a supply of NADH. When lactate is the gluconeogenic substrate the NADH is supplied by the lactate dehydrogenase reaction, and it is supplied by the malate dehydrogenase reaction when pyruvate is the substrate. Secondly, 1 mole of glyceraldehyde-3-phosphate must be isomerized to DHAP and then a mole of DHAP can be condensed to a mole of glyceraldehyde-3-phosphate to form 1 mole of fructose-1,6-bisphosphate in a reversal of the aldolase reaction. Most non-hepatic tissues lack glucose-6-phosphatase and so the glucose-6-phosphate generated in these tissues would be a substrate for glycogen synthesis. In hepatocytes the glucose-6-phosphatase reactions allows the liver to supply the blood with free glucose. Remember that due to the high K_m of liver glucokinase most of the glucose will not be phosphorylated and will flow down its' concentration gradient out of hepatocytes and into the blood. Place mouse over intermediate names to see structures.

The three reactions of glycolysis that proceed with a large negative free energy change are bypassed during gluconeogenesis by using different enzymes. These three are the pyruvate kinase, phosphofructokinase-1(PFK-1) and hexokinase/glucokinase catalyzed reactions. In the liver or kidney cortex and in some cases skeletal muscle, the glucose-6-phosphate (G6P) produced by gluconeogenesis can be incorporated into glycogen. In this case the third bypass occurs at the glycogen phosphorylase catalyzed reaction. Since skeletal muscle lacks glucose-6-phosphatase it cannot deliver free glucose to the blood and undergoes gluconeogenesis exclusively as a mechanism to generate glucose for storage as glycogen.

Pyruvate to Phosphoenolpyruvate (PEP), Bypass 1

Conversion of pyruvate to PEP requires the action of two mitochondrial enzymes. The first is an ATP-requiring reaction catalyzed by pyruvate carboxylase, (PC). As the name of the enzyme implies, pyruvate is carboxylated to form oxaloacetate (OAA). The CO₂ in this reaction is in the form of bicarbonate (HCO₃^-) . This reaction is an anaplerotic reaction since it can be used to fill-up the TCA cycle. The second enzyme in the conversion of pyruvate to PEP is PEP carboxykinase (PEPCK). PEPCK requires GTP in the decarboxylation of OAA to yield PEP. Since PC incorporated CO₂ into pyruvate and it is subsequently released in the PEPCK reaction, no net fixation of carbon occurs. Human cells contain almost equal amounts of mitochondrial and cytosolic PEPCK so this second reaction can occur in either cellular compartment.

For gluconeogenesis to proceed, the OAA produced by PC needs to be transported to the cytosol. However, no transport mechanism exist for its' direct transfer and OAA will not freely diffuse. Mitochondrial OAA can become cytosolic via three pathways, conversion to PEP (as indicated above through the action of the mitochondrial PEPCK), transamination to aspartate or reduction to malate, all of which are transported to the cytosol.

If OAA is converted to PEP by mitochondrial PEPCK, it is transported to the cytosol where it is a direct substrate for gluconeogenesis and nothing further is required. Transamination of OAA to aspartate allows the aspartate to be transported to the cytosol where the reverse transamination occurs yielding cytosolic OAA. This transamination reaction requires continuous transport of glutamate into, and α-ketoglutarate out of, the mitochondrion. Therefore, this process is limited by the availability of these other substrates. Either of these latter two reactions will predominate when the substrate for gluconeogenesis is lactate. Whether mitochondrial decarboxylation or transamination occurs is a function of the availability of PEPCK or transamination intermediates.

Mitochondrial OAA can also be reduced to malate in a reversal of the TCA cycle reaction catalyzed by malate dehydrogenase (MDH). The reduction of OAA to malate requires NADH, which will be accumulating in the mitochondrion as the energy charge increases. The increased energy charge will allow cells to carry out the ATP costly process of gluconeogenesis. The resultant malate is transported to the cytosol where it is oxidized to OAA by cytosolic MDH which requires NAD⁺ and yields NADH. The NADH produced during the cytosolic oxidation of malate to OAA is utilized during the glyceraldehyde-3-phosphate dehydrogenase reaction of glycolysis. The coupling of these two oxidation-reduction reactions is required to keep gluconeogenesis functional when pyruvate is the principal source of carbon atoms. The conversion of OAA to malate predominates when pyruvate (derived from glycolysis or amino acid catabolism) is the source of carbon atoms for gluconeogenesis. When in the cytoplasm, OAA is converted to PEP by the cytosolic version of PEPCK. Hormonal signals control the level of PEPCK protein as a means to regulate the flux through gluconeogenesis.

