For years investigators have sought an assay for insulin which would combine virtually absolute specificity with a high degree of sensitivity, sufficiently exquisite for measurement of the minute insulin concentrations usually present in the circulation. Methods in use recently depend on the ability of insulin to exert an effect on the metabolism of glucose in vivo or in excised muscle or adipose tissue. Thus, the insulin concentration in plasma has been estimated: a) from the degree of hypoglycemia produced in hypophysectomized, adrenalectomized, alloxan-diabetic rats (1); b) from the augmentation of glucose uptake by isolated rat hemidiaphragm (2); or c) from the increased oxidation of glucose-1-C14 by the rat epididymal fat pad (3). Since there have been reports indicating the presence, in plasma, of inhibitors of insulin action (4) and of noninsulin substances capable of inducing an insulin-like effect (5,6), these procedures, while yielding interesting information regarding the effects of various plasmas on glucose metabolism in tissues, are of doubtful specificity for the measurement of insulin per se (5).

Immunoassay of endogenous plasma insulin in man

In six patients with acanthosis nigricans variable degrees of glucose intolerance, hyperinsulinemia and marked resistance to exogenous insulin were found. Studies of insulin receptors on circulating monocytes suggest that the insulin resistance in these patients was due to a marked decrease in insulin binding to its membrane receptors. When these patients were fasted, there was a fall in plasma insulin but no increase in insulin binding, suggesting that the receptor defect was not secondary to the hyperinsulinemia. The clinical features shared by these cases and several similar ones previously reported may be divided into two unique clinical syndromes: Type A, a syndrome in younger females with signs of virilization or accelerated growth, in whom the receptor defect may be primary, and Type B, a syndrome in older females with signs of an immunologic disease, in whom circulating antibodies to the insulin receptor are found.

The syndromes of insulin resistance and acanthosis nigricans. Insulin-receptor disorders in man.

Integrated insulin secretion rates calculated from peripheral venous C-peptide measurements by two-compartment kinetic analysis were measured in six young normal subjects after increasing oral glucose loads of 25, 50, and 100 g and respective isoglycemic glucose infusions. The differences in B-cell secretory responses between oral and iv glucose challenges were attributed to factors other than glycemia itself (incretin effect). Both insulin and C-peptide concentrations as well as calculated integrated insulin secretion rates increased with increasing oral glucose loads. Due to the similarity in the glucose profiles after all oral loads, almost identical amounts of iv glucose (approximately 20 g) were infused in all "isoglycemic" infusion experiments, with resulting similar hormone profiles and insulin secretion rates. The percent contribution of incretin factors to total immunoreactive insulin responses after 25, 50, and 100 g glucose (85.6%, 74.9%, and 93.0%; response to oral load, 100%) was significantly higher than their contribution to integrated C-peptide responses (27.6-62.9%) or calculated integrated insulin secretion rates (19.2-61.0%). These findings indicate that the degree of incretin stimulation of insulin secretion depends on the amount of glucose ingested. A discrepancy between the estimates of the incretin effect derived from peripheral venous insulin responses, on the one hand, and C-peptide responses or calculated insulin secretion rates, on the other hand, exists. Inasmuch as peripheral insulin values reflect both insulin secretion and hepatic insulin removal, this discrepancy suggests that elimination kinetics of insulin differ between oral and iv glucose administration. This difference can be related to a significantly reduced fractional hepatic insulin extraction after oral (46.9-54.6%) compared to iv (63.4-76.5%) glucose administration when calculated by a three-compartment kinetic model. This reduction in fractional hepatic insulin extraction could be caused by gastrointestinal factors (hormones or nerves) stimulated in the course of glucose ingestion.

Incretin effects of increasing glucose loads in man calculated from venous insulin and C-peptide responses

This review is aimed at providing readers with a comprehensive reference article about the distribution and function of P2 receptors in all the organs, tissues, and cells in the body. Each section provides an account of the early history of purinergic signaling in the organ/cell up to 1994, then summarizes subsequent evidence for the presence of P2X and P2Y receptor subtype mRNA and proteins as well as functional data, all fully referenced. A section is included describing the plasticity of expression of P2 receptors during development and aging as well as in various pathophysiological conditions. Finally, there is some discussion of possible future developments in the purinergic signaling field.

