History of glucagon
Glucagon, like insulin, was named before it was discovered. Its existence was first suspected when crude insulin extracts were seen to cause a brief rise in plasma glucose before the subsequent fall. This was postulated to be due to the presence of another pancreatic hormone, christened the "glucose agonist", abbreviated to glucagon. From the point of view of the insulin chemists, this was a contaminant, only finally eliminated by the mid-century. Chemists at Eli Lilly finally succeeded in isolating glucagon in the 1950s. Glucagon is however more than a dancing partner for insulin, since (according to the bihormonal hypothesis), type 2 diabetes is characterised as much by glucagon excess as by insulin deficiency.
The discovery of glucagon
In 1869 the medical student Paul Langerhans identified small isolated clumps of cells scattered throughout the tissue of the exocrine pancreas, but did not appreciate their significance. The islets, which make up 1-2% of the mass of the pancreas (average weight 70-100 grams), were named in his honour in the 1890s. By this time many investigators already guessed that they were the source of an unknown substance that regulated the amount of glucose produced by the liver. In 1909 de Meyer, a Belgian physician, proposed the name insulin (the islet hormone) for this unknown substance, which was not isolated successfully until 1921-2.
When insulin was first injected it was noted that glucose rose initially before falling again. This, as we now know, was because crude pancreatic extracts contain glucagon as well as insulin. The existence of a second pancreatic hormone was first postulated in 1923 when Kimball and Murlin were studying methods of insulin extraction from the pancreas and found that acetone precipitated a fraction soluble in 95% alcohol, a procedure which separated the unknown substance from insulin. Injection of this largely insulin-free fraction into dogs and rabbits caused a rapid rise in glucose. They inferred from this that the preparation contained a second pancreatic hormone, and named this the glucose agonist, hence glucagon.
Attempts to study this further were limited by difficulties of extraction, and were mainly driven by the efforts of the insulin manufacturers to identify and remove this contaminant (known as the H-G or hyperglycaemic-glycogenolytic factor) from their products. This was thought to have been achieved by the crystallization of insulin by John Jacob Abel in 1926, and the H-G factor was long dismissed as a contaminant. The question was re-examined after World War 2, when the H-G factor was found to be present in all insulin preparations other than those produced by Novo.
The work was taken up by Earl Sutherland and Christian de Duve at the Lilly Laboratories, who found that the H-G factor was present in the pancreas and gastric mucosa of the dog, with traces in the jejeunum and ileum. It was also present in fetal pancreas, and was not destroyed by alloxan treatment of experimental animals which was known to destroy the insulin-producing beta cells. From this they inferred that it was produced by the alpha cells. They speculated that the new factor might be a second hormone involved in glucose metabolism, but felt that such speculation was premature until the substance was shown to be secreted into the blood stream and to be involved in glucose metabolism. They were puzzled by the presence of the H-G factor in the mucosa of the upper part of the stomach (in the dog, but not in other species examined) and elsewhere in the gut.
The name glucagon was reintroduced to replace the H-G factor (probably by de Duve in 1953). By the early 1950s chemists working with Eli Lilly had succeeded in crystallizing and partially characterizing a pure form of glucagon, which then became available for physiological investigation, and later as an injectable treatment for hypoglycaemia.
Where did glucagon come from? Investigators in the early part of the century had were able to distinguish two sub-populations of cells in the pancreatic islets named alpha and beta cells. The dominant beta cell population was linked to insulin secretion and (as noted above) the alpha cells were tentatively identified as the source of glucagon in the 1950s, later confirmed by immunohistochemical staining for the two hormones. These two cell populations constitute the majority of cells within the pancreatic islet.
Later study has identified further cell types: delta cells that produce somatostatin, PP cells that produce pancreatic polypeptide, and epsilon cells that produce ghrelin. These three cell types constitute 5-10% of the islet content. In humans, beta cells constitute ~82% of the mass of the healthy dorsal pancreas, which constitutes 90% of the pancreatic mass. The smaller ventral pancreas (10% of total mass) is largely composed of PP cells. Alpha cells make up ~13% of the human islets in the dorsal pancreas, and the other three cell types make up the remaining ~5%.
The development of radioimmunoassays for glucagon in the 1960s opened up exciting possibilities for investigators, but it was soon appreciated that substances produced in the wall of the intestine were interfering with the assay. This led, by degrees, to the discovery of a range of glucagon-like peptides produced outside the pancreas, and so to identification of a precursor molecule – proglucagon – within which the glucagon sequence was embedded. Later studies, described elsewhere, finally elucidated the molecular biology of proglucagon and its daughter hormones. This in turn led on to discovery of the glucagon-like peptides GLP 1 and GLP2.
The bihormonal hypothesis
The first practical application of glucagon injection was as a safe and effective way of reversing insulin-induced hyperglycaemia, but the discovery of a glycogenolytic hormone which antagonised the actions of insulin led to early speculations as to its possible role in the pathogenesis of diabetes. The development of radioimmunoassay and special staining techniques for insulin and glucagon next made it clear that type 2 diabetes was not just a disease of insulin deficiency, but also of glucagon excess. This was demonstrated by the presence of elevated glucagon levels in the plasma and by increased prominence of alpha cells in the pancreatic islets, although it was not clear whether this reflected true alpha cell hyperplasia or declining numbers of beta cells.
These observations led Roger Unger to propose in 1975 that type 2 diabetes is a bihormonal disorder, driven by glucagon excess as well as insulin deficiency. To understand this hypothesis, we need to take a closer look at the anatomical arrangement of the alpha and beta cells within the pancreas. As noted, beta cells constitute about 80% of the cell population of the islets in the dorsal pancreas. These beta cells are in physical contact with one another, and communicate directly with one another via special cell-to-cell “gateways” known as gap junctions. This enables them to function as a syncytium, or single functional unit, rather than in isolation. This is most strikingly demonstrated in the ability of the islet mass to secrete insulin in synchronised pulses, most probably regulated by neural pathways linking the islets.
The glucagon-secreting alpha cells are scattered among the beta cells in such a way that almost all beta cells are in close proximity to an alpha cell. It has long been appreciated that this arrangement has functional implications, and that alpha and beta cells function reciprocally. To use a simple analogy, glucagon is the accelerator pedal controlling the rate at which the liver produces glucose, and insulin is the brake. The controls for this system are designed to ensure that both pedals are not pressed simultaneously, and are highly sophisticated. Three components are recognised: (1) by local or paracrine effects; i.e. the production of each hormone (or other signal molecules produced at the same time) inhibits secretion of the other; (2) neural control via autonomic nerve fibres (which densely innervate the islets); (3) endocrine, via molecules secreted elsewhere in the body.
These developments have led to the increasing recognition that type 2 diabetes is a complex disorder resulting in disordered function of a closely integrated network of chemical and neural signals
^ Staub, A., Sinn, L., and Behrens, 0. K., Characterization of glucagon (hyperglycemic-glycogenolytic factor). Federation Proc., 1954, 13, 303.
^ Sutherland, E. W., Cori, C. F., Haynes, R., and Olsen, N. S., Purification of the hyperglycaemic-glycogenolytic factor from insulin and from gastric mucosa. J. Biol. Chem., 1949, 180, 825.
^ De Duve C. Glucagon; the hyperglycaemic glycogenolytic factor of the pancreas. Lancet 1953;Jul 18;265(6777):99-104
^ Unger RH, Orci L. The essential role of glucagon in the pathogenesis of diabetes mellitus. Lancet 1975;i,14-16