The net result of the PC and PEPCK reactions is:

Pyruvate + ATP + GTP + H₂O ——> PEP + ADP + GDP + P_i + 2H⁺

Fructose-1,6-bisphosphate to Fructose-6-phosphate, Bypass 2

Fructose-1,6-bisphosphate (F1,6BP) conversion to fructose-6-phosphate (F6P) is the reverse of the rate limiting step of glycolysis. The reaction, a simple hydrolysis, is catalyzed by fructose-1,6-bisphosphatase (F1,6BPase). Like the regulation of glycolysis occurring at the PFK-1 reaction, the F1,6BPase reaction is a major point of control of gluconeogenesis (see below).

Glucose-6-phosphate (G6P) to Glucose (or Glycogen), Bypass 3

G6P is converted to glucose through the action of glucose-6-phosphatase (G6Pase). This reaction is also a simple hydrolysis reaction like that of F1,6BPase. Since the brain and skeletal muscle, as well as most non-hepatic tissues, lack G6Pase activity, any gluconeogenesis that occurs in these tissues is not utilized for blood glucose supply. In the kidney, muscle and especially the liver, G6P be shunted toward glycogen if blood glucose levels are adequate. The reactions necessary for glycogen synthesis are an alternate bypass series of reactions.

Phosphorolysis of glycogen is carried out by glycogen phosphorylase, whereas, glycogen synthesis is catalyzed by glycogen synthase. The G6P produced from gluconeogenesis can be converted to glucose-1-phosphate (G1P) by phosphoglucose mutase (PGM). G1P is then converted to UDP-glucose (the substrate for glycogen synthase) by UDP-glucose pyrophosphorylase, a reaction requiring hydrolysis of UTP.

Substrates for Gluconeogenesis

Lactate:

Lactate is a predominate source of carbon atoms for glucose synthesis by gluconeogenesis. During anaerobic glycolysis in skeletal muscle, pyruvate is reduced to lactate by lactate dehydrogenase (LDH). This reaction serves two critical functions during anaerobic glycolysis. First, in the direction of lactate formation the LDH reaction requires NADH and yields NAD⁺ which is then available for use by the glyceraldehyde-3-phosphate dehydrogenase reaction of glycolysis. These two reaction are, therefore, intimately coupled during anaerobic glycolysis. Secondly, the lactate produced by the LDH reaction is released to the blood stream and transported to the liver where it is converted to glucose. The glucose is then returned to the blood for use by muscle as an energy source and to replenish glycogen stores. This cycle is termed the Cori cycle.

The Cori cycle invloves the utilization of lactate, produced by glycolysis in non-hepatic tissues, (such as muscle and erythrocytes) as a carbon source for hepatic gluconeogenesis. In this way the liver can convert the anaerobic byproduct of glycolysis, lactate, back into more glucose for reuse by non-hepatic tissues. Note that the gluconeogenic leg of the cycle (on its own) is a net consumer of energy, costing the body 4 moles of ATP more than are produced during glycolysis. Therefore, the cycle cannot be sustained indefinitely.

Pyruvate:

Pyruvate, generated in muscle and other peripheral tissues, can be transaminated to alanine which is returned to the liver for gluconeogenesis. The transamination reaction requires an α-amino acid as donor of the amino group, generating an α-keto acid in the process. This pathway is termed the glucose-alanine cycle. Although the majority of amino acids are degraded in the liver some are deaminated in muscle. The glucose-alanine cycle is, therefore, an indirect mechanism for muscle to eliminate nitrogen while replenishing its energy supply. However, the major function of the glucose-alanine cycle is to allow non-hepatic tissues to deliver the amino portion of catabolized amino acids to the liver for excretion as urea. Within the liver the alanine is converted back to pyruvate and used as a gluconeogenic substrate (if that is the hepatic requirement) or oxidized in the TCA cycle. The amino nitrogen is converted to urea in the urea cycle and excreted by the kidneys.

