Alanine released from muscle in fasting

Even in the fasting state there is a small uptake of glucose by muscle, but much of the glucose uptake will presumably have come from red blood cells, as will the output of lactate.

There is a considerable output of glucose by the liver. Some of this will come from liver glycogen, but much will come from gluconeogenesis after an overnight fast.

There will be a small amount of lactate, mainly from red blood cells, although resting muscle takes up glucose and metabolises it to some extent anaerobically, putting out lactate. However, the main substrates for gluconeogenesis in the fasting state are amino acids.

The diagram shows the arterial - femoral vein differences in concentrations of amino acids

(From data reported by Wahren J. et al Journal of Clinical Investigation 51: 1870 1972)

In most cases the arterio-venous difference is negative, showing that there is an output of most amino acids form muscle in the fasting state, although it takes up moderate amounts of serine and cysteine.

The output of alanine is 3-fold higher than that of any other amino acid, amounting to ~60% of the total amino acid output.

Where have these amino acids come from?

They must have come from catabolism of muscle protein.

The diagram below shows the amino acid composition of muscle protein.

Although the output of alanine by muscle in fasting is 3-fold higher than any other amino acid, this does not reflect the amino acid composition of muscle proteins. Alanine accounts for ~60% of the total amino acid output from muscle, yet it accounts for only 10% of the amino acids in muscle proteins.

The diagram below shows the arterial - hepatic vein differences in concentrations of amino acids

The liver takes up most amino acids in the fasting state (although it puts out small amounts of valine, leucine and aspartate). Again the uptake of alanine is considerably greater than that of any other amino acid, and alanine accounts for more than half the total amino acid uptake by the liver.

The structures of alanine, pyruvate and lactate are shown on the right.

There is a family of enzymes, transaminases, that transfer the amino group from an amino acid onto a keto-acid, so forming the amino acid corresponding to that amino acid, and leaving the keto-acid corresponding to the amino acid substrate. The keto-acid corresponding to alanine is pyruvate, which, as we have already seen, is a substrate for gluconeogenesis.

In the liver the substrate keto-acid is commonly ketoglutarate, forming glutamate. This glutamate can then undergo oxidative deamination to reform ketoglutarate and ammonium ions. The ammonium is then used for the synthesis of urea, which is excreted.

Note that although compounds such as pyruvate and ketoglutarate (and also the carbon skeletons of other amino acids) are commonly referred to as keto-acids, correctly they should be called oxo-acids. The C=O group is an oxo-group, and these compounds are not chemically ketones.

It is unfortunate that oxo-acids are more commonly called keto-acids, because this often causes confusion between these compounds and the ketone bodies (acetoacetate, hydroxybutyrate and acetone) which are formed from fatty acids in the fasting state in the liver to provide metabolic fuel for muscle and (in prolonged starvation) the brain. Elevated blood concentrations of ketone bodies leads to keto-acidosis, as seen in poorly controlled diabetes mellitus and prolonged starvation.

The reactions of transaminases are freely reversible, so that alanine can readily be formed in muscle from pyruvate. The amino donors are various of the amino acids released by catabolism of muscle protein, and their keto-acids are metabolised in muscle as metabolic fuel.

This leaves us with the problem of where the pyruvate comes from to form alanine to be exported from muscle.

Many endurance athletes (marathon runners, racing cyclists, etc) practice what is know as carbohydrate loading. In preparation for an event they exercise to exhaustion, which occurs when muscle glycogen is more or less completely depleted, then consume a large meal that is high in carbohydrate, commonly a meal of pasta, providing as much as 60 - 70% of energy. Sometimes they consume a low carbohydrate diet (less than 10% of energy) to deplete muscle glycogen for 2 - 3 days before the event, rather than exercising to exhaustion.

The large intake of carbohydrate leads to rapid synthesis of glycogen, and increases the total amount of glycogen in muscle, so permitting them to continue for longer before becoming exhausted.

In a study of carbohydrate loading, volunteers underwent a muscle biopsy to measure glycogen, then exercised to exhaustion, and consumed a meal of pasta that also contained [13C]glucose. As expected, this led to an increase in muscle glycogen, which was labelled with 13C.

Blood samples were taken at intervals for the next 24 hours, during which they were allowed water but no food.

The 13C label in blood glucose fell over the first 2 - 5 hours after the meal, then began to rise again.

The fall in labelling of blood glucose reflects the utilisation of glucose for glycogen synthesis. The rise after about 5 hours of fasting reflects the mobilisation of liver glycogen to maintain the blood concentration of glucose. Remember that muscle cannot release glucose from its glycogen stores because it lacks glucose 6-phosphatase (see the exercise on Fasting hypoglycaemia in an infant - and poor exercise tolerance in two brothers).

After about 18 hours of fasting, There was significant labelling of plasma alanine with 13C .

This suggests that the carbon skeleton of the alanine put out by muscle in the fasting state has come from glucose 1-phosphate released from muscle glycogen. We can therefore draw up a glucose-alanine cycle, in which muscle glycogen is metabolised by glycolysis in the muscle, but the pyruvate is neither oxidised to acetyl CoA (as occurs in aerobic metabolism) nor reduced to lactate (as occurs in anaerobic glycolysis), but undergoes transamination to alanine. This alanine is taken up by the liver and used for gluconeogenesis.

There is an inter-organ glucose-alanine cycle.

In this way, although muscle cannot release glucose from its glycogen reserves, it can indirectly provide a source of glucose for other tissues when liver glycogen is more or less exhausted.

For more on this topic see: Felig, P. The glucose-alanine cycle. Metabolism 22(2): 179-207 (1973).

Key points from this exercise:

• Although most of the body's glycogen is in muscle, this cannot provide a direct source of blood glucose in the fasting state because muscle lacks glucose 6-phosphatase. (In an adult the liver contains ~ 90 g of glycogen in the fed state is and the muscle ~245 g).
• In the fasting state muscle puts out amino acids that have arisen form catabolism of muscle protein. Alanine is about 60% of total amino acid output, yet it accounts for only 10% of the total amino acids in muscle proteins.
• In the fasting state liver takes up alanine from the bloodstream and uses it as a substrate for gluconeogenesis.
• Alanine undergoes transamination to yield pyruvate; the amino group is transferred onto ketoglutarate, forming glutamate, which then undergoes oxidative deamination to reform ketoglutarate and release ammonium ions, which are used for synthesis of urea for excretion.
• Muscle glycogen yields glucose 1-phosphate, which undergoes glycolysis to pyruvate. Pyruvate then undergoes transamination to yield alanine, which is exported to the liver. The amino groups come from the amino acids released by catabolism of muscle proteins. The carbon skeletons of the amino acids (the keto-acids) are metabolised as metabolic fuel in muscle.