Metabolism Flashcards

Question

LO 3.2 Describe the clinical condition of glucose 6-phosphate dehydrogenase deficiency and explain the biochemical basis of the signs and symptoms

Answer 1

Glucose 6-phoshate dehydrogenase is the rate-limiting enzyme in the pentose phosphate pathway, used to increase NADPH concentration. Deficiency in this enzyme is caused by a point mutation in the X-linked gene coding for the enzyme. The mutation results in reduced activity of the enzyme and therefore low levels of NADPH. The structural integrity and functional activity of proteins in RBCs depends on free –SH groups. –SH groups tend to form disulphide bridges unless prevented by NADPH. In G6PD deficiency the NADPH levels are sometimes too low to prevent the formation of these disulphide bridges. Haemoglobin and other proteins then become cross-linked by disulphide bonds to form insoluble aggregates called Heinz bodies. This leads to premature destruction of the RBCs (haemolysis).

Answer 2

Pyruvate does not enter stage 3 of catabolism directly; instead it is converted to Acetyl~CoA. The enzyme responsible for this is Pyruvate dehydrogenase (PDH) a multi-enzyme complex. The PDH reaction is irreversible in the cell. This means that the loss of CO2 from Pyruvate is irreversible and Acetyl~CoA cannot be converted back to Pyruvate for use in gluconeogenesis to produce glucose. PDH is subject to control mechanisms: o Acetyl~CoA from the -oxidation of fatty acids rather than from glucose is used in stage 3 catabolism (acetyl~CoA allosterically inhibits PDH) o The reaction is energy sensitive. ATP/NADH inhibit and ADP promotes allosterically. o The enzyme is activated when there is plenty of glucose to be catabolised (insulin activates the enzyme by promoting its Dephosphorylation).

Answer 3

The TCA cycle (Krebs cycle, citric acid cycle, stage 3 catabolism) is a central pathway in the catabolism of sugars, fatty acids, ketone bodies, alcohol and amino acids. It is an oxidative pathway that occurs in mitochondria. o The pathway requires NAD+, FAD and oxaloacetate. o Main function is to break the C-C bond in acetate (as acetyl~CoA) and oxidise the C atoms to CO2. o The H¬+ and e- removed from the acetate are transferred to NAD+ and FAD. o The pathway is of fundamental importance to the major energy requiring tissues of the body, and does not function in the absence of oxygen. o There are no known defects in the pathway, as any would be lethal. It is estimated that the TCA cycle leads to the production of 32 molecules of ATP per molecule of glucose. The chemical strategy of the pathway is to produce intermediates (C6 tricarboxylic acids and C5 keto-acids) that readily lose CO2 producing C4 acids that are interconvertible. As well as these catabolic functions the pathway as anabolic functions. o C5 and C4 intermediates used for the synthesis of non-essential amino acids o C4 intermediates used for the synthesis of haem and glucose o C6 intermediates used for the synthesis of fatty acids

Answer 4

The oxidation of Acetyl~CoA linked to the reduction of NAD+ and FAD by the TCA cycle is essential for the generation of ATP in all mitochondria-containing tissues. Therefore two major signals feed information to the TCA cycle on the rate of ATP use: o ATP/ADP ratio o NADH/NAD+ ratio One of the irreversible steps in the TCA (catalysed by isocitrate dehydrogenase) is allosterically inhibited by the high-energy signal NADH and activated by the low-energy signal ADP.

Answer 5

Oxidative Phosphorylation is Stage 4 of catabolism. The complete oxidation of glucose: C6H12O6 + 6O2  6CO2 + 6H2O G = -2,870 kJ/mole By the end of stage 3 (TCA cycle): o All C-C bonds have been broken, and C-atoms oxidised to CO2 o All C-H bonds have been broken, and H-atoms (H¬+ and e-) transferred to NAD+ and FAD. All of the energy from the breaking of these bonds has gone to: o ATP/GTP formation (2 in glycolysis, 2 in the TCA cycle) o Chemical bond energy of the e- in NADH/FAD2H NADH and FAD2H contain high energy electrons that can be transferred to oxygen through a series of carrier molecules, releasing large amounts of free energy. NADH + H+ + O  NAD+ + H2O G = -220 kJ/mole FAD2H + O  FAD + H2O G = -152 kJ/mole This energy can be used to drive ATP synthesis in the final stage of catabolism (oxidative phosphorylation), occurring in the inner mitochondrial membrane. o Electron Transport, electrons in NADH and FAD2H are transferrerd through a series of carrier molecules to oxygen, releasing free energy. o ATP synthesis, the free energy released in electron transport drives ATP synthesis from ADP + Pi

Answer 6

Electron transport o Carrier molecules transferring electrons to molecular oxygen are organized into a series of four highly specialized protein complexes spanning the inner mitochondrial membrane. o Electrons are transferred from NADH (and FAD2H) sequentially through the series of complexes to molecular oxygen with the release of free energy. o Three of the complexes, in addition to transferring electrons, also act as proton translocation complexes (proton pump). Proton Motive Force (PMF) o Free energy from electron transport is used to move protons from the inside to the outside of the inner mitochondrial membrane via p.t.complexes. o The membrane itself is impermeable to protons and as electron transport continues the concentration of protons outside the inner membrane increases. o The proton translocating complexes therefore transform the chemical bond energy of the electrons into an electro-chemical gradient. o This is known as the Proton Motive Force. o NADH has more energy than FAD2H and so uses all three p.t.complexes while FAD2H only uses two. o This process requires oxygen, as it is the last electron acceptor. ATP Synthesis o ATP Hydrolysis results in the release of energy (G = -31kJ/mol). Therefore for the synthesis of ATP from ADP and Pi, + 31 kJ/mol of energy is required to drive the reaction. o This energy is derived from the pmf that has been produced across the inner mitochondrial membrane by electron transport. o Protons can normally only re-enter the mitochondrial matrix via the ATP synthase complex, driving the synthesis of ATP from ADP and Pi. The greater the PMF the more ATP synthesised The oxidation of 2 moles of NADH gives 5 moles of ATP The oxidation of 2 moles of FAD2h gives 3 moles of ATP Coupling of Electron Transport and ATP synthesis ET and ATP Synthesis are tightly coupled. One does not occur without the other. The mitochondrial concentration of ATP plays an important role in regulating both processes. When ATP concentration is high: - The ADP concentration is low and the ATP synthase stops (lack of substrate) - This prevents H+ transport back into the mitochondria - The H+ concentration outside increases to a level that prevents more protons being pumped to the outside - In the absence of proton pumping, electron transport stops

Answer 7

Some substances (eg dinitrophenol, dinitrocresol) increase the permeability of the inner mitochondrial membrane to protons. Therefore protons being pumped out by electron transport can re-enter without passing through the ATP synthase complex. The two processes become uncoupled so the p.m.f. is dissipated as heat. Proton leak is physiologically important and accounts for 20-25% of the BMR. Uncoupling Proteins (UCPs) The function of UCPs is to uncouple ET from ATP production to produce heat. The proteins are located in the inner mitochondrial membrane and allow a leak of protons across the membrane. UCP1 - (previously known as thermogenin) is expressed in brown adipose tissue and involved in non-shivering thermogenesis enabling mammals to survive the cold. UCP2 – Quite widely distributed in the body. Research suggest it is linked to diabetes, obesity, metabolic syndrome and heart failure. UCP3 – Found in skeletal muscle, brown adipose tissue and the heart. It appears to be involved in modifying fatty acid metabolism and in protecting against ROS damage. Noradrenaline – Is released from the sympathetic nervous system and stimulates lipolysis releasing fatty acids to provide fuel for oxidation in brown adipose tissue. NADH and FAD2H are formed as a result of -oxidation of the fatty acids. NADH and FAD2H drive ET and increase p.m.f. However, noradrenaline also activates UCP1, allowing protons to cross the inner mitochondrial membrane without passing through the ATP synthase complex. The higher p.m.f. is dissipated as heat.

Answer 8

Oxidative Phosphorylation Requires membrane associated complexes (inner mitochondrial membrane) Energy coupling occurs indirectly through generation and subsequent utilisation of a proton gradient (p.m.f.) Cannot occur in the absence of oxygen Major process for ATP synthesis in cells that require large amounts of energy Substrate Level Phosphorylation Requires soluble enzymes. (Cytoplasmic and mitochondrial matrix) Energy coupling occurs directly through formation of a high energy of hydrolysis bond (phosphoryl-group transfer) Can occur to a limited extent in absence of oxygen Minor process for ATP synthesis in cells that require large amounts of energy

Answer 9

Lipids are a structurally diverse group of important compounds that are generally insoluble in water (hydrophobic) but are soluble in organic solvents. There is no general formula but most contain C, H and O (phospholipids also contain P and N). They are more reduced than carbohydrates (contain less O and more H per C-atom). Classes of lipids 1. Fatty acid derivatives o Fatty Acids – Fuel molecules o Triacylglycerols – Fuel storage and insulation o Phospholipids – Components of membranes and plasma lipoproteins o Eicosanoids – Local mediators 2. Hydroxy-methyl-glutaric acid derivatives (C6 compound) o Ketone bodies (C4) – Water soluble fuel molecules o Cholesterol (C27) – Membranes and steroid hormone synthesis o Cholesterol esters – Cholesterol storage o Bile acids and salts (C24) – Lipid Digestion 3. Fat Soluble Vitamins o A, D, E and K

Answer 10

Triacylglycerols are the major dietary lipids (butter, vegetable oils) and are hydrolysed by pancreatic lipase in the small intestine to release glycerol and fatty acids. This is a complex process requiring bile salts and a protein factor called colipase. Glycerol metabolism Glycerol derived from the hydrolysis of dietary triacylglycerols enters the blood stream and is transported to the liver where it is metabolized. Fatty Acids The most common fatty acids in the body are long-chain molecules that contain an even number of C-atoms: CH3(CH2)nCOOH (n=14 to 18). They are hydrophobic and highly reduced molecules, properties that make them ideal for energy storage. They may be saturated or unsaturated (unsaturated contain C=C bonds). Saturated fatty acids are a non-essential part of the diet as they can be synthesised from carbohydrates and certain amino acids. Over 50% of fatty acids in the body are unsaturated and contain between 1 and 4 C=C bonds. Certain polyunsaturated fatty acids are essential components of the diet as they cannot be synthesised in the body. Arachidonic acid (C20:4) is an important polyunsaturated fatty acid as it is the starting point for the synthesis of the eicosanoids.

Answer 11

There are three ketone bodies produced in the body: o Acetoacetate – CH3COCH2COO- o Acetone – CH3COCH3 (Spontaneous non-enzymatic decarboxylation of above) o B-hydroxybutyrate – CH3CHOHCH2COO- Acetoacetate and B-hydroxybutyrate are synthesised in the liver from Acetyl~CoA. Normally the concentration of ketone bodies in the cirulation is low (10mM – pathological ketosis) Ketone bodies are water-soluble molecules, allowing high plasma concentration and their excretion in urine (ketonuria). Acetoacetate and B-hydroxybutyrate are relatively strong organic acids and when in high concentration in the plasma they may cause acidosis (ketoacidosis). Acetone is volatile and may be excreted via the lungs (acetone smell on breath of untreated type 1 diabetes). Ketone body synthesis/Regulation Hydroxymethyl glutaryl CoA lysase/reductase enzymes are controlled by the insulin/glucagon ratio. Ketone synthesis occurs when glucose concentration is low. Therefore Glucose Glucagon Lyase Ketones and vice versa The synthesis of ketone bodies requires both of the following: o Fatty acids to be available for oxidation in the liver following excessive lipolysis in adipose tissue – this supplies the substrate. o The plasma insulin/glucagon ratio to be low, usually due to a fall in plasma insulin – this activates the lyase and inhibits the reductase. Ketone bodies are important fuel molecules that can be used by all tissues containing mitochondria including the nervous system. The rate of utilisation is proportional to the plasma concentration. They are converted to acetyl~CoA and this is subsequently oxidised via stage 3 catabolism (TCA cycle).

Answer 12

Acetyl~CoA is produced by the catabolism of: o Fatty Acids o Sugars o Alcohol o Certain amino acids And can be oxidised via stage 3 of catabolism (TCA cycle). Acetyl~CoA is an important intermediate in lipid biosynthesis the major site of which is the liver (some in adipose tissue).