The glucose-alanine cycle is used primarily as a mechanism for skeletal muscle to eliminate nitrogen while replenishing its energy supply. Glucose oxidation produces pyruvate which can undergo transamination to alanine. This reaction is catalyzed by alanine transaminase, ALT (ALT used to be referred to a serum glutamate-pyruvate transaminase, SGPT). Additionally, during periods of fasting, skeletal muscle protein is degraded for the energy value of the amino acid carbons and alanine is a major amino acid in protein. The alanine then enters the blood stream and is transported to the liver. Within the liver alanine is converted back to pyruvate which is then a source of carbon atoms for gluconeogenesis. The newly formed glucose can then enter the blood for delivery back to the muscle. The amino group transported from the muscle to the liver in the form of alanine is converted to urea in the urea cycle and excreted.

Amino Acids:

All 20 of the amino acids, excepting leucine and lysine, can be degraded to TCA cycle intermediates as discussed in the metabolism of amino acids. This allows the carbon skeletons of the amino acids to be converted to those in oxaloacetate and subsequently into pyruvate. The pyruvate thus formed can be utilized by the gluconeogenic pathway. When glycogen stores are depleted, in muscle during exertion and liver during fasting, catabolism of muscle proteins to amino acids contributes the major source of carbon for maintenance of blood glucose levels.

Glycerol:

Oxidation of fatty acids yields enormous amounts of energy on a molar basis, however, the carbons of the fatty acids cannot be utilized for net synthesis of glucose. The two carbon unit of acetyl-CoA derived from β-oxidation of fatty acids can be incorporated into the TCA cycle, however, during the TCA cycle two carbons are lost as CO₂. Thus, explaining why fatty acids do not undergo net conversion to carbohydrate.

The glycerol backbone of lipids can be used for gluconeogenesis. This requires phosphorylation to glycerol-3-phosphate by glycerol kinase and dehydrogenation to dihydroxyacetone phosphate (DHAP) by glyceraldehyde-3-phosphate dehydrogenase (G3PDH). The G3PDH reaction is the same as that used in the transport of cytosolic reducing equivalents into the mitochondrion for use in oxidative phosphorylation. This transport pathway is called the glycerol-phosphate shuttle.

The glycerol phosphate shuttle is a secondary mechanism for the transport of electrons from cytosolic NADH to mitochondrial carriers of the oxidative phosphorylation pathway. The primary cytoplasmic NADH electron shuttle is the malate-aspartate shuttle. Two enzymes are involved in this shuttle. One is the cytosolic version of the enzyme glycerol-3-phosphate dehydrogenase (glycerol-3-PDH) which has as one substrate, NADH. The second is is the mitochondrial form of the enzyme which has as one of its' substrates, FAD⁺. The net result is that there is a continual conversion of the glycolytic intermediate, DHAP and glycerol-3-phosphate with the concomitant transfer of the electrons from reduced cytosolic NADH to mitochondrial oxidized FAD⁺. Since the electrons from mitochondrial FADH₂ feed into the oxidative phosphorylation pathway at coenzyme Q (as opposed to NADH-ubiquinone oxidoreductase [complex I]) only 2 moles of ATP will be generated from glycolysis. G3PDH is glyceraldehyde-3-phoshate dehydrogenase.

The glycerol backbone of adipose tissue stored triacylgycerols is ensured of being used as a gluconeogenic substrate since adipose cells lack glycerol kinase. In fact adipocytes require a basal level of glycolysis in order to provide them with DHAP as an intermediate in the synthesis of triacyglycerols.

Propionate:

Oxidation of fatty acids with an odd number of carbon atoms and the oxidation of some amino acids generates as the terminal oxidation product, propionyl-CoA. Propionyl-CoA is converted to the TCA intermediate, succinyl-CoA. This conversion is carried out by the ATP-requiring enzyme, propionyl-CoA carboxylase then methylmalonyl-CoA epimerase and finally the vitamin B₁₂ requiring enzyme, methylmalonyl-CoA mutase. The utilization of propionate in gluconeogenesis only has quantitative significance in ruminants.

Conversion of Propionyl-CoA to Succinyl-CoA

Regulation of Gluconeogenesis

Obviously the regulation of gluconeogenesis will be in direct contrast to the regulation of glycolysis. In general, negative effectors of glycolysis are positive effectors of gluconeogenesis. Regulation of the activity of PFK-1 and F1,6BPase is the most significant site for controlling the flux toward glucose oxidation or glucose synthesis. As described in control of glycolysis, this is predominantly controlled by fructose-2,6-bisphosphate, F2,6BP which is a powerful negative allosteric effector of F1,6Bpase activity.

Regulation of glycolysis and gluconeogenesis by fructose 2,6-bisphosphate (F2,6BP). The major sites for regulation of glycolysis and gluconeogenesis are the phosphofructokinase-1 (PFK-1) and fructose-1,6-bisphosphatase (F-1,6-BPase) catalyzed reactions. PFK-2 is the kinase activity and F-2,6-BPase is the phosphatase activity of the bi-functional regulatory enzyme, phosphofructokinase-2/fructose-2,6-bisphosphatase. PKA is cAMP-dependent protein kinase which phosphorylates PFK-2/F-2,6-BPase turning on the phosphatase activity. (+ve) and (-ve) refer to positive and negative activities, respectively.

The level of F2,6BP will decline in hepatocytes in response to glucagon stimulation as well as stimulation by catecholamines. Each of these signals is elicited through activation of cAMP-dependent protein kinase (PKA). One substrate for PKA is PFK-2, the bifunctional enzyme responsible for the synthesis and hydrolysis of F2,6BP. When PFK-2 is phosphorylated by PKA it acts as a phosphatase leading to the dephosphorylation of F2,6BP with a concomitant increase in F1,6Bpase activity and a decrease in PFK-1 activity. Secondarily, F1,6Bpase activity is regulated by the ATP/ADP ratio. When this is high, gluconeogenesis can proceed maximally.

Gluconeogenesis is also controlled at the level of the pyruvate to PEP bypass. The hepatic signals elicited by glucagon or epinephrine lead to phosphorylation and inactivation of pyruvate kinase (PK) which will allow for an increase in the flux through gluconeogenesis. PK is also allosterically inhibited by ATP and alanine. The former signals adequate energy and the latter that sufficient substrates for gluconeogenesis are available. Conversely, a reduction in energy levels as evidenced by increasing concentrations of ADP lead to inhibition of both PC and PEPCK. Allosteric activation of PC occurs through acetyl-CoA. Each of these regulations occurs on a short time scale, whereas long-term regulation can be effected at the level of PEPCK. The amount of this enzyme increases in response to prolonged glucagon stimulation. This situation would occur in a starving individual or someone with an inadequate diet.

The mechanism of carbohydrate metabolism regulation by nervous and endocrine systems

Тhe role of the central nervous system in regulation of carbohydrate metabolism

We are dealing here with the major action of epinephrine to mobilize glycogen from liver (shown), and also muscle (which lacks PFK-2 and glucose-6-phosphatase). The context is acute stress, so the liver's role is to release fuel into the blood, while muscle would then use the fuel, from either the liver or from its own glycogen stores, for the "fight or flight response".

The coordinate regulation of glucose metabolism is via phosphorylation, the opposite of insulin's action via dephosphorylation. As you will recall, the G_s/cAMP/protein kinase A pathway is involved in many important phosphorylation events, including the ones here. Recall also that epinephrine, unlike the peptide hormone glucagon,is a catecholamine and thus must bind to an adrenergic receptor.

Which adrenergic receptor subtype(s) act via G_s?

Primary Action of Epinephrine in a Liver Cell

This is essentially the same figure as for insulin action in liver (question 1 of this module), and the five enzymes in question are denoted with question marks. The key for arriving at the correct answer is to determine which enzymes are missing in muscle.

Insulin Affects both Glucose and Lipid Metabolism.

The authors present information about "cross-talk" between various organs.  Here, the signal initiating "cross-talk" is either an increase in blood sugar or fatty acids levels. This can occur following a meal and uptake from the small intestine or as a result of stimulation of glucose release from the liver.  The figure is simplified to aid understanding, but remember, changes in glucagon usually oppose alterations in insulin levels.  Thus, gluconeogenesis and glycogenolysis are often initiated by rising glucagon and falling insulin levels.

Increased circulating glucose levels stimulate pancreatic secretion of insulin.  This has several immediate effects:

1.  Increased skeletal muscle glucose uptake.

2.  Inhibition of hepatic gluconeogenesis and glycogenolysis and stimulation of glucose uptake in the liver (not shown).

3.  Inhibition of lipolysis in fat tissue. 

Muscle tissue and liver do not just take up glucose.  They must "do something with it" or it will diffuse over the cell membrane and return to the circulation.  Both tissues have glycogen reserves and these will be filled when glucose is taken up.  Further, skeletal muscle, which makes up over 50% of the body, will use glucose as a substrate for "aerobic glycolysis", that is, complete glucose "burning" from the sugar phosphate and mitochondrial metabolism to CO₂ and water.  Approximately  25% of the carbohydrate content of a meal will normally be used as an energy source in skeletal muscles.  You can click here for more information about carbohydrate metabolism after meals.