Answer 13

Type of Fue -> Weight (kg -> Energy Content (kJ) Triacylglycerol -> ~ 15 -> ~ 600,000 Glycogen -> ~ 0.4 -> ~ 4,000 Muscle Protein -> ~ 6 -> ~ 100,000

Answer 14

Glycogen Synthesis (glycogenesis) The pathway of glucose to glycogen involves a number of steps: 1. Glucose -> Glucose 6-Phosphte (catalysed by hexokinase and using ATP) 2. Glucose 6-Phosphate -> Glucose 1-Phosphate (catalysed by phosphoglucomutase) 3. Glucose 1-Phosphate + UTP + H2O -> UDP-Glucose + 2 Pi (UTP is structurally similar and energetically equivalent to ATP. UDP-Glucose is a highly activated form of glucose. Interconversion of glucose to galactose. 4. Glycogen (n residues) + UDP-Glucose -> Glycogen (n+1 residues) + UDP This irreversible reaction is catalysed by glycogen synthase adding non-branched (alpha 1,4 glycosidic bonds) subunits and branching enzyme adding branched subunits (Alpha 1,6 glycosidic bonds) about every 10 units. Glycogen Breakdown Glycogen is degraded in skeletal muscle in response to exercise and in the liver in response to fasting (or from stress response: “fight or flight”). The pathway is not a reversal of glycogen synthesis. The complete degradation of glycogen is shown by: Glycogen (n residues) + nPi -> 0.9n Glucose 6-Phosphate + 0.1n Glucose Glycogen is never degraded fully, a small amount of primer is always preserved. The Degradative pathway consists of the following steps: 1. Glycogen (n residues) + Pi -> Glucose 1-Phosphate + Glycogen (n-1 residues) The reaction is catalysed by glycogen phosphorylase that attacks the alpha 1,4 bonds. The bonds are subjected to phosphorolysis, not hydrolysis resulting in glucose residues released as glucose 1-phosphate rather than free glucose. (Glycogen phosphorylase doesn’t attack the alpha 1,6 branch points as this requires de-branching enzyme. De-branching enzyme produces free glucose) 2. Glucose 1-Phosphate -> Glucose 6-Phosphate (catalysed by phosphoglucomutase) The glucose 6-phosphate enters glycolysis and is used to provide energy for exercising muscle. Muscle glycogen represents a store of glucose 6-phosphate only used by muscle In the liver, glucose 6-phosphate is converted to glucose using glucose 6-phosphatase (absent in muscle): 3. Glucose 6-phosphate + H2O -> Glucose + Pi (catalysed by glucose 6-phosphatase) The glucose is released into the blood stream and transported to other tissues. Therefore, liver glycogen represents a glucose store that can be made available to all tissues of the body.

Answer 15

Liver Glycogen – Glucose store for all tissues of the body | Muscle Glycogen – Glucose 6-phosphate store, only used by muscle cells

Answer 16

Inherited glycogen metabolism disorders result from an abnormality in one or other of the enzymes of glycogen metabolism. 1. Glycogen phosphorylase 2. Phosphoglucomutase 3. Glucose 6-phosphatase (liver) Clinical severity depends on what enzyme/tissue is affected. o Increased/Decreased amounts of glycogen - Tissue damage if excessive storage - Fasting hypoglycaemia (low blood glucose) - Poor exercise tolerance o Glycogen structure may be abnormal o Usually liver and/or muscle are affected

Answer 17

Gluconeogenesis allows the production of glucose when carbohydrates are absent. This is for glucose-dependent tissues (E.g. CNS). Initially glucose will come from stores of glycogen but these stores can only last for 8-10 hours of fasting. The liver is the main site of gluconeogenesis. Possible substrates for gluconeogenesis - Pyruvate, lactate and glycerol can be converted to glucose - Essential and non-essential amino acids whose metabolism involves pyruvate or intermediates if the TCA cycle can be converted to glucose - Acetyl~CoA cannot be converted to glucose as PDH is irreversible The pathway of gluconeogenesis from pyruvate uses some of the steps of glycolysis. Overall: 2 Pyruvate + 4 ATP + 2 GTP + 2 NADH  Glucose + 2 NAD+ + 4 ADP + 2 GDP + 6 Pi + 2H+ Reversible steps of glycolysis are used in gluconeogenesis and irreversible bypassed. Steps 1 & 3 are by-passed by thermodynamically spontaneous reactions catalysed by phosphatases (glucose 6-phosphatase and fructose 1,6-bisphophatase): Glucose 6-phosphate + H2O  Glucose + Pi G = -ve Fructose 1,6-phosphate + H2O  Fructose 6-phosphate + Pi G = -ve Step 10 is by-passed by two reactions driven by ATP and GTP hydrolysis and catalysed by pyruvate carboxylase and phosphoenolpyruvate caroxykinase (PEPCK) respectively: Pyruvate + CO2 + ATP + H2O  Oxaloacetate + ADP + Pi + 2 H+ G = -ve Oxaloacetate + GTP + 2 H+  Phosphoenolpyruvate + GDP + CO2 G = -ve The last reaction provides a link between the TCA cycle and gluconeogenesis and enables the products of amino acid catabolism that are intermediates of the TCA cycle to be used to synthesise glucose. Regulation of Gluconeogenesis Gluconeogenesis is part of stress response, and is largely under hormonal control. The major control sites are PEPCK and Fructose 1,6-bisphosphonate. PEPCK Kinase activity is increased by – Glucagon, Cortisol is decreased by – Insulin Fructose 1,6-bisphosphonate activity is increased by – Glucagon is decreased by – Insulin Therefore, the insulin/anti-insulin ratio plays a major role in determining the rate of gluconeogenesis. In the absence of adequate levels of biologically effective insulin, (diabetes) increased gluconeogenesis rates contribute significantly to hyperglycaemia.

Answer 18

Triacylglycerols are the major dietary and storage lipid in the body. Consisting of three fatty acids (usually long: n=16) esterified to glycerol: Hydrophobic and stored in an anhydrous form in a highly specialised storage tissue (adipose tissue). Function largely as a store of fuel molecules (fatty acids, glycerol) for prolonged aerobic exercise, stress situations (e.g. starvation, pregnancy). Storage is controlled hormonally: o Storage promotion by insulin o Storage depletion activated by glucagon, adrenaline, cortisol, growth hormone and thyroxine.

Answer 19

The major dietary lipids are hydrolysed by pancreatic lipase in the small intestine to release glycerol and fatty acids. Glycerol Metabolism Glycerol derived from the hydrolysis of dietary triacylglycerols enters the blood stream and is transported in chylomicrons to adipose tissue to be stored as TAGs:

Answer 20

B-Oxidation of fatty acids The oxidation of fatty acids occurs via a sequence of reactions (B-oxidation pathway) that oxidises the fatty acid and removes the C2 unit (acetate). The shortened fatty acid is cycled through this reaction repeatedly removing a C2 unit each turn until only two carbons remain. The reaction sequence requires mitochondrial NAD+ and FAD. It cannot occur in the absence of oxygen since this is required for stage 4 (ox.p/ET) of catabolism to re-oxidise the NADH and FAD2H formed. There is no direct synthesis of ATP by the pathway. All the intermediates are linked to coenzyme A and the C-atoms of the fatty acid are converted to acetyl~CoA. Fatty Acid Synthesis (lipogenesis) Fatty acids are synthesised from acetyl~CoA (from carbohydrates/amino acids) at the expense of ATP and NADPH. The pathway occurs in the cytoplasm: 8 CH3CO~CoA + 7 ATP + 14 NADPH + 6 H+ goes to CH3(CH2)14COOH + 14 NADP+ + 8 CoA + 7 ADP + 7 Pi + 6 H2O NADPH is produced in the cytoplasm by the pentose phosphate pathway. Acetyl~CoA comes from the mitochondria when cleaved from citrate – releasing oxaloacetate and Acetyl~CoA. Most steps of the pathway are carried out by a multi-enzyme complex known as the fatty acid synthase complex. The fatty acids are built up sequentially from acetyl~CoA by a cycle of reactions that adds C2 per turn of the cycle to the growing fatty acid. The reactions therefore appear to act in the reverse of those in the B-Oxidation pathway, however this is not the case. The C2 units are added to fatty acid chains in the form of malonyl~CoA (a C3 compound) with the subsequent loss of CO2. Malonyl~CoA is produced from Acetyl~CoA by the enzyme acetyl~CoA carboxylase in a reaction that requires biotin: CH3CO~CoA + CO2 ATP -> CH2(COOH)CO~CoA + ADP + Pi (Acetyl~CoA carboxylase is not a component of the fatty acid synthase complex) It plays an important role in controlling the rate of fatty acid synthesis and can be regulated by: o Allosteric regulation (citrate activates and AMP inhibits) o Regulation by covalent modification of protein structure (Reversible phosphorylation/dephosphorylation) Insulin activates by promoting dephosphorylation (removing bulky PO4) Glucagon & Adrenaline inhibit the enzyme by promoting phosphorylationq

Answer 21

In a typical western diet more protein is eaten than is needed to supply the essential amino acids so excess amino acids are broken down in stage 2 catabolism: Catabolism of Amino Acids Each amino acid found in protein has it’s own pathway of catabolism – over 20 pathways. o They include an early step when the (-NH2) amino group is removed – transamination/deamination. This is converted to urea (CO(NH2)2) and excreted in the urine. o The remaining C-skeleton can be converted to one or more of the following: - Pyruvate - Oxaloacetate - Fumarate - -Ketoglutarate - Succinate - Acetyl~CoA o Amino acids that produce acetyl~CoA (e.g. leucine, lysine) are described as ketogenic as the acetyl~CoA can be used to synthesise ketone bodies. o Amino acids that produce the other products are described as glucogenic as they can be used for glucose synthesis by gluconeogenesis. o Some AAs are both ketogenic and glucogenic (isoleucine, threonine, phenylalanine) Transamination The enzymes involved are called aminotransferases and are specific for individual amino acids/similarly structured groups. o Most transaminases use ¬-ketoglutarate as the keto acid2 that is converted to glutamate. e.g. 1 Alanine aminotransferase (ALT) = glutamate-pyruvate transaminase (GPT) alanine + ¬-ketoglutarate  pyruvate + glutamate e.g. 2 Aspartate aminotransferase (AST) = glutamate-oxaloacetate transaminase (GOT) aspartate + ¬-ketoglutarate  oxaloacetate + glutamate o When oxaloacetate is used as keto acid2 it is converted to aspartate – an important intermediate in urea synthesis o Cortisol stimulates transaminase synthesis in the liver. Deamination L & D-amino acid oxidases are low specificity enzymes that convert amino acids to keto acids and NH3. Human liver cells have a high activity of D-amino acid oxidase. D-amino acids are found in plant and bacterial cell :. enter body through diet. They must not be used for protein synthesis as the proteins would be structurally abnormal and non-functional. The enzyme converts them to keto acids that are not optically active. Glutaminase is a high specificity enzyme that converts glutamine to glutamate + NH3 Glutamate dehydrogenase is a high specificity enzyme that catalyses: Glutamate + NAD+ + H2O  ¬-ketoglutarate + NH4+ + NADH + H+ It is important in amino acid metabolism by the liver as it is involved in both the disposal of amino acids (glutamate to ¬-ketoglutarate) and the synthesis of non-essential amino acids (¬-ketoglutarate to glutamate).

Answer 22

Phenylketonuria (PKU) is an inherited disorder in which the urine contains large amounts of phenylketones produced from phenylalanine. The first step in the metabolism of phenylalanine is its oxidation to tyrosine by the enzyme phenylalanine hydroxylase. This enzyme is defective in most PKU cases. As a result, phenylalanine accumulates in tissues and blood. It is metabolised by other pathways to produce various products including phenylpyruvate that is excreted in the urine. PKU is diagnosed by the detection of phenylketones in the urine or high phenylalanine blood concn (normal is

Answer 23

Creatinine is a breakdown product of Creatine. It is produced at a constant rate by a spontaneous reaction unless muscle is wasting (atrophy), or in a high protein diet. Creatinine is excreted via the kidneys into urine. The amount of excretion of 24hrs is proportional to the muscle mass of the individual. This provides a measure of muscle mass.