What happens after a meal?  Insulin stimulates muscular glucose uptake and metabolism, and forces the tissue to reduce the use of fat as an energy substrate.  After all, acetyl-CoA, the common substrate formed from both sugars and fats for entry into mitochondrial metabolism is used at a constant rate as long as the work load does not change.    I will come back to control of fatty acid use soon but will point out here that insulin inhibits release of fatty acids from fat cells (inhibits lipolysis as shown in the figure above).  Muscle takes up and uses fatty acids in proportion to the concentration of fatty acids in blood.  Thus, insulin speeds up glucose uptake and metabolism, while setting down the rate of lipolysis and release of fatty acids from fat cells to the circulation.   This sounds like a simple rule, but things are not so simple.  Remember, insulin swings markedly after a meal, increasing from basal to maximal concentrations during the first hour after eating, then falls rapidly.  At the same time, glucagon levels swing in the opposite direction and have effects opposing insulin.  The united effect on metabolism is always the result of the balance between these two hormones.  Metabolism and the associated choice of energy substrate follow an integrated response to the hormonal picture.

We can also see from the figure that free fatty acids reduce insulin's effect on glucose uptake.  Free fatty acids are involved in "insulin resistance", that is, the concentration of fatty acids in the blood is inversely correlated with tissue responsiveness to insulin.  Chronically increased serum fatty acids as seen in overweight and obesity are implicated in the development of diabetes type 2, where we see a decreased response to insulin, often counteracted by markedly increased insulin levels.   

Fat cells produce a number of peptide hormones (adipokines) that have been identified during the past five to ten years.  These are involved in regulation of tissue response to hormones.   Resistin appears to dampen muscle, liver and fat cell responses  to insulin as does TNF-α.  Adiponectin sensitizes receptor-cells to insulin.  Leptin regulates metabolism and appetite. 

Control of Carbohydrate Metabolism; the "Phosphofructokinase-Fructose bisphosphate phosphatase Couple".

Blood sugar, or glucose, is the major source of energy for many tissues.  Blood cells and the brain are normally completely dependent upon blood sugar.  Their metabolism is locked to this substrate and they have no reserve carbohydrate.  Glycogen stores are not found in these tissues.  And, while skeletal muscle can cover much of its energy requirement through oxidation of fats, hard working muscle uses carbohydrates.  Muscle tissue can, in fact, take up so much glucose from the circulation that hypoglycemia and loss of consciousness results. 

We can get an overview of regulation of carbohydrate by studying hepatic metabolism.  We find all of the hormone and enzyme functions that control carbohydrate metabolism there.  The major control points in glycolysis and gluconeogenesis are the enzymes which catalyze the reactions between fructose-6-phosphate and fructose-1,6-bisphosphate.  Phosphofructokinase-1 (PFK-1) and fructose bisphosphate phosphatase are regulated by allosteric "feedback" mechanisms and by hormones.  They are regulated by common signal substances.  However, these have opposite effects on these two enzymes and, therefore, upon metabolism. 

Let us look at PFK-1 first.  The PFK-1 step is the slowest in glucose metabolism (glycolysis).  It is, therefore, very well suited as THE primary controlling point in this process.  PFK-1 is inhibited by ATP and stimulated by its breakdown product, 5´-AMP.  We have previously seem that ATP levels are surprisingly stabile while AMP swings markedly during energy utilization.  PFK-1 is sensitive to the physiological concentrations of these nucleotides and its activity increases as AMP levels increase. 

PFK-1 is also sensitive to citrate which is released from the mitochondria to the cytosol when the liver uses fatty acids.  This occurs between meals and is a part of the "fatty acids spare carbohydrate" business.  Not only does fatty acid oxidation turn off pyruvate dehydrogenase and pyruvate uptake to the mitochondria;  it also turns off the source of pyruvate. 

Both PFK-1 and fructose-1,6-bisphosphate phosphatase are regulated by another of those "fructose-diphosphate" things.  A hormone sensitive kinase, phosphofructokinase-2, produces the 2,6 bisphosphate from fructose-6-phosphate.  This kinase is subject to cyclic AMP-stimulated phosphorylation.  The phosphorylated form has phosphatase activity, not kinase activity.  The phosphorylated form uses fructose-2,6-bisphosphate as its substrate, thus reversing the effects of the non-phosphorylated PFK-2. 

Fructose-2,6-bisphosphatase controls carbohydrate metabolism by regulating the activities of PFK-1 and fructose bisphosphate phosphatase.  Hormones that increase the rate of glycolysis increase the level of fructose-2,6-bisphosphate.  Hormones that phosphorylate PFK-2 reduce the levels of fructose-2,6-bisphosphate and favor gluconeogenesis. 

The liver is sensitive to several hormones that increase cyclic AMP.  These are glucagon, adrenalin and noradrenalin.  These inhibit glycolysis by reducing the concentrations of an activator of PFK-1 (fructose-2,6-bisphosphate).  The same hormones stimulate gluconeogenesis by removing an inhibitor of the key enzyme (by inhibiting the action of an inhibitor). 

The liver is also responsive to insulin which increases  breakdown of cyclic AMP through activation of phosphodiesterase.  Thus, insulin activates glycolysis by increasing the activity of PFK-2 and synthesis of fructose-2,6-bisphosphate.  This is coordinated with inhibition of gluconeogenesis at the fructose bisphosphate phosphatase step by the same signaling substance. 

In summary, it is fructose-2,6-bisphosphate levels that are a major regulator of carbohydrate metabolism.  This is a control substance synthesized in answer to stress or hunger and geared towards stabilizing blood glucose levels.  Reducing synthesis of fructose-2,6-bisphosphate  turns off carbohydrate "burning" and starts up glucose production from smaller substances. 

Тhe insulin effect on the carbohydrate metabolism

Тhe role of cyclic AMP in mechanism of epinephrine and glucagon action

Transduction has many steps and can result in amplification of signal. (Small signal produces big response…)

1) Transduction pathways relay information. The ligand binds to a receptor on the plasma membrane, causing a conformational change in the receptor, which triggers a cascade of events. The activated receptor protein activates a series of other proteins that relay the information.
2) Protein phosphorylation is major mechanism of signal transduction in cells.

Protein kinase--

    Protein phosphotase--

3) Second messengers play a role in signal transduction pathways. Small, water-soluble, and nonprotein, they can spread throughout cell by diffusion. Two most common cyclic AMP and Ca++.

Cyclic AMP

b. Calcium ions / Inositol Triphosphate:

Diabetes mellitus: types, causes, disorders of metabolism, symptoms, treatment

Types 1

Causes, incidence, and risk factors

Diabetes is a lifelong disease for which there is not yet a cure. There are several forms of diabetes. Type 1 diabetes is often called juvenile or insulin-dependent diabetes. In this type of diabetes, cells of the pancreas produce little or no insulin, the hormone that allows glucose to enter body cells.

Without enough insulin, glucose builds up in the bloodstream instead of going into the cells. The body is unable to use this glucose for energy despite high levels in the bloodstream. This leads to increased hunger.

In addition, the high levels of glucose in the blood cause the patient to urinate more, which in turn causes excessive thirst. Within 5 to 10 years, the insulin-producing beta cells of the pancreas are completely destroyed and the body can not longer produce insulin.

Type 1 diabetes can occur at any age. Many patients, however, are diagnosed after age 20.

The exact cause is unknown. Genetics, viruses, and auto-immune problems may play a role.

Definition

Type 2 diabetes is a life-long disease marked by high levels of sugar in the blood. It occurs when the body does not respond correctly to insulin, a hormone released by the pancreas. Type 2 diabetes is the most common form of diabetes.

Causes, incidence, and risk factors

Diabetes is caused by a problem in the way your body makes or uses insulin. Insulin is needed to move glucose (blood sugar) into cells, where it is used for energy.

If glucose does not get into the cells, the body cannot use it for energy. Too much glucose will then remain in the blood, causing the symptoms of diabetes.

There are several types of diabetes. This article focuses on type 2, which is usually accompanied by obesity and insulin resistance.

Insulin resistance means that insulin produced by your pancreas cannot get inside fat and muscle cells to produce energy. Since the cells are not getting the insulin they need, the pancreas produces more and more. Over time, abnormally high levels of sugar build up in the blood. This is called hyperglycemia. Many people with insulin resistance have hyperglycemia and high blood insulin levels at the same time. People who are overweight have a higher risk of insulin resistance, because fat interferes with the body's ability to use insulin.

Type 2 diabetes usually occurs gradually. Most people with the disease are overweight at the time of diagnosis. However, type 2 diabetes can also develop in those who are thin, especially the elderly.

Family history and genetics play a large role in type 2 diabetes. Low activity level, poor diet, and excess body weight (especially around the waist) significantly increase your risk for type 2 diabetes.