Answer 24

98.5% of ammonia in the body (pH 7.4) is in the form of the ammonium ion (NH4) Many tissues produce ammonia and it is also absorbed from the gut, being toxic it is normally rapidly detoxified and removed from the body. The peripheral blood concentration is normally kept very low (25-40M). Toxicity of ammonia The central nervous system is very sensitive to ammonia and hyperammonaemia is associated with blurred vision, tremors, slurred speech, coma and eventually death. The toxic effect of ammonia may involve its reaction with Alpha-ketoglutarate to form glutamate in mitochondria via glutamate dehydrogenase removing Alpha-ketoglutarate from the TCA cycle, which slows, disrupting the energy supply to brain cells. Also affects pH inside cells of the CNS and interferes with neurotransmitter synthesis/release. Ammonia detoxification Hyperammonaemia is seen in liver disease. Ammonia can be detoxified either by synthesis of N-compounds such as glutamine or by conversion to urea. Both urea and ammonia can be excreted from the body in the urine. Glutamine Synthesis – Glutamine is non-toxic (normal blood concentration 0.5mM) and can be synthesised from ammonia and glutamate via glutamine synthetase, requiring ATP. NH4+ + Glutamate + ATP -> Glutamine + ADP + Pi Glutamine is transported to the liver and kidney where it is hydrolysed by glutaminase releasing ammonia that is disposed of in the urine (kidney) and converted to urea (liver). Glutamine -> NH4+ + Glutamate Urea Synthesis Urea is very soluble in water, therefore is excreted in urine. It is non-toxic, metabolically inert and has a high nitrogen-content (47%) and so is a good way of disposing of unwanted nitrogen. Synthesis occurs in the liver by the urea cycle (5 enzyme series) and transported via the blood to the kidneys for excretion: HCO3- + NH4+ Aspartate + 3 ATP -> CO(NH2)2 + Fumarate + 2 ADP + AMP + 4 Pi NH2 groups of urea come from NH4+ and aspartate. NH4+ comes from the deamination of AA’s releasing NH3 as well as from NH3 produced by gut bacteria that enters the liver via the portal circulation. Aspartate is formed from oxaloacetate by transamination. Regulation of urea synthesis The enzymes of the cycle are not subject to feedback inhibition because the function of the cycle is to dispose of ammonia as urea. The enzymes of the cycle are inducible – they are induced by a high-protein diet, and repressed by low-protein/starvation. If treating starvation/low protein diet, gradual re-introduction to prevent hyperammonaemia. Inherited diseases of the urea cycle All defects cause: 1. Hyperammonaemia (high blood NH4+ concentration) 2. Accumulation and/or excretion of a particular urea cycle intermediate(s) Depends on the extent of the defect, the amount of protein eaten and includes vomiting, lethargy and irritability. There is usually mental retardation and in severe cases seizures, comas and eventually death. Treatment consists of a low protein diet and replacing the essential amino acids with keto acids that use up NH4+ when converted to amino acids, therefore lowering NH4+ concentration. Hyperammonaemia may also arise as a secondary consequence of liver disease such a cirrhosis where the liver’s ability to remove NH3 from the portal blood is impaired. Metabolic Fate of Urea Diffuses from the liver cells to the blood and is carried to the kidney where it is filtered and excreted in the urine. Some urea diffuses across the intestinal wall into the intestine, here bacteria breaks it down releasing ammonia that can be reabsorbed. In kidney failure where the concentration of urea in the blood is high, the production of ammonia from urea by the gut bacteria can contribute to the hyperammonaemia

Answer 25

Lipid Transport Various classes of lipids including triacylglycerols, fatty acids, cholesterol, cholesterol esters and phospholipids are normally found in blood. As they’re insoluble in water they’re carried in the plasma in association with protein. - ~98% carried as highly specialised non-covalent assemblies called lipoprotein particles - ~2% mostly fatty acids are carried bound non-covalently to albumin. The albumin bound fatty acids are fatty acids released from adipose tissue during lipolysis and are used as a fuel by tissues e.g. muscle. Albumin has limited capacity for fatty acids, therefore blood fatty acid levels do not normally exceed ~3mM. Plasma lipoproteins have great significance in medicine since disorders in the metabolism are associated with a number of important diseases E.g. atherosclerosis and coronary artery disease. Plasma Lipoprotein Particles Several classes of lipoproteins are found which differ from each other in the lipid being transported, the origins of the lipid and its destination. The protein components are apoproteins. Structurally, the apoproteins are involved in packaging non-water soluble lipids into soluble form. This is possible as they contain hydrophobic regions that interact with lipid molecules and hydrophilic regions that interact with water. Functionally, apoprotein molecules may be involved in the activation of enzymes or the recognition of cell surface receptors. The apoprotein composition of a lipoprotein particle determines its function. Lipoprotein Structure o Spherical particles that consist of a surface coat (shell) and a hydrophobic core. o The surface coat contains the phospholipids, cholesterol and apoproteins. o The hydrophobic core contains the triacylglycerol and cholesterol esters. o Lipoproteins are only stable if they maintain their spherical shape – this is dependant on the ratio of core to surface lipids. Therefore as the lipid from the hydrophobic core is removed the surface coat must be reduced. o Many components of the surface coat are free to transfer. The core components can only be removed by special proteins e.g. lipases and transfer proteins. Lipoprotein Functions Lipoprotein Transport Function Chylomicrons Transport dietary triacylglycerols from the intestine to tissues such as adipose tissue. VLDL Transport of triacylglycerols synthesised in the liver to adipose tissue for storage. LDL Transport of cholesterol synthesised in the liver to tissues. HDL Transport of excess tissue cholesterol to the liver for disposal as bile salts. Chylomicrons Dietary lipids absorbed from the intestine transported to tissues (e.g. adipose tissue) require Chylomicrons. Dietary triacylglycerols cannot be absorbed directly and are hydrolysed in the small intestine by the enzyme pancreatic lipase that release fatty acids and glycerol. The fatty acids enter the epithelial cells of the small intestine where they’re re-esterified back to triacylglycerols using glycerol phosphate from glucose metabolism. The triacylglycerols are then packaged with other dietary lipids (e.g. cholesterol, fat soluble vitamins) into chylomicrons. Chylomicrons are released from the epithelial cells into the blood stream via the lymphatic system and carried to tissues (e.g. adipose) that express extracellular enzyme lipoprotein lipase. The enzyme hydrolyses the triacylglycerols to release fatty acids that enter the cell where they’re converted to triacylglycerols for storage.

Answer 26

Lipoprotein Metabolism Lipoprotein Lipase is the enzyme responsible for removing the core triacylglycerols from lipoproteins such as chylomicrons and VLDLs. It’s attached to the inner surface of capillaries in tissues such as adipose tissue and muscle. Insulin increases the synthesis of the enzyme. The enzyme hydrolyses triacylglycerols in lipoprotein particles, releasing fatty acids and glycerol. Tissues then take up the fatty acids and the glycerols go to the liver. Lecithin:Cholesterol Acyltransferase (LCAT) restores the stability of lipoproteins. This is done by the conversion of surface lipid to core lipid. LCAT converts cholesterol to cholesterol ester using fatty acid derived from lecithin (phophatidylcholine). Deficiency of LCAT results in unstable lipoproteins of abnormal structure, therefore general failure of lipid transport. Lipid deposits occur in many tissues and atherosclerosis is a serious problem. LDL Metabolism Tissues obtain the cholesterol they need from LDLs by the process of receptor-medicated endocytosis. LDL particles are taken up by the cell and the cholesterol released inside the cell. All cells (except erythrocytes) are able to synthesise cholesterol from acetyl~CoA and could satisfy their requirements by biosynthesis. In practice, all cells appear to prefer the uptake of pre-formed cholesterol circulating in plasma lipoproteins. o Cells requiring cholesterol synthesise LDL receptors that are exposed on the cell surface. o These receptors recognise and bind to specific apoproteins (Apo B100) on the surface of the LDL. o The LDL receptor with its bound LDL is then endocytosed by the cell and subjected to lysosomal digestion. o Cholesterol esters are converted to free cholesterol that is released within the cell. o The cholesterol can be stored (as cholesterol esters) or used by the cell o This also inhibits the synthesis of cholesterol by the cell and reduces the synthesis and exposure of LDL receptors. This prevents the cell from accumulating too much cholesterol.

Answer 27

Type I Chylomicrons in fasting plasma No link to coronary artery disease Caused by defective lipoprotein lipase Type IIa Raised LDL Associated with coronary artery disease that may be severe Caused by defective LDL receptor Type IIb Raised LDL and VLDL Associated with coronary artery disease Defect unknown Type III Raised IDL and chylomicrons remnants Associated with coronary artery disease Caused by defective apoprotein (Apo. E) Type IV Raised VLDL Associated with coronary artery disease Defect unknown Type V Raised chylomicrons and VLDL in fasting plasma Associated with coronary artery disease Defect unknown Plasma lipoproteins are of great significance in medicine since disorders in their metabolism are associated with a number of important diseases including atherosclerosis and coronary artery disease. ``` Familial hypercholesterolaemia Familial hypercholesterolaemia (Type IIa hyperlipoproteinaemia) is a condition in which there may be an absence (homozygous) or deficiency (heterozygous) of functional LDL receptors. The condition is characterised by elevated levels of LDL and cholesterol in the plasma. Homozygotes develop extensive atherosclerosis early in life and heterozygotes develop the condition later in life. ```

Answer 28

Diet and lifestyle modifications (e.g. increase in exercise) are considered first to treat hyperlipoproteinaemia. Aiming to reduce/eliminate cholesterol from the diet and reduce the intake of triacylglycerols (especially those with saturated fatty acids). In some patients this will have little effect. Drug therapy – statins (e.g. simvastin) are a group of drugs that may lower plasma cholesterol by reducing the synthesis of cholesterol in tissues. The liver can dispose of cholesterol by converting it into bile salts and secreting a small amount directly in the bile. The bile salt sequestrants (e.g. cholestyramine) lower plasma cholesterol by increasing its disposal from the body. They act by binding to bile salts in the GI tract preventing them from being reabsorbed into the hepatic-portal circulation and promoting their loss in the faeces.

Answer 29

Mitochondria produce over 90% of a cell’s ATP via oxidative phosphorylation. This involves an electron transport chain, with the final destination for electrons being oxygen, which is then combined with protons to form water. However, some electrons (0.1 – 2%) are leaked from the chain (complexes 1 and 3). These electrons prematurely reduce oxygen to form superoxide radicals (O2-). These free radicals, with an unpaired electron, are highly reactive and are known as Reactive Oxygen Species (ROS).

Answer 30

ROS cause damage to cells that includes DNA, protein and membrane damage. The constant production of superoxide by mitochondria therefore needs to be handled. The enzyme Superoxide Dismutase (SOD) catalyses the reaction of superoxide radicals together, to form oxygen and hydrogen peroxide. However, hydrogen peroxide is itself a ROS. The enzyme Catalase therefore rapidly breaks it down into molecular oxygen and water. Hydroxyl Radicals can also be produced from Hydrogen Peroxide with the addition of iron ions (Fenton Reaction). These highly reactive agents also cause damage to cells, particularly membranes, but cannot be eliminated by an enzymatic reaction. Some cells of the immune system, such as neutrophils and monocytes when stimulated can rapidly produce a release of ROS, which is known as an oxidative burst. They do this by using the enzyme NADPH oxidase. In doing so the cell is usually destroyed, but surrounding bacteria or fungal cells are also destroyed. Oxidising agents can also be produced by ionising radiation (UV light, X-rays, -rays). Some chemicals also produce oxidising agents, e.g. Primaquine (anti-malarial) and Paraquat (banned herbicide). ``` Nitric Oxide (NO.) is also a free radical, produced from Arginine by the inducible enzyme nitric oxide synthase. Nitric Oxide and Oxygen react together to produce the free radical Peroxynitrite, which is involved in inflammation. ```

Answer 31

Cells have other defences against ROS apart from SOD and Catalase, which include NADPH and glutathione (GSH), antioxidant vitamins, flavenoids and minerals. NADPH is a reducing agent that is mostly produced by the pentose phosphate pathway. Glutathione (GSH) is a tripeptide that is usually abundant in cells and acts as an important antioxidant. The thiol (-SH) group in cysteine can donate its H and therefore act as a reducing agent. The oxidised form of GSH is glutathione disulphide (GSSG), and it is recycled via reduction by NADPH to deal with more ROS. These reactions are catalysed by the enzymes glutathione peroxidise and glutathione reductase.

Answer 32

Condition Cancer - DNA damaged by ROS Emphysema - Lung tissue destruction by ROS Pancreatitis - Pancreas damaged by ROS CV Disease - Lipid oxidation by ROS Crohn’s Disease - Bowel inflammation caused by ROS Rheumatoid Arthritis - Joint inflammation caused by ROS Diabetes Mellitus - (Type 1)  cell destruction by ROS Alzheimer’s Disease - Protein damage and misfolding by ROS The reaction of unsaturated lipids with ROS forms lipid peroxides. This damage to cell membranes is thought to be involved in the early stage of cardiovascular disease.

Answer 33

Pharmacodynamics – what a drug does to the body (remember, D’s) Pharmacokinetics – what the body does to the drug ``` Metabolites of drugs are usually less pharmacologically active with a few exceptions: - Primidone  Phenobarbitone - Pethidine  Norpethidine - Codeine  Morphine Essentially pharmacokinetics covers the: Absorption Distribution Metabolism Elimination… of a drug ADME ``` Phase I Most drug molecules are stable and relatively unreactive (a pro-drug) so in Phase I a reactive group is exposed on the parent molecule or added to the molecule. This generates a reactive intermediate that can be conjugated (in Phase II) with a water-soluble molecule to form a water-soluble complex. The most common chemical reactions in Phase I are oxidation, reduction and hydrolysis. The process requires a complex enzyme system called the cytochrome P450 (CYP) system and a high-energy cofactor, (NADPH). Some drugs already have a reactive group on their molecule so they can bypass Phase I. Morphine is a good example of this. Phase II The reactive intermediate from Phase I is conjugated with a polar molecule to form a water-soluble complex. The process is also known as conjugation. Glucoronic acid is the most common conjugate, as it’s an available by-product of cell metabolism. Drugs can also be conjugated with sulphate ions and glutathione. Phase II metabolism requires specific enzymes and a high-energy cofactor, uridine diphosphate glucuronic acid (UDPGA) First Pass Effect Substances absorbed from the lumen of the ileum enter the venous blood, which drains into the hepatic portal vein and is transported directly to the liver. Unfortunately the liver is the main site of drug metabolism as it contains all of the necessary enzyme systems, so any drug absorbed from the ileum may be extensively metabolised during this first pass through the liver – the first pass effect. E.g. 90% of an oral dose of paracetamol is usually metabolised by the first pass effect.