Other risk factors include:

• Race/ethnicity (African-Americans, Hispanic-Americans, and Native Americans all have high rates of diabetes)
• Age greater than 45 years
• Previously identified impaired glucose tolerance by your doctor
• High blood pressure
• HDL cholesterol of less than 35 mg/dL or triglyceride level of greater than 250 mg/dL
• History of gestational diabetes

Symptoms

Often, people with type 2 diabetes have no symptoms at all. If you do have symptoms, they may include:

• Increased thirst
• Increased urination
• Increased appetite
• Fatigue
• Blurred vision
• Frequent or slow-healing infections
• Erectile dysfunction

Biochemical differentiation of diabetes mellitus, diabetes insipidus, steroid diabetes,

liver diabetes , kidney diabetes .

Nephrogenic Diabetes Insipidus (NDI) is characterised by the inability of the kidneys to concentrate urine in response to arginine vasopressin (AVP). Such patients typically experience polyuria and polydipsia because of this inability to autoregulate their water balance. This provides a perioperative challenge that could lead to a life-threatening situation. This article documents a patient with NDI who underwent an elective bowel re-anastomosis. Two peak serum sodium values were attained. The first when the patient was retaining sodium due to an inappropriate fluid regimen and the second due to hypovolaemia. The literature is reviewed and principles for NDI perioperative management are proposed

INTRODUCTION

Nephrogenic diabetes insipidus (NDI) is characterised by the inability of the kidneys to concentrate urine in response to arginine vasopressin (AVP).1 In the majority of cases NDI is acquired, but X-linked congenital NDI (due to a defect in the arginine vasopressin receptor 2) and autosomal recessive NDI (due to a genetic defect in the aquaporin 2 protein) have been described.

Lithium therapy is the commonest cause of acquired NDI. It is estimated that NDI is a feature of approximately 12% of patients receiving lithium, with a further 20-40% having urinary concentration dysfunction.

NDI patients classically experience polyuria and polydipsia because of this inability to autoregulate their water balance. This provides a perioperative challenge that could lead to a life-threatening situation.

Normal water homeostasis consists of a series of complex interactions regulated by AVP that is essential for our survival. Nephrogenic diabetes insipidus is a disorder of water balance that results from the kidney being unable to concentrate urine in response to AVP stimulation. In NDI the renal tubules are totally or, more often, partially resistant to the action of AVP.

Figure 2: The pathway of arginine vasopressin (AVP) - mediated water resorption.

Lithium impairs the AVP stimulatory effect on adenylate cyclase.^4,5 This leaves the aquaporins, or water channels, unable to fully open resulting in decreased water resorption. ^6,7 Most renal impairment resolves with lithium withdrawal but the effect can persist, with one case documenting impairment eight years after stopping lithium.⁸ Furthermore, renal impairment can become severe in a small number of cases, leaving patients requiring dialysis.

Nephrogenic diabetes insipidus is characterised by polyuria and polydipsia. These in turn can result in: dehydration (potentially life-threatening), hypernatraemia, hydronephrosis and seizures. In addition, abrupt correction of hypernatraemia can result in pontine demyelination, coma and death.

In this case report, pre-operative fluids and electrolytes were given to correct the serum sodium and potassium levels. With no signs of pronounced hypovolaemia pre-operatively, the patient became hypotensive after the induction of anaesthesia. Initially, it was thought that the hypotension was due to hypovolaemia and the patient received large infusions of saline and Hartmann’s with little improvement. However, a more likely explanation is that the patient had a hypotensive response as a result of the temporary vasodilatation induced by the anaesthetic (remifentanil and propofol). Every anaesthetic attempts to compensate for the stress of surgical stimulation. Following induction and intubation, a period follows when there is no surgical stimulus as the patient is prepared and draped. If the depth of the anaesthesia is too great at this time, profound cardiovascular depression can occur. The compensatory mechanisms for hypovolaemia are overcome by the vasodilating effects that are inherent in most anaesthetic agents. This patient’s CVP was found to be low on insertion of the central line, but was refractory to generous fluid resuscitation. The systemic blood pressure finally responded to a low dose norepinepherine infusion supporting vasodilatation as a major factor in her hypotension.

Post-operatively, the patient had a CVP of 10cm H20, normal serum urea and was 7480ml positive, yet had a serum sodium of 166mmol/l. This last result led to the continuation of fluid resuscitation for hypovolaemia with further 0.9% saline, which the patient responded to by increasing their serum sodium to 172mmol/l. At this point, the fluid regimen was changed to predominantly 5% dextrose as it was decided that the sodium concentration was increasing as a result of the 0.9% saline infusion, rather than haemoconcentration from dehydration.

This regimen was successful. The fundamental defect in NDI is the inability to excrete concentrated urine. Thus, these patients are unable to excrete the solute load presented to them when given large amounts of 0.9% saline. This results in iatrogenic hypernatraemia due to sodium retention, which occurred in this patient. Therefore, when hypernatraemia is present in NDI patients, the major component of the fluid replacement should be in the form of 5% dextrose with oral water.

On day six the patient had an episode of vomiting that resulted in an increase in the serum urea to 8.6mmol/l. The patient was in negative fluid balance and the serum sodium increased to 153mmol/l. The elevation in serum sodium in this episode was not due to the patient retaining sodium, but due to dehydration.

These two peaks of serum sodium represent two different scenarios: euvolaemia and hypovolaemia. This case report documents both scenarios in the one patient and emphasises that the sodium values must always be interpreted in the hydration context of the patient and not always assumed to be secondary to hypovolaemia.

Steroid diabetes is a medical term referring to prolonged hyperglycemia due to glucocorticoid therapy for another medical condition. It is usually, but not always, a transient condition.

The most common glucocorticoids which cause steroid diabetes are prednisone and dexamethasone given systemically in "pharmacologic doses" for days or weeks. Typical medical conditions in which steroid diabetes arises during high-dose glucocorticoid treatment include severe asthma, organ transplantation, cystic fibrosis, inflammatory bowel disease, and induction chemotherapy for leukemia or other cancers.

Glucocorticoids oppose insulin action and stimulate gluconeogenesis, especially in the liver, resulting in a net increase in hepatic glucose output. Most people can produce enough extra insulin to compensate for this effect and maintain normal glucose levels, but those who cannot develop steroid diabetes.

The diagnostic criteria for steroid diabetes are those of diabetes (fasting glucoses persistently above 125 mg/dl (7 mM) or random levels above 200 mg/dl (11 mM)) occurring in the context of high-dose glucocorticoid therapy. Insulin levels are usually detectable, and sometimes elevated, but inadequate to control the glucose. In extreme cases the hyperglycemia may be severe enough to cause nonketotic hyperosmolar coma.

Treatment depends on the severity of the hyperglycemia and the estimated duration of the steroid treatment. Mild hyperglycemia in an immunocompetent patient may not require treatment if the steroids will be discontinued in a week or two. Moderate hyperglycemia carries an increased risk of infection, especially fungal, and especially in people with other risk factors such as immunocompromise or central intravenous lines. Insulin is the most common treatment.

Steroid diabetes must be distinguished from stress hyperglycemia, hyperglycemia due to excessive intravenous glucose, or new-onset diabetes of another type. Because it is not unusual for steroid treatment to precipitate type 1 or type 2 diabetes in a person who is already in the process of developing it, it is not always possible to determine whether apparent steroid diabetes will be permanent or will go away when the steroids are finished. More commonly undiagnosed cases of type 2 diabetes are brought to clinical attention with corticosteroid treatment because subclinical hyperglycemia worsens and becomes symptomatic. Generally, steroid diabetes without preexisting type 2 diabetes will resolve upon termination of corticosteroid administration.

Diabetic nephropathy (nephropatia diabetica), also known as Kimmelstiel-Wilson syndrome and intercapillary glomerulonephritis, is a progressive kidney disease caused by angiopathy of capillaries in the kidney glomeruli. It is characterized by nephrotic syndrome and nodular glomerulosclerosis. It is due to longstanding diabetes mellitus, and is a prime cause for dialysis in many Western countries.

The glucose-tolerance test

Glucose tolerance test To test for diabetes, patients are often given a glucose tolerance test. In this test, they drink a quantity of glucose and then their blood glucose is checked to see:

· How high it rises

· How long it takes for their blood glucose to return to a normal level.

At the right you can see a typical glucose tolerance test for a healthy person.
Note that a similar tolerance test using fat does not raise blood glucose

The need for replacement of the missing early-phase insulin response after an oral glucose tolerance test (OGTT) is readily apparent, even more so than under normal physiologic conditions. Compared with regular human insulin, administration of insulin lispro resulted in a significantly improved plasma insulin profile, lower plasma glucose levels, and lower plasma C-peptide levels over the course of nearly 6 hours.