Answer 34

o There are about 50 different haem-containing enzymes in the cytochrome P450 system (CYP) and there are polymorphisms in the human population. o The isoform CYP3 A4 is the most important, accounting for ~55% of drug metabolism. o The CYP cofactor is NADPH

Answer 35

Genetic Factors We all differ slightly in the level of expression of metabolic enzymes. Therefore drug effects can vary from person to person. Some individuals may lack the gene that codes for a crucial enzyme (E.g. CYP3 A4) and this can have a large effect on drug metabolism. E.g. some people are described as slow acetylators, because they lack the main enzyme responsible for the acetylation reaction in Phase II. This has a large effect on the rate of metabolism of drugs requiring this pathway. E.g. some individuals have relatively low levels of pseudocholinesterase enzymes in the plasma, which affects their ability to metabolise drugs containing an ester bond, such as suxamethonium, a muscle relaxant used during anaesthesia. Environmental Factors If two drugs are given together to a patient then the metabolism of one drug may affect the metabolism of the other. Enzyme inhibition or induction can occur. Ethanol, nicotine and barbiturates are well known enzyme inducers.

Answer 36

Example 1 – Paracetamol Paracetamol is a widely available antipyretic drug. At therapeutic levels, paracetamol conjugates with glucuronide or sulphate in Phase II. If a toxic dose is taken these pathways become quickly saturated, paracetamol undergoes Phase I metabolism. This produces the toxic metabolite N-acetyl-p-benzo-quinone imine (NAPQI). Not only is this toxic to hepatocytes but it also undergoes Phase II conjugation with glutathione, which is an important anti-oxidant. Liver failure occurs over a period of several days. Example 2 - Alcohol The major site of alcohol metabolism is the liver. Alcohol  alcohol dehydrogenase  Acetaldehyde  aldehyde dehydrogenase  Acetate (low specificity enzyme) This complete oxidation requires NAD+ and forms reduced NADH The acetate is converted with ATPAMP + 2Pi to acetyl~CoA. Aldehyde Dehydrogenase has a low Km and so keeps the toxic Acetaldehyde to a minimum. With prolonged alcohol consumption Acetaldehyde can accumulate causing liver damage and the NAD+/NADH ratio and acetyl~CoA effect liver metabolism. CYP2E1 is an inducible enzyme that also metabolises alcohol, via oxidation. Low NAD+/NADH ratio NAD+ is used for: 1. Fatty acid oxidation 2. Conversion of Lactate  Pyruvate 3. Metabolism of Glycerol Accumulation of Lactate in the blood due to low NAD+ may cause lactic acidosis. It will also reduce the kidney’s ability to excrete uric acid. Therefore due to Uric acid levels, crystals of urate may accumulate in tissues producing the painful condition gout. Gluconeogenesis cannot be activated (NAD+/lactate use/glycerol use) and fasting hypoglycaemia becomes a serious problem. High Acetyl~CoA Acetyl~CoA cannot be oxidised due to the NAD+/NADH ratio and so synthesis of fatty acids and ketone bodies. The fatty acids are converted to triacylglycerols but cannot be transported because there’s a lack of lipoproteins. Therefore Fatty Liver develops. Sometimes the production of ketone bodies is enough to cause keto-acidosis Decrease in Liver Function (due to cell damage from toxic acetaldehyde) Damaged cells have a leaky plasma membrane and can lose enzymes such as transaminases and gamma glutamyl transpeptidase. Their appearance in the blood is an indicator of liver cell damage (liver function tests). Reduced liver function can result in: • Uptake of conjugate bilirubin  Hyperbilirubinaemia  Jaundice • Urea production  Hyperammonaemia and Glutamine • Protein synthesis  Albumin Clotting Factors Lipoproteins Serum Albumin may produce oedema Clotting factors increases blood-clotting time Lipoproteins causes lipid build up (liver main site of lipogenesis)  Fatty liver Direct/Indirect Effects of Alcohol Indirect - there is likely to be vitamin and mineral deficiencies and there may also be inadequate protein and carbohydrate uptake. Direct effect on GI tract - there is often loss of appetite/diarrhoea/impaired absorption of nutrients (Vit. K, folic acid, pyridoxine and thiamine) Vitamin deficiency symptoms are often seen in alcoholics. Thiamine deficiency can lead to Wernicke-Korsakoff syndrome with mental confusion and unsteady gait. Treatment of Alcohol Dependency Disulfiram is a drug that can be used as an addition in the treatment of chronic alcohol dependency. It is an inhibitor of the aldehyde dehydrogenase enzyme: If the patient drinks alcohol, Acetaldehyde accumulates in blood, giving ‘hangover’ symptoms.

Answer 37

Glucose/Glycogen Glucose can be used by all cells and is the preferred fuel. Only about 12g is present in solution in the body fluids, capable of supporting the CNS for ~2hours. More glucose (~300g) is stored as glycogen, in the liver and muscle. Only the glucose stored in the liver (~100g) can be made available to the CNS. ``` Fatty Acids/Ketone Bodies Many cells (excluding RBCs and those in the CNS) can also use fatty acids as fuels. Derived from triacylglycerol stored in adipose tissue. In a typical 70kg individual there is about 10-15kg of fat, enough to supply the body’s fuel needs for ~2 months. This makes up ~80% of the total fuel reserve. Fatty acids can be converted to ketone bodies in the liver to be used as fuel for the CNS when glucose low during starvation. ``` Protein/Amino Acids Protein in muscle (~6kg) can be broken down (proteolysis) to amino acids that can also be used to provide fuel in times of shortage, either by conversion to glucose and ketone bodies or by direct oxidation.

Answer 38

Blood glucose concentration is controlled via the endocrine system by regulating the rates of entry of glucose into the blood and removal from it. The Central Nervous System The CNS must receive adequate supply of glucose. Glucose has to be available at all times in the CNS (~140g/24hr) and other glucose-dependant tissues (40g/24hr) as metabolism proceeds at a constant rate throughout the day. Since glucose uptake by the CNS relates to the blood glucose concentration then to supply the CNS adequately then blood glucose concentration is maintained within a particular range (4.0-6.0mM) Hypoglycaemia When blood glucose 7.0mM a patient is classed as hyperglycaemic. The chronic effects of hyperglycaemia reduce both the quality and duration of life. Many systems including the nervous, cardiovascular and renal systems may be affected. Glucose appears in the urine as the renal threshold is exceeded. More water is lost in the urine as a result of the osmotic effects of the glucose ‘polyuria’ An increased thirst follows ‘polydipsia’ There is increased non-enzymatic glycosylation of plasma proteins such as lipoproteins that leads to disturbances in their function.

Answer 39

Effects of Insulin - Anabolic Increases glucose uptake and utilisation by muscle and adipose tissue Promotes storage of glucose as glycogen in the liver and muscle Promotes lipogenesis and storage of fatty acids as triacylglycerols in adipose tissues Promotes amino acid uptake and protein synthesis in liver and muscle Effects of Glucagon - Catabolic Gluconeogenesis to maintain supplies of glucose for the brain Glycogenolysis in the liver to maintain blood glucose for glucose-dependant tissues e.g. Brain Lipolysis in adipose tissue to provide fatty acids for use by tissues

Answer 40

Effects of Feeding The absorption of glucose, amino acids and lipids from the gut raises their blood concentration. The increases stimulate the endocrine pancreas to release insulin. Effects of Fasting As blood glucose levels falls insulin secretion is depressed. This reduces the uptake of glucose by adipose tissue and muscle. The falling blood glucose concentration also stimulates glucagon secretion i.e.  insulin/anti-insulin ratio.

Answer 41

Starvation Gluconeogenesis offset to spare protein 1. At first blood glucose falls, but is maintained at an adequate level (3.5mM) by the actions of glucagon, which stimulates the breakdown of hepatic glycogen. As these stores last only a few hours, the continued decrease of blood glucose stimulates the pituitary to release ACTH and consequently blood cortisol is elevated. This hormone acts to maintain blood glucose by stimulating gluconeogenesis and making gluconeogenic substrates available (mainly alanine and glycerol) by stimulating the breakdown of protein and fat. Glucagon also stimulates gluconeogenesis and the actions of both hormones involve amount/activity of key enzymes of the gluconeogenic pathway in liver cells. 2. Lipolysis occurs at a high rate due to insulin and lipolytic hormones e.g. glucagon/cortisol/growth hormone Free fatty acids in blood rise to about 2mM (normal ~0.3mM). The continuing action of cortisol stimulates fat breakdown and prevents most cells from using glucose, the fatty acids produced are preferentially metabolised. Glycerol then provides an important substrate for gluconeogenesis, reducing the need for breakdown of proteins. 3. Stimulated by the insulin/anti-insulin ratio, fatty acids are also oxidised in the liver to produce ketone bodies, which can replace glucose as a fuel for the brain. This further reduces the need for gluconeogenesis in the liver sparing protein. Ketone concentrations rise from 0.01mM in the fed state to 2-3mM after three days of starvation and 6-7mM after 1-2 weeks (physiological ketosis) As starvation continues: • The brain becomes able to use ketones as fuel, reducing its glucose requirement from 140g/day to 40g/day. • The kidneys begin to contribute to gluconeogenesis The brains use of ketones reduces the need for gluconeogenesis, after 4-5 weeks starvation gluconeogenesis has fallen to ~30% of that seen during the early period of starvation. Urinary nitrogen excretion initially about 12g/day (mostly urea) eventually falls to about 4g/day (approx. equal amounts of urea and NH4+) 4. The reduction in urea synthesis leads to decrease in the amount and activity of the enzymes involved in the process in liver cells. This is important for re-feeding a starved individual; a gradual increase of protein content is required.

Answer 42

Hormones are chemical messengers that travel via the bloodstream.

Answer 43

Hormones may be classified by chemical type: o Polypeptide hormones (largest group) – short or long chain(s) of amino acids E.g. Insulin, glucagon, growth hormone, placental lactogen o Glycoprotein hormones – large protein molecules, with carbohydrate side chains E.g. the ‘anterior pituitary hormones’ – luteinizing hormone (LH), follicle stimulating hormone (FSH), thyroid stimulating hormone (TSH) o Amino acid derivatives – small molecules synthesised from amino acids E.g. adrenaline (a catecholamine) and the thyroid hormones - thyroxine, tri-iodothyronine o Steroids – all derived from cholesterol E.g. cortisol, aldosterone, testosterone, oestrogen

Answer 44

Transport Polypeptide hormones, glycoprotein hormones and adrenaline are relatively hydrophilic and are transported in the bloodstream dissolved in the plasma. Steroid hormones and thyroid hormones are relatively hydrophobic (lipophylic) and need specialised transport proteins. Acting on the Target Cells Generally speaking, the effect that a hormone has on a target cell depends upon its concentration in the blood stream. Often lipophilic hormones (steroids and thyroid hormones) bind specifically or non-specifically to proteins in the blood and in this case it is the concentration of unbound or free hormone that matters. • If the physiologically effective concentration of the hormone in the blood is too low, the subject will suffer hormone deficiency. Example: Growth hormone secretion – failure to grow properly • If the physiologically effective concentration of the hormone in the blood is too high, then sign & symptoms of excess will occur. Example: Acromegaly; GH secretion – change shape of face and body and other metabolic effects. To change the activity of a target cell, hormones must interact chemically with the target cell. 1. Hormone binds to a specific, high affinity receptor on or in the cell. The location of the receptor in the target cell depends upon the chemical nature of the hormone: - Hormones that can cross cell membranes (i.e. lipophilic) bind to receptors inside the cells (cytoplasmic and/or nuclear). - Hormones that cannot readily cross cell membranes (i.e. hydrophilic) bind to receptors on the cell surface. 2. The binding hormone to a receptor triggers changes in the target cells, which may be in the activity of the enzyme/other proteins or in gene expression. When hormones bind to receptors on the cell surface a second messenger is often released within the cell, which goes on to influence the cell’s activity. - Target tissues usually respond rapidly to hormones that work by altering the activity of functional proteins e.g. enzymes, membrane transport proteins (seconds-minutes) - Target tissues responding to hormones that work by changing gene expression occurs over a longer time period (minutes-hours) and may even occur after the hormone concentration has returned to normal. Some hormones appear to have one major target tissue (e.g. TSH - thyroid gland) while others have a number of important target tissues. (e.g. insulin - liver/muscle/adipose tissue)

Answer 45

Negative Feedback Hormones are constantly lost from the circulation as they are excreted or broken down, therefore secretion rate must be adjusted to maintain an appropriate blood concentration. The rate of secretion of a hormone is usually controlled by negative feedback. The rate of secretion of the hormone is affected by the blood concentration directly, as soon as the concentration/the effect of the hormone falls below a critical level hormone secretion increases, until the correct level is achieved again. E.g. The pancreatic -cells secrete insulin. This acts on liver, muscle and adipose tissue to remove glucose from the blood. The -cells are directly sensitive to blood glucose concentration. If it rises above 5mM (i.e. following a meal) then insulin is secreted until the blood glucose concentration falls until it reaches the normal level again, when insulin secretion is switched off. One Hormone Controlling Another In many cases, the secretion of one hormone is controlled by another. The controlling hormone is known as the trophic hormone, these are mostly secreted by the anterior pituitary gland. E.g. secretion of cortisol from the cortex of the adrenal gland is controlled by adrenocorticotrophic hormone (ACTH), from the anterior pituitary. The anterior pituitary secretes (6 main) hormones: - Thyroid Stimulating Hormone  Thyrotrophin (TSH) – affects thyroid gland - Adrenocorticotrophic Hormone  Corticotrophin (ACTH) – affects adrenal gland - Growth Hormone  Somatotrophin (GH) – affects metabolism - Luteinizing Hormone (LH)  Affects ovary and testis function - Follicle Stimulating Hormone (FSH)  Affects ovary and testis function - Prolactin  Affects breast development and milk production TSH, ACTH, LH and FSH are trophic hormones controlling hormone production in endocrine gland. In some cases the concentration of the hormone they control affects the secretion by negative feedback. E.g. TSH – when the concentration of thyroid hormone in the blood gets too high less TSH is secreted, so thyroid hormone production falls. Conversely if thyroid hormone (T3/T4) concentration is too low, then more TSH is secreted, so more thyroid hormone is produced. Releasing or Inhibiting Hormones These hormones come from the nerves cells in the hypothalamus and travel to the gland via specialised blood vessels known as the hypophyseal portal vessels. This allows the brain to control hormone secretion. E.g.: Thyrotrophin Releasing Hormone (TRH) – stimulates TSH release Corticotrophin Releasing Hormone (CRH) – stimulates ATCH release Somatotrophin Releasing Hormones (SRH) – stimulates GH release Somatostatin – inhibits GH release Inactivation of Hormones This occurs in the liver and kidney and sometimes in target tissues. - Steroid hormones are inactivated by a relatively small change in chemical structure that increases their water solubility enabling them to be excreted from the body in the urine or via the bile. - Protein hormones undergo more extensive chemical changes and are degraded to amino acids that are reused.

Answer 46

Action of Insulin Insulin is stored in -cells storage granules as a crystalline-zinc complex. Dissolves in the plasma and circulates as a free hormone. Target tissue: liver, skeletal muscle and adipose tissue. Interacting with cell surface receptors and therefore stimulates enzymes/proteins inside the cell to act. ANABOLIC - affects carbohydrate, lipid and amino acid metabolism. Short term – clearing absorbed nutrients from the blood following a meal. Long term – effects on cell growth/cell division that relate to its ability to stimulate protein synthesis and DNA replication. Major actions on carbohydrates, lipids and amino acid metabolism are: -  Glucose transport into adipose tissue/skeletal muscle -  Glycogenesis and  Glycogenolysis in liver/muscle -  Gluconeogenesis in liver -  Glycolysis in liver/adipose tissue -  Lipolysis in adipose tissue - Lipogenesis and esterification of fatty acids in liver and adipose tissue -  Ketogenesis in liver -  Lipoprotein lipase activity in the capillary bed of tissues such as adipose tissue -  Amino acid uptake and protein synthesis in liver, muscle and adipose tissue -  Proteolysis in liver, skeletal muscle and adipose tissue Action of Glucagon Single chain polypeptide hormone, lacking disulphide bridges so has flexible 3D structure It takes up its active conformation on binding to its receptor on the surface of target cells. Synthesised as a larger precursor molecule (pre-proglucagon) that undergoes post-translational processing to produce the biologically active molecule. CATABOLIC Binds to a specific glucagon receptor in the cell membrane, a type of receptor termed a G protein-coupled receptor (GPCR). Binding to the receptor activates the enzyme adenylate cyclase, which increases cyclic AMP (cAMP) intracellularly. High levels of cAMP activate protein kinase A (PKA), which phosphorylates and :. activates important enzymes within the target cell. Major actions of glucagon are: - Glycogenolysis in liver -  Glycogenesis in liver - Gluconeogenesis in liver -  Ketogenesis n liver -  Lipolysis in adipose tissue

Answer 47

A typical islet (Islets of Langerhans) contains a number of cell types that produce different polypeptide hormones. The major cell types are -cells (~75%) that produce insulin and the -cells (~20%) that produce glucagon. Both these cell types store their hormonal products intracellularly in the membrane-limits vesicles (storage granules) prior to secretion – each cell may contain ~13,000. The cells have ultrastructural features characteristic of tissues that synthesise proteins for export – e.g. RER, well-defined Golgi,  Mitochondria and a system of microtubules and microfilaments.

Answer 48

Diabetes Mellitus is a group of metabolic disorders characterised my chronic hyperglycaemia due to insulin deficiency, insulin resistance or both. Type 1 This is more present in the teenage years (but the age related rate is otherwise similar up to old), there are substantially different rates between countries. There is a strong seasonal variation suggesting a link with a viral infection acting as a trigger. It’s likely that a genetic predisposition to the disease interacts with an environmental trigger to produce immune activation. This leads to the production of killer lymphocytes and macrophages and antibodies that attack and progressively destroy -cells (an auto-immune process). The genetic predisposition is associated with the genetic markers HLA DR3 and HLA DR4. Typically for type 1 - a lean young person with a recent history of viral infection who present a triad of symptoms: - Polyuria – excess urine production. Large quantities of glucose in the blood are filtered by the kidney, so not all of it is reabsorbed. The extra glucose in the nephron places an extra osmotic load on it, meaning that less water is reabsorbed to maintain osmotic pressure. - Polydipsia – thirst and drinking a lot, due to polyuria. - Weight loss - as fat and protein are metabolised because insulin is absent. Diagnosis of Type 1 Type 1 can be diagnosed by measurement of plasma glucose levels. Blood glucose is elevated because of the lack of insulin. The lack of insulin causes: -  Uptake of glucose into adipose tissue and skeletal muscle -  Storage of glucose as glycogen in muscle and liver -  Gluconeogenesis in liver The high blood glucose will lead to the appearance of glucose in the urine (glycosuria) and if not dealt with rapidly the individual will progress to a life-threatening crisis (diabetic ketoacidosis) Type 2 Type 2 is relatively common in all populations enjoying an affluent life-style. The estimated prevalence in the UK is about 2%, typically in older and often overweight. At diagnosis patients retain about 50% of their -cells and as these fall (ultimately to none) patients develop disorders of insulin secretion or insulin resistance so blood glucose is raised.

Answer 49

``` Type 1 Commonest type in the young. Characterised by the progressive loss of all or most of pancreatic -cells. Is rapidly fatal if not treated. Must be treated with Insulin. ``` Type 2 Affects a large number of usually older individuals. Characterised by the slow progressive loss of -cells but with disorders of insulin secretion and tissue resistance. May be present for a long time before diagnosis. May not initially need treatment with Insulin, but all do eventually.

Answer 50

Type 1 People can be found with the relevant HLA markers and auto-antibodies but without glucose or insulin abnormalities. They may then develop impaired glucose tolerance, then diabetes (sometimes initially diet controlled) before becoming totally insulin dependant. Type 2 People can be found with insulin resistance then as insulin production fails they develop impaired glucose tolerance. Finally they will develop diabetes that can initially be controlled by diet, then tablets, then insulin. If the process continues long enough they may lose all insulin production. Diagnosis of Diabetes Diabetes is diagnosed in the presence of symptoms i.e. polyuria, polydipsia and unexplained weight loss plus: - A random venous plasma glucose concentration > 11.1 mmol/l or - A fasting plasma glucose concentration > 7.0 mmol/l (whole blood > 6.1 mmol/l) or - Plasma glucose concentration > 11.1 mmol/l 2 hours after 75g anhydrous glucose in a oral glucose tolerance test (OGTT) With no symptoms diagnosis should not be based on a single glucose determination but requires confirmatory venous plasma glucose determination. At least one additional glucose test result on another day with a value in the diabetic range is essential, either fasting, from a random sample or from the two hour post glucose load. A diagnosis of diabetes has important legal and medical implications for the patient and it is therefore essential to be secure in the diagnosis and an appropriate laboratory must undertake the plasma glucose measurement.

Answer 51

High rates of -oxidation of fats in the liver coupled to the low insulin/anti-insulin ratio leads to the production of huge amounts of ketone bodies (such as acetoacetone, acetone and -hydroxybutyrate). Acetone, which is volatile may be breathed out, and smelt on the patient’s breath. The H+ associated with ketones produce a metabolic acidosis – ketoacidosis. The features of ketoacidosis are: prostration, hyperventilation, nausea, vomiting, dehydration and abdominal pain. Ketoacidosis is a very dangerous condition. It is most important to test for ketones in the urine when assessing diabetes control.

Answer 52

Hypoglycaemia A diabetic can become hypoglycaemic (plasma glucose 10 mmol/l. Symptoms include polyuria, polydipsia, weight loss, fatigue, blurred vision, dry or itchy skin, poor wound healing. Plasma proteins may become glycosylated, affecting their functions LO 8.6 Describe, in broad outline, the principles of management of diabetes Type 1 diabetes cannot be cured and must be managed for the rest of the patient’s life. Insulin is used to treat Type 1 diabetics and comes cases of Type 2. It must be injected subcutaneously as it’s a peptide hormone that can be digested in the stomach. Patients must be educated to treat themselves at appropriate times with appropriate doses so as to mimic as closely as possible the behaviour of pancreatic islets in controlling blood glucose. On occasion, if the patient has an infection or suffered a trauma, insulin dosage needs to be increased or there is a risk of ketoacidosis. The social and psychological implications are huge, and the degree of control achieved by patients can be very variable. Dietary management and regular exercise are vital components of the treatment regime. The management of blood glucose requires frequent blood glucose measurement. A small amount of blood from a finger prick is sufficient to measure blood glucose using the BM stick and reader. There is always a risk that blood glucose will fall too low – hypoglycaemia – so both patients and their associates need to be aware of the signs and symptoms of hypoglycaemia, which can occasionally be fatal unless treated with glucose either my mouth or by infusion. Type 2 diabetes can sometimes be managed by diet or by “oral hypoglycaemic” drugs such as sulphonylureas that increase insulin release from the remaining -cells, and reduce insulin resistance and particularly metformin that reduce gluconeogenesis.

Answer 53

Persistent hyperglycaemia is associated with the abnormal metabolism of glucose to products that may be harmful to cells. Uptake of glucose into cells of tissues such as peripheral nerves, the eye and the kidney does not require insulin and is determined by the extracellular glucose concentration. Therefore, during hyperglycaemia the intracellular concentration of glucose in these tissues increases and glucose is metabolised via the enzyme aldose reductase which catalyses the reaction: Glucose + NADPH + H+  Sorbitol + NADP+ This reaction depletes NADPH and leads to increased disulphide bond formation in cellular proteins altering their structure and function. The accumulation of sorbitol causes osmotic damage to cells. Glycosylation Hyperglycaemia is also associated with increased non-enzymatic glycosylation of plasma proteins (e.g. lipoproteins) that leads to disturbances in their function. Glucose reacts with free amino groups in proteins to form stable covalent linkages. The extent of glycosylation depends on the glucose concentration and the half-life of the protein. Glycosylation changes the net charge on the protein and the 3D structure of the protein, therefore affects the function of the protein. Glycosylated Haemoglobin Glucose in the blood will react with the terminal valine of the haemoglobin molecule to produce glycosylated haemoglobin (HbA1c). The percentage of HBA1c is a good indicator of how effective blood glucose control has been. As RBCs normally spend ~3 months in the circulation the %HbA1c is related to the average blood glucose concentration over the preceding 2-3 months. Poorly controlled diabetics can have a HbA1c value above 10%.

Answer 54

Macrovascular complications include: o  risk of stroke o  risk of myocardial infarction o Poor circulation to the periphery – particularly the feet Microvascular complications include: o Diabetic eye disease: Changes in the lens due to osmotic effects of glucose (glaucoma) and possibly cataracts. More important problem is diabetic retinopathy – damage to blood vessels in the retina, which can lead to blindness. Damages blood vessels may leak and form protein exudates on the retina or they can rupture and cause bleeding in the eye. New vessels may form (proliferative retinopathy) - these are weak and can easily bleed. o Diabetic kidney disease (nephropathy): This is due to damage to the glomeruli, poor blood supply because of change in kidney blood vessels, or damage from infections of the urinary tract (more common in diabetics – excess glucose for bacteria to thrive). An early sign of nephropathy is a protein in the urine (microalbuminuria). o Diabetic neuropathy: Damage to the peripheral nerves which directly absorb glucose causing changes of or loss of sensation, and changes due to alteration in the function of the autonomic nervous system. o Diabetic feet: Poor blood supply, damage to nerves and increase risk of infection all sum up to make the feet of a diabetic vulnerable. In the past, loss of feet through gangrene was not uncommon. Care is needed to keep feet in good condition.

Answer 55

The appetite centre is located in the arcuate nucleus in the hypothalamus. This is a group of neurones consisting of two types: - Primary neurones - sense metabolite levels/respond to hormones - Secondary neurones - synthesise input from 10 neurones and co-ordinate a response via the vagus nerve. Primary Neurones can further be sub-divided into: - Excitatory – stimulate appetite via the release of neuropeptide Y (NPY) and agouti-related peptide (AgRP). - Inhibitory – suppress appetite by releasing pro-opiomelanocortin (POMC) POMC is a polypeptide prohormone which can be enzymatically cleaved to produce: - -endorphin - Adrenocorticotrophin hormone (ACTH) - -Melanocyte stimulating hormone (-MSH) -MSH acting on melanocortin 4 receptors is involved in suppressing appetite. There is also a feedback system. In response to the stomach being filled, POMC is released in the brain to suppress appetite. With this -endorphin (from the POMC) gives feelings of euphoria and tiredness.

Answer 56

Ghrelin I a peptide hormone released from the wall of the empty stomach, which activates the stimulatory neurones in the arcuate nucleus - appetite. Stretch of the stomach wall caused by food intake inhibits Ghrelin release. Leptin is a peptide hormone released into the blood by adipocytes in fat stores. Leptin acts by stimulating inhibitory neurones and inhibiting stimulatory neurones in the arcuate nucleus. Leptin acts as a feedback mechanism from the body’s fat stores – a lack of leptin/insensitivity to leptin has been associated with obesity. Leptin induces the expression of uncoupling proteins in mitochondria, which leads to the production of heat rather than ATP. PYY is a peptide hormone released from the wall of the small intestine appetite. Insulin hormone involved in short/long term regulation of body weight. Insulin appetite via the same mechanism as leptin (however leptin seems to be more important in this role). Insulin resistance in associated with obesity and often leads to Type 2 Diabetes. Amylin is a peptide hormone secreted with insulin from the -cells, known to appetite, glucagon secretion and slow gastric emptying. Pramlintide is undergoing trials as a hypoglycaemic agent in early Type 2 Diabetes.

Answer 57

Metabolic syndrome arose from the observation of a common pattern of symptoms in obese people. It is defined as: “A group of symptoms including insulin resistance, dyslipidaemia, glucose intolerance and hypertension associated with central adiposity. The co-occurrence in the same individual of a number of cardiovascular risk factors such as dyslipidaemia and hypertension, usually in association with overweight or obesity and a sedentary life style is known as the ‘metabolic syndrome’” BMJ 2003;327:61-2 The major factors that predispose to insulin resistance are obesity and a sedentary life-style. Insulin resistance is associated with the development of a dyslipidaemic profile (VLDL LDL HDL) that is highly atherogenic. It is also associated with a risk of hypertension. The WHO criteria for metabolic syndrome are: - Central obesity with a waist:hip ratio >0.9 (men) >0.85 (women) - BMI above 30kg/m2 - Blood pressure > 140/90 mmHg - Triglycerides > 1.7 mmol/L - HDL cholesterol 7.8 mmol/L - Glucose uptake during hyperinsulinaemic euglycaemic clamp in lowest quartile for population. Metabolic syndrome is controversial and has not been universally accepted in the medical profession, although it is gaining ground in terms of acceptance. (see pages 29-30 of Metabolism Workbook 2)

Answer 58

Barker Hypothesis, from David Barker. A study (and then repeated studies) showed that the incidence of adult diseases such as coronary heart disease, hypertension and Type 2 Diabetes are related to low birth weight. This suggests that the experience of the foetus in utero during development somehow determines the future health of the individual. It was discovered that biochemical adaptation took place in the foetus according to the supply of nutrients via the placenta and that these adaptations were ‘programmed in’ for adult life. Further research showed that the programming involved the switching on/off of genes at critical times during foetal development. One puzzling feature was that low birth weight could apparently be passed down generations. An explanation for this has been provided by epigenetics: ‘an epigenetic trait is a stably inherited phenotype resulting from changes in a chromosome without alterations in the DNA sequence’ From research with rodents, the mechanism appears to involve methylation of DNA at crucial positions and altering the histone structure causing suppression of gene transcription targeting the promoter region of specific genes.

Answer 59

The gland is located in the neck, anterior to the lower larynx and upper trachea. It is inferior to the thyroid cartilage. The recurrent laryngeal and the external branch of the superior laryngeal nerves lie close to the thyroid. It is highly vascularised with three arteries and veins supplying and draining it – superior, middle and anterior thyroid arteries/veins. Two lateral lobes joined by a central isthmus. 2-3cm across and weighs 15-20g making it one of the largest endocrine glands. Two major cells types are found in the gland: - Follicular cells - arranged in units called follicles separated by connective tissue. The follicles are spherical and are lined with epithelial (follicular) cells surrounding a central space (lumen) containing protein - colloid. - Parafollicular (C-cells) - found in the connective tissue.

Answer 60

The thyroid produces 3 hormones: o Thyroxine (T4) produced in the follicular cells o Tri-idothyronine (T3) produced in the follicular cells - T3 and T4 are small molecules derived from tyrosine with the addition of iodine. - T3 is the active form of the hormone. - T4 is much more stable, so much more T4 is released and then converted to T3 o Calcitonin produced in the parafollicular cells - Calcitonin is a polypeptide hormone involved in calcium metabolism. Synthesis of T3 & T4 o Transport of iodide into the epithelial cells against a concentration gradient (coupled with 2 Na+ ions – Sodium Iodide Symporter) o Synthesis of a tyrosine rich protein (thyroglobulin) in the epithelial cells o Secretion (exocytosis) of thyroglobulin into the lumen of the follicle o Oxidation of iodide to produce an iodinating species o Iodination of the side chains of tyrosine residues in thyroglobulin to form: - MIT (monoiodotyrosine) - DIT (di-iodotyrosine) o Coupling of DIT with MIT or DIT to form T3 & T4 respectively within the thyroglobulin. o T3:T4 production in the ratio of ~1:10 Storage T3 & T4 are stored extracellularly in the lumen of the follicles as part of the thyroglobulin molecules. The amounts stored (T3=~0.4moles T4=~6moles) are considerable and would last several months at normal rates of secretion. Secretion of T3 & T4 Thyroglobulin is taken into the epithelial cells from the lumen of the follicles by the process of endocytosis. Here proteolytic cleavage of thyroglobulin occurs to release T3 & T4, which diffuse from the epithelial cells into circulation. Transport of T3 &T4 T3 & T4 are hydrophobic molecules, therefore are bound to proteins Thyroxine binding globulin (TBG), pre-albumin and albumin

Answer 61

T3 & T4 Secretion is controlled by the hypothalamus and anterior pituitary gland. Thyrotrophin-Releasing Hormone (TRH) is a tri-peptide released from cells in the dorsomedial nucleus of the hypothalamus. This is influenced by T3 & T4 (negative feedback). Stress TRH Temperature TRH TRH travels in the hypothalamic/pituitary portal system to stimulate secretion of Thyroid Stimulating Hormone (TSH) from the thyrotrophs in the anterior pituitary. TSH travels in the blood to affect the follicular cells. Thyroid Stimulating Hormone (TSH) is a glycoprotein consisting of two non-covalently linked subunits ( & -subunits). It’s released in low-amplitude pulses following a circadian rhythm - during the night  early morning. TSH interacts with receptors on the surface of the follicle cells and stimulates (all aspects of the) synthesis and secretion of T3/T4. TSH has trophic effects on the gland that result in vascularity, size/number of follicle cells. These trophic effects can lead to an enlarged thyroid (goitre) that may be over/underactive.

Answer 62

Effects of Thyroid hormones on the body T3 and T4 increase the metabolic rate of many tissues: - Increased Glucose uptake and metabolism - Stimulate mobilization and oxidation of fatty acids - Stimulate protein metabolism Since the metabolic effects of T3 and T4 are mainly catabolic, this leads to: - Increased BMR - Increased Heat Production (UCPs) - Increased O2 consumption T3 and T4 are important for normal growth/development - T3 and T4 directly affect bone mineralisation and increase the synthesis of heart muscle protein. - The CNS is sensitive to T3/T4, especially during development as they’re required for the development of cellular processes of nerve cells, hyperplasia of cortical neurons and myelination of nerve fibres. In the absence of thyroid hormone from birth-puberty the child remains mentally & physically retarded (cretinism). If the deficiency isn’t corrected within a few weeks of birth there’s irreversible damage. All newborns have their thyroid function assessed soon after birth. In adults lack of thyroid hormones is characterised by poor concentration and memory, lack of initiative. - T3 and T4 are also indirectly related to interactions with hormones and neurotransmitters. T3/T4 stimulate hormone and neurotransmitter receptor synthesis in tissues (i.e. heart muscle, GI) that can increase responsiveness of these tissues to regulatory factors. - In heart muscle – tachycardia. In the GI tract – increased motility. - T3 and T4 have a permissive role in the actions of hormones such as FSH and LH. - Ovulation fails to occur in the absence of thyroid hormones. Mechanism of action of Thyroid hormones T3 & T4 act within the target cell, interacting with high-affinity (10x greater) receptors located in the nucleus and possibly mitochondria. - Binding of T3 to the hormone-binding domain (is thought to) produce a conformational change in the receptor that unmasks the DNA-binding domain. - Interaction of the hormone-receptor complex with DNA increases the rate of transcription of specific genes that are then translated into proteins. - The rate of protein synthesis stimulates oxidative energy metabolism in the target cells to provide the extra energy required for protein synthesis. - Also, protein synthesis produces more specific functional proteins, therefore increases cell activity and demand for energy. Conversion of T4 to T3 T4 can be converted to T3 by the removal of the 5’-iodide. This helps to regulate the amount the amount of active (free) hormone in cells, as T3 is 10x more active than T4. Removal of the 3’-iodide produces the inactive reverse T3 (rT3)

Answer 63

Hypothyroidism Hashimoto’s disease Affecting ~1% of the population, mostly women. An autoimmune disease resulting in: a) The destruction of follicles or b) The production of an antibody that blocks the TSH receptor on follicle cells Patients are generally treated with oral thyroxine – (Over treat  Hyperthyroidism) Other Causes of Hypothyroidism - Post-surgery - Radioactive Iodine - Anti-thyroid drugs - Secondary (Lack of TSH) - Congenital - Iodine deficiency Signs and symptoms of hypothyroidism in adults: - Cold intolerance and reduced BMR - Weight gain - Tiredness and lethargy - Bradycardia (abnormally slow HR) - Neuromuscular system – weakness, muscle cramps and cerebellar ataxia (clumsiness) - Skin dry and flaky - Alopecia (hair loss) - Voice is deep and husky Hyperthyroidism Grave’s disease Affecting ~1% of the population, mostly women. An autoimmune disease, in which antibodies are produced that stimulate TSH receptors on follicle cells resulting in increased production/release of T3 and T4. Patients may be treated with carbimazole that inhibits the addition of iodine into thyroglobulin. Other Causes of Hyperthyroidism - Toxic (overproducing T3/T4), multinodular goiter - Excessive T4/T3 therapy - Excess iodine – amiodarone - Thyroid carcinoma (99% don’t cause hyper/hypothyroidism) - Ectopic thyroid tissue Signs and symptoms of hyperthyroidism: - Heat intolerance, increased oxygen consumption and increased BMR - Weight loss - Physical and mental hyperactivity - Tachycardia (increased HR >100) - Intestinal hyper-mobility (see effects of thyroid hormones on body) - Exopthalmos - Skeletal and cardiac myopathy giving rise to tiredness, weakness and breathlessness - Osteoporosis due to increased bone turnover and preferential resorption

Answer 64

``` Hyper-thyroidism BMR & Catabolic Activity - UP Sympathetic & CNS Activity - UP (GI Tract, CNS) Direct effects on tissues - CVS Free T4 - UP TSH - DOWN Hyperthyroidism Most common form is Graves Disease Autoimmune Antibody that stimulates TSH receptors Treated with carbimazole Heat Intolerance Increased BMR and O2 consumption Hyperactivity (mental and physical) Tachycardia (increased HR > 100) Goitre Exopthalmos Intestinal hyper-mobility Skeletal and cardiac myopathy Osteoporosis (Inc. turnover, osteoclasts > osteoblasts) ``` ``` Hypo-thyroidism BMR & Catabolic Activity - DOWN Sympathetic & CNS Activity - DOWN (GI Tract, CNS) Direct effects on tissues - CVS (SUBCUTAENOUS) Free T4 - DOWN TSH - UP Most common form is Hashimoto’s Disease Autoimmune Antibody that blocks TSH receptors Destruction of follicles Treated with oral thyroxine (T4) Cold Intolerance Reduced BMR Tiredness and lethargy Bradycardia (abnormally slow HR) Goitre Weakness, muscle cramps and cerebellar ataxia Skin dry and flaky Alopecia Voice is deep and husky ```

Answer 65

Calcium plays a critical role in many cellular processes: - Hormone secretion - Nerve conduction - In/activation of enzymes - Muscle contraction - Exocytosis Therefore, the body very carefully regulates the plasma concentration of free ionised calcium ([Ca2+]), the physiologically active form of the metal, and maintains free plasma [Ca2+] within a narrow range (1.0 to 1.3mM, or 4.0 to 5.2mg/dl). In plasma, calcium exists as: - Free ionised species - Bound to/associated with anionic sites on serum proteins (especially albumin) - Complexed with low-molecular-weight organic anions (e.g. citrate and oxaloate) Phosphate – part of the adenosine triphosphate molecule, therefore plays a crucial role in cellular energy metabolism and in the activation and inactivation of enzymes. Unlike calcium the plasma phosphate concentration is not strictly regulated and fluctuates during the day (i.e. after meals). Calcium and phosphate homeostasis are linked because: - Calcium and phosphate are the major components of hydroxyapatite crystals [Ca10(PO4)6(OH)2], which constitute the major portion of the mineral in bone. - They’re regulated by the same hormones, primarily parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D (calcitriol) and, to a lesser extent, the hormone calcitonin. These hormones act on; the bone, the kidneys and the gastrointestinal tract to control levels of these two ions in plasma. The actions of these hormones on calcium and phosphate are opposed

Answer 66

There are two key hormones involved in the regulation of serum calcium: - Parathyroid Hormone (PTH) - Vitamin D (Active form, Calcitriol) These both raise serum calcium concentrations. There is a third hormone Calcitonin, which in animals lowers serum calcium but suggested only to preserve the maternal skeleton during pregnancy (i.e. serum calcium/osteoclast acitivity)

Answer 67

Changes in Ca2+ concentration alter PTH by negative feedback. Chief cells have unique G-protein calcium receptors on the cell surface. Increased Ca2+ binds to the G-protein receptors and stimulates Phospholipase C (PLC) inhibiting adenylate cyclase, which leads to reduced cAMP and reduced PTH release. High Ca2+   PLC   Adenylate Cylase   cAMP   PTH Reverse occurs when Ca2+ is low.

Answer 68

Vitamin D is formed in the skin or absorbed in the gut from the diet. As it has a short T½, it is converted to Calciferol (25-hydroxyvitamin D) in the liver (T½~2 weeks) Vitamin D is not regulated – it’s final conversion is in the Kidney to Calcitriol is regulated by PTH

Answer 69

Vitamin D is synthesised in the skin in the presence of sunlight. PTH regulates calcitriol (the active form of Vitamin D). Calcium levels regulate PTH with negative feedback. Vitamin D2 -> Absorbed by Gut -> Nil (Prohormone) Vitamin D3 -> Skin(UV light) -> Nil (Prohormone) Calciferol ->Liver(1st Hydroxylation of Vit D) -> Nil (Prohormone) Calcitriol Kidney -> (2nd Hydroxylation of Vit D) -> increased Ca2+ absorption (Binds to Ca2+ in the Gut) Parathyroid Hormone -> Parathyroid Gland -> Conversion of Calciferol -> Calcitriol Ca2+ release from bone Ca2+ reabsorption in kidney

Answer 70

PTH affects tubular cells within the kidney, increasing Ca2+ reabsorption in the distal convoluted tubule (DST). Pi is removed from circulation by inhibition of Kidney proximal tubule (PT) reabsorption, this prevents calcium stone formation.

Answer 71

Hypocalcaemia Results in hyper-excitability in the nervous system, including the neuromuscular junction, leading to paraesthesia (tingling sensation), then tetany (involuntary muscle contraction), paralysis and even convulsions. This is due to the low amount of Ca2+ bound to the NMJ membrane, allowing Na+ to depolarise it much more readily. Hypercalcaemia – MOANS, GROANS, STONES May result in the formation of kidney stones (renal calculi), constipation, dehydration, kidney damage, tiredness and depression. Parathyroid hormone related peptide (PTHrP) is a peptide hormone produced in tumours, which may lead to Hypercalcaemia. The measurement of PTHrP can be of assistance in determining the cause of an otherwise unexplained hypercalcaemia. PTHrP is secreted by some cancer cells leading to humeral hypercalcaemia of malignancy (HHM). Commonly in patients with breast/prostrate cancer and occasionally with myeloma. PTHrP is similar to PTH; calcium release from bone, renal calcium excretion and renal phosphate reabsorption. However PTHrP does not increase C-1 hydroxylase activity and therefore does not increase calcitriol concentration.

Answer 72

Anterior Pituitary o TSH is produced in the Thyrotrophs (see session 9 – the Thyroid gland) o ACTH produced in the Corticotrophs o Growth hormone produced in the Somatotrophs (largest number of cells) o LH and FSH are produced in the Gonadotrophs o Prolactin is produced in the Lactotrophs Adrenal Gland Cortex: There are 3 zones of the Adrenal Cortex, and 3 types of steroid hormone secreted: ``` o Zona Glomerulosa  Mineralocorticoids E.g. Aldosterone (C21 steroid) o Zona Fasciculata  Glucocorticoids E.g. Cortisol and Corticosterone (C21 steroids) major steroids produced o Zona Reticularis  Androgens E.g. Testosterone (C19 steriod) ``` ``` Medulla: o Adrenaline (epinephrine) ``` Action/Mechanism of ACTH ACTH is hydrophilic and interacts with high affinity receptors on the cell surface in the zona fasiculata and reticularis. The binding of ACTH to specific receptors - a type of melanocortin receptor (type 2), known as MC2 or corticotropin receptor (this receptor uses cAMP as a secondary messenger) - leads to activation of cholesterol esterase increasing the conversion of cholesterol esters to free cholesterol. It also stimulates other steps in the synthesis of cortisol from cholesterol. Actions of Cortisol Cortisol is an important component of the stress response and it has a number of important effects on metabolism. In the starved and stressed states it affects the availability of all major substrates by increasing proteolysis, lipolysis and gluconeogenesis. The metabolic actions of cortisol include:  Amino acid uptake,  Protein synthesis &  Proteolysis (not in liver)   Amino acids  Hepatic Gluconeogenesis and Glycogenolysis   Glucose  Lipolysis in Adipose tissue   Fatty acids (N.B. high levels of cortisol  lipogenesis in adipose tissue) Peripheral uptake of glucose (anti-insulin) In addition it also has direct effects on cardiac muscle, bone and the immune system. Aldosterone Stimulates Na+ reabsorption in the kidney in exchange for K+ (or H+).  Over secretion of Aldosterone increases Na+ and water retention and loss of K+ causing hypertension and muscle weakness.  Under secretion of Aldosterone does the opposite causing hypotension. Androgens Stimulate the growth and development of male genital tract and male secondary sexual characteristics including height, body shape, facial and body hair, and lower voice pitch. They also have anabolic actions especially on muscle protein.  Over secretion of adrenal androgens produces effects in the female that include: hair growth (hirsuitism), acne, menstrual problems, and virilisation, increased muscle bulk, deepening voice. Oestrogens Stimulate growth and development of female genital tract, breasts and female secondary characteristics including broad hips, accumulation of fat in breasts and buttocks, body hair distribution. They are weakly anabolic and decrease circulating cholesterol levels.

Answer 73

``` Structure of Cortisol Cortisol is a member of the C21 steroid family that differ from other steroids in: - The number of C-atoms - Presence of functional groups - Distribution of C=C double bonds ``` All the steroid hormones are lipophilic and must be transported bound to plasma proteins (~90% transcortin, ~10% free and active) are synthesised from cholesterol via progesterone in a series of enzyme-catalysed reactions.

Answer 74

Mechanism of Action of Cortisol upon its Target Cells (Not to be confused with mechanism of ACTH that stimulates cortisol secretion) Cortisol can cross the plasma membranes (lipophilic/hydrophobic) of target cells and bind to cytoplasmic receptors. The hormone/receptor complex then enters the nucleus to interact with specific regions of DNA. This interaction changes the rate of transcription of specific genes and may take some time. Mechanism of Action of Adrenaline upon its Target Cells Adrenaline does not cross cell membranes; instead it binds to adrenoreceptor on the outside of the cell. A secondary messenger then affects cell activity.

Answer 75

Adrenocorticotrophic Hormone (ACTH or Corticotrophin) secreted from corticotrophs of the anterior pituitary is the main factor controlling the release of cortisol. The secretion of ACTH is under the control of Corticotrophin Releasing Hormone (CRH) – a peptide produced in the Hypothalamus. CRH is secreted in response to physical (temperature/pain), chemical (hypoglycaemia) and emotional stressors. There is also negative feedback by Glucocorticoids on both the Hypothalamus and Pituitary i.e. on both ACTH and CRH

Answer 76

Catecholamine synthesis The catecholamines are synthesised in a series of enzyme-catalysed steps that convert Tyrosine to Dopa and then Dopamine. Dopamine is then converted to Noradrenalin (neurotransmitters dopamine & noradrenaline) and noradrenaline to Adrenaline by methylation. The catecholamines are stored in the medullary cells in vesicles. Tyrosine  Dopa  Dopamine  Noradrenaline  Adrenaline Actions of Adrenaline Adrenaline is released as part of the fright, flight or fight response in man and it is secreted in response to stress situations. It has effects on: - Cardiovascular System ( Cardiac Output,  Blood supply to muscle) - Central Nervous System ( Mental alertness) - Carbohydrate Metabolism ( Glycogenolysis in liver and muscle) - Lipid Metabolism ( Lipolysis in adipose tissue) ``` Clinical consequences of over-secretion of adrenaline Overproduction by the adrenal medulla, usually due to a tumour (Phaemochromocytoma), may be associated with: o Hypertension o Palpitations o Sweating o Anxiety o Pallor o Glucose intolerance ```

Answer 77

Adrenocorticotrophic Hormone (ACTH or corticotrophin) secreted from corticotrophs of the anterior pituitary is the main factor controlling the release of cortisol. The secretion of ACTH is under the control of Corticotrophin Releasing Hormone (CRH) – a peptide produced in the Hypothalamus. CRH is secreted in response to physical (temperature/pain), chemical (hypoglycaemia) and emotional stressors. There is also negative feedback by Glucocorticoids on both the Hypothalamus and Pituitary i.e. on both ACTH and CRH

Answer 78

ACTH is a single chain polypeptide hormone, the initial biosynthetic precursor is a large protein called pro-opiomelanocortin (POMC). Post-translational processing of POMC at different sites produces biologically active peptides including ACTH, MSH (Melanocyte Stimulating Hormone) and endorphins. The MSH sequence is contained within the ACTH sequence in POMC giving ACTH some MSH-like activity when present in excess. The clinical consequences of over-secretion of ACTH (this would occur when there is low/no negative feedback from cortisol to the anterior pituitary – Addison’s disease) relate to the direct effects of ACTH on tissues (increased pigmentation due to partial MSH activity) and the effects of ACTH on the adrenal cortex.

Answer 79

Mechanism of Action of Cortisol (Not to be confused with mechanism of ACTH that stimulates cortisol secretion) Cortisol can cross the plasma membranes (lipophilic/hydrophobic) of target cells and bind to cytoplasmic receptors. The hormone/receptor complex then enters the nucleus to interact with specific regions of DNA. This interaction changes the rate of transcription of specific genes and may take some time.

Answer 80

Hyperactivity – Increased Secretion of Glucocorticoids Cushing’s Syndrome May be due to: - Increased activity of the adrenal cortex due to a tumour (adenoma) - Ectopic secretion of ACTH (Uncontrolled  glucocorticoids secretion - no negative feedback loop with tumour) Clinical Effects of Excess Cortisol Secretion Signs and symptoms may include: o Increased muscle proteolysis and hepatic gluconeogenesis that may lead to hyperglycaemia with associated polyuria and polydipsia (“steroid diabetes”) o Increased muscle proteolysis leads to wasting of proximal muscles, producing thin arms and legs and muscle weakness. o Increased lipogenesis in adipose tissue leading to deposition of fat in abdomen, neck and face and producing characteristic body shape, moon-shaped face and weight gain. o Purple striae on lower abdomen, upper arms and thighs reflecting the catabolic effects on protein structures in the skin and leading to easy bruising because of thinning of skin and subcutaneous tissue. o Immunosupressive, anti-inflammatory and anti-allergenic reactions of cortisol leading to increased susceptibility to bacterial infections and producing acne. o May be back pain and collapse of ribs due to osteoporosis caused by disturbances in calcium metabolism and loss of bone matrix protein. o Mineralocorticoid effects of excess cortisol may produce hypertension due to sodium and fluid retention ``` Deficiency Excess Low Glucose Weight Loss Nausea Hypotension High Glucose Weight Gain Increased Appetite Hypertension Cushingoid ``` Hypoactivity – Decreased activity of the adrenal cortex Addison’s Disease May be due to: - Diseases of the adrenal cortex (auto-immune destruction) reduce glucocorticoids and mineralocorticoids. - Disorders in pituitary or hypothalamus that lead to decreased secretion of ACTH or CRH – only affects glucocorticoids. Clinical effects of too little cortisol secretion Too little cortisol secretion, caused by autoimmune destruction of the adrenal gland, would also involve the loss of mineralocorticoids producing a complex situation that may present as: Acute emergency (Addisonian crisis) Chronic debilitating disorder (Addison’s Disease) o Insidious onset with initial non-specific symptoms of tiredness, extreme muscular weakness, anorexia, vague abdominal pain, weight loss and occasional dizziness. o Extreme muscular weakness and dehydration o A more specific sign is the increased pigmentation, particularly on the exposed areas of the body, points of friction, buccal mucosa, scars and palmar creases due to ACTH-mediated melanocyte stimulation (see LO 11.2) o Decreased blood pressure due to sodium and fluid depletion o Postural hypotension due to fluid depletion o Hypoglycaemia episodes especially on fasting -  cortisol  catabolic stimulation These effects may be exacerbated by stress such as trauma or severe infection and lead to nausea, vomiting, extreme dehydration, hypotension, confusion, fever and even coma (Addisonian crisis). This constitutes a clinical emergency that must be treated with intra-venous cortisol and fluid replacement (dextrose in normal saline) to avoid death.

Answer 81

Measure ACTH/Cortisol Levels Measurement of plasma cortisol and ACTH levels and the 24hr (circadian rhythm) urinary excretion of cortisol and its breakdown products (17-hydroxysteroids) are important in investigating suspected adrenocortical disease. Dexamethasone Suppression Test Dexamethasone is a potent synthetic steroid that, when given orally would normally suppress (by feedback inhibition) the secretion of ACTH and therefore cortisol. Dexamethasone suppression of plasma cortisol by > 50% is characteristic of Cushing’s disease as even though the diseased pituitary is relatively insensitive to cortisol it does retain some sensitivity to potent synthetic steroids. Suppression does not normally occur in adrenal tumours or ectopic ACTH production. Synacthen Test The administration of Synacthen (a synthetic analogue of ACTH) intramuscularly, would normally increase plasma cortisol by >200 nmol/l. A normal response usually excludes Addison’s disease.

Answer 82

Steroid hormone receptor homology The steroid receptors form part of a family of nuclear DNA-binding proteins (steroids pass through the cell membrane acting in the cell) that include the thyroid and vitamin D receptors. They all have three main regions: - A hydrophobic hormone-binding region - A DNA-binding region rich in cysteine and basic amino acids - A variable region There is sequence homology in the hormone-binding region of the receptors. The percentage homology of the hormone-binding region of the glucocorticoid receptor with receptors is: - Mineralocorticoid is ~64% - Androgen is ~62% - Oestrogen is ~31% - Thyroid is ~24% Therefore, cortisol will bind to the mineralocorticoid and androgen receptors with low affinity. This binding may become significant when high levels of the hormone are present.

Answer 83

A typical net weight gain by the end of pregnancy is ~8kg as the mother supplies everything required for the growth of the foetus (nutrients, vitamins, minerals, oxygen and water). These requirements increase as growth proceeds and exert an ever-increasing impact on maternal metabolism. The rate of transfer of nutrients across the placenta to the foetus is dependant on their concentration gradient Mother:Foetus (like that of glucose conc.CNS). The environment in which the foetus develops is controlled by maternal metabolism This changes as pregnancy proceeds to ensure that: - The foetus is supplied with the range of nutrients it requires - These nutrients are supplied at the appropriate rate for each stage of development - This is achieved with minimal disturbances to maternal nutrient homeostasis - The foetus is buffered from any major disturbances in maternal nutrient supply The metabolism of all major maternal nutrients is affected during pregnancy. These changes are long-term adaptive responses of maternal metabolism that are hormonally mediated. The hormones involved are: Maternal insulin It’s concentration in the maternal circulation increases as pregnancy proceeds and it acts to promote the uptake and storage of nutrients, largely as fat in maternal adipose tissue. Foetal-placental hormones These become increasingly important as pregnancy proceeds, they largely oppose the actions of insulin i.e. they are “anti-insulin”. They maintain the glucose concentration gradient to ensure it’s constant supply. Metabolic changes during the first half of pregnancy The changes in the first 20 weeks are related to a preparatory increase in maternal nutrient stores (especially adipose tissue) ready for the more rapid growth of the foetus, birth and subsequent lactation. Increasing levels of insulin (insulin/anti-insulin ratio) promote an anabolic state in the mother that results in  nutrient storage. Metabolic changes during the second half of pregnancy This is characterised by a marked increase in growth of the placenta and foetus. Maternal metabolism adapts to meet an increasing demand by the foetal-placental unit for nutrients as well as meeting requirements of maternal tissues. The foetal-placental unit demands are met by keeping the concentration of nutrients in the maternal circulation relatively high. By: • Reducing the maternal utilisation of glucose by switching tissues to fatty acids • Delaying the maternal disposal of nutrients after meals • Releasing fatty acids from the stores built up during the first half of pregnancy Changes in maternal metabolism are controlled by changes in insulin/anti-insulin ratio. Maternal insulin levels continue to increase but the production of the anti-insulin hormones by the foetal-placental unit increases at an even faster rate, therefore the insulin/anti-insulin ratio falls producing the required metabolic changes. Maternal Ketogenesis The marked decrease in the insulin/anti-insulin ratio during the second half of pregnancy affects maternal ketogenesis (insulin/anti-insulin ratio = ketogenesis). The increased availability of fatty acids to the liver resulting from the mobilisation of maternal adipose, coupled with the fall in the insulin/anti-insulin ratio switches on the production of ketone bodies by the maternal liver. These are used to fuel the developing foetal brain.

Answer 84

Maternal insulin is a major factor in controlling the metabolic response to pregnancy. The rate of secretion of insulin (both basal and stimulated) normally increases as pregnancy proceeds. The ability of -cells to meet this increased demand for insulin secretion is achieved by -cell hyperplasia and hypertrophy as well as the increased rate of insulin synthesis in the -cell. Gestational Diabetes In some women, the endocrine pancreas is unable to respond to the metabolic demand of pregnancy and the pancreas fails to release the increased amounts of insulin required. As a consequence there is a loss of control of metabolism, blood glucose increases and diabetes results (Gestational Diabetes). After birth, when the increased metabolic demands of pregnancy are removed and hormone levels change, the endocrine pancreas can respond adequately and the diabetes disappears. Women who experience gestational diabetes are more likely to develop over diabetes later in life.

Answer 85

The metabolic response to exercise ensures: - The increased energy demands of skeletal and cardiac muscle are met by mobilisation of fuel molecules from energy stores - There are minimal disturbances to homeostasis by keeping the rate of mobilisation equal to the rate of utilisation - The glucose supply to the brain is maintained (prevent hypoglycaemia) - The end products of metabolism are removed as quickly as possibly The nature/extent of the metabolic response depends on: - Type of exercise (muscles used) - Intensity and duration of exercise - Physical condition and nutritional status of the individual - During high intensity activities of short duration skeletal muscle works under anaerobic conditions as the supply of oxygen to the muscle is inadequate for aerobic metabolism. - During lower intensity activities of longer duration the oxygen supply to muscle is adequate to allow aerobic metabolism. Energy Requirments ATP, C~P, substrate level phosphorylation The energy for muscle contraction comes from the hydrolysis of ATP: ATP + H2O  ADP + Pi + energy (RE: sliding filament theory) The ATP concentration does not fall by more than 20% as it is regenerated from ADP. Initially it is regenerated from the creatine phosphate (C~P) present in muscle: Creatine~P + ADP  ATP + Creatine Next, ADP must be rapidly converted back to ATP by coupling it to the oxidation of fuel molecules (substrate level phosphorylation). Fuel Molecules Glycogen The glycogen stores of muscle could provide the muscles with enough energy - Under aerobic conditions for ~60min of low intensity exercise - Under anaerobic conditions, where the end product is lactic acid for ~2min This huge difference reflects the different amounts of ATP produced - 33moles of ATP/mole of glucose as glycogen under aerobic conditions - 3 moles of ATP/mole of glucose as glycogen under anaerobic conditions The liver stores glycogen, however this store of glucose is required to prevent hypoglycaemia and the associated impairment of CNS function. Using muscle glycogen is advantageous: - Availability not affected by blood supply - No need for membrane transport into muscle cells - Produces G-6-P without using ATP Glycogen + Pi  Glucose 1-Phosphate  Glucose 6-Phosphate In the liver G6-P is converted, catalyse by Glucose 6-phosphatase, to Glucose whereas the enzyme is absent in muscle and G6-P enters glycolysis. [see LO 4.5]) - Mobilisation can be very rapid – highly branched structure allows many sites for enzyme attack and glycogen phosphorylase activity can be changed rapidly by a mixture of covalent modification (phosphorylation) and allosteric activation (ADP and Ca2+). Limiting the anaerobic metabolism of glucose in muscle is from the build-up of lactate and H+. The accumulation of H+ is so dramatic (2 moles H+/mole of glucose) that it exceeds the buffering capacity of the muscle cells and impairs their function producing fatigue. Mechanisms of muscle impairment by H+ include: - Inhibition of glycolysis by H+ - H+ interferes with actin/myosin interaction - H+ causes sarcoplasmic reticulum to bind calcium (inhibits contraction) Triacylglycerol Triacylglycerol stores can provide muscles with fatty acids, the oxidation of which (under aerobic conditions) would provide sufficient energy for ~48 hours of low intensity exercise. There are a number of factors that limit the use of fatty acids in muscle. These include: - Rate of fatty acid release from adipose tissue (rate of lipolysis) - Limited capacity of the blood to transport fatty acids (requires albumin) - Rate of fatty acid uptake into muscle cells and into cell mitochondria - Fatty acid oxidation required more oxygen/mole of ATP produced than glucose - Fatty acids can only be metabolised under aerobic conditions Metabolic Response to Short Duration High Intensity Exercise Confined to skeletal muscle that works anaerobically and is controlled by the nervous system (noradrenaline) with some input endocrine systems (adrenaline). Metabolic response includes: - Muscle ATP and C~P are use initially (~5 secs) - Muscle glycogen is rapidly mobilised to provide glucose 6-phosphate (~5sec) - Glucose 6-P is metabolised via glycolysis to provide ATP from ADP by substrate level phosphorylation. - Glycolysis is carried out under anaerobic conditions as oxygen supply to muscle is inadequate for aerobic metabolism - Dramatic increase in rate of anaerobic glycolysis (1,000 times) produces lactate and H+ (max. rate ~20 mmol of H+/second are produced) - Build up of H+ produces fatigue Metabolic Response to Medium Duration Intensity Exercise The body regenerates ATP by ~60% aerobic and ~40% anaerobic metabolism of glycogen. The body must eliminate a large amount of CO2 but no major problem with H+ as it can be buffered. Three phases to the race: - The initial sprint which uses ATP, C~P and anaerobic glycogen metabolism. - A long middle phase, which ATP is produced aerobically from glycogen in muscle. This relies on an adequate O2 supply. - A finishing burst, which relies on the anaerobic metabolism of glycogen and produces lactate. Metabolic Response to Long Duration Low Intensity Exercise The carbohydrate stores in the body are insufficient to provide enough energy to complete the distance and muscle cells have to oxidise fatty acids. The metabolic changes are more gradual and involve several tissues. The major features of the metabolic response are: - The muscle work aerobically (supply of oxygen increased by cardiovascular response) and can use all types of fuel molecules (not just glucose) - The origin and type of fuel changes as exercise proceeds Control of these changes is largely hormonal (insulin, adrenaline, growth hormone, glucagon and Cortisol) with some input from the nervous system (noradrenaline). The fuels used are: - Initially muscle glycogen, if this was the sole source it would last ~60mins aerobically. Many runners try to prolong this period by eating carbohydrate rich diets to increase their glycogen stores – this is most effective after exercise as exercise promotes the storage of glucose as muscle glycogen rather than as lipids. - Increased utilisation of circulating blood glucose by muscles. The blood glucose concentration stays relatively constant. The glucose used is replaced with glucose from the Liver (~75% glycogen store, ~25% gluconeogenesis). There are limited substrates for liver gluconeogenesis – eventually blood glucose will fall. - Due to the aerobic conditions, muscle cells are able to use fatty acids. This utilisation increases with time.

Answer 86

Body composition changes (adipose reduced and muscle increased ) Glucose tolerance improves (muscle increased) -increased glucose transport proteins in cell membrane (GLUT 4) Insulin sensitivity of tissues increased -increased skeletal muscle vascularisation? Blood Triacylglycerol decrease (VLDL & LDL decreased , HDL increased) Blood pressure falls (vascularisation?) Psychological effects – feeling of “well-being”