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Cellular distribution and functions of P2 receptor subtypes in different systems.

An approach based on the polymerase chain reaction has been devised to clone new members of the family of genes encoding guanosine triphosphate-binding protein (G protein)-coupled receptors. Degenerate primers corresponding to consensus sequences of the third and sixth transmembrane segments of available receptors were used to selectively amplify and clone members of this gene family from thyroid complementary DNA. Clones encoding three known receptors and four new putative receptors were obtained. Sequence comparisons established that the new genes belong to the G protein-coupled receptor family. Close structural similarity was observed between one of the putative receptors and the 5HT1a receptor. Two other molecules displayed common sequence characteristics, suggesting that they are members of a new subfamily of receptors with a very short nonglycosylated (extracellular) amino-terminal extension.

https://science.sciencemag.org/content/sci/244/4904/569.full.pdf

Selective amplification and cloning of four new members of the G protein-coupled receptor family.

Publisher Summary Pertussis and cholera toxins are important tools to investigate functional and structural aspects of the stimulatory (Ns) and inhibitory (Ni) regulatory components of adenylyl cyclise. Cholera toxin acts on Ns by ADP-ribosylating its αs subunit. It uses NAD+ as a cosubstrate. ADP-ribosylation of Ns alters its properties so that its guanosine triphosphate (GTP) hydrolyzing capacity is inhibited and the action of GTP is potentiated. In intact cells, this leads to the increases in cyclic adenosine monophosphate (AMP) levels. Both pertussis and cholera toxin are hexameric multisubunit molecules. Cholera toxin is composed of one A and 5 B subunits; pertussis toxin is formed of one S1, one S2, one S3, two S4, and one S5 subunits. This chapter presents a set of protocols, as developed in the laboratory, that can be used to study simultaneously and comparatively the susceptibility of Ns and Ni to be ADP-ribosylated by cholera and pertussis toxin.

ADP-ribosylation of membrane components by pertussis and cholera toxin.

Hypoglycemia after ingestion of ethanolic beverages has been reported (1-4). In some cases (1, 2), hypoglycemia was attributed to the nonethanol ingredients in such mixtures as \"smoke\" and \"solox\" (methyl alcohol, and gasoline and ethyl acetate). Cummins, however, suggested that hypoglycemia might be caused by ethanol itself (3). He described a 6-year-old boy who had convulsions and hypoglycemia after ingestion of gin. Neame and Joubert also ascribed hypoglycemia observed in their patients to ethanol rather than other ingredients (4). They also emphasized the inadequate dietary intake before the ethanolic ingestion and the development of hypoglycemia. Recently, we studied a chronic alcoholic patient who had several documented episodes of hypoglycemia and minimal evidence of abnormal liver function. Our findings indicated that his hypoglycemia was probably induced by ethanol consumption in combination with a poor dietary intake. Studies were also performed with normal control subjects which indicated that ethanol ingestion after a 2-day fast caused hypoglycemia. During ethanol-induced hypoglycemia, conversion of fructose to glucose was normal, but glycogen synthesis appeared to be inhibited, from the failure of an adequate hyperglycemic response to glucagon. Animal studies suggested that ethanol caused hypoglycemia by interfering with gluconeogenesis as well as glycogen synthesis. Recent studies reported in abstract form have also proposed that the mechanism of ethanol-induced hypoglycemia might be an interference in gluconeogenesis (5).

/pdf/studies-on-the-mechanism-of-ethanol-induced-hypoglycemia-lzgypjoy07.pdf

Studies on the mechanism of ethanol-induced hypoglycemia.

https://www.amjmed.com/article/0002-9343(71)90322-6/pdf

Impaired water excretion in myxedema

Extraction of insulin by the liver after administration of glucose in the duodenum has been studied in fourteen anesthetized dogs. Plasma insulin and glucose were measured in the portal vein hepatic vein and hepatic artery. During the control period 40+/-3% of the approximately 11 mU of insulin presented to the liver/min was removed during a single transhepatic passage. Within 5 min after glucose administration, the amount of insulin reaching the liver increased significantly. In some animals this increase preceded any significant increase in the glucose concentration of the femoral artery. After glucose administration, hepatic extraction of insulin remained unchanged in five animals and rose significantly in nine. In five of the latter animals, the increase may have been more apparent than real due to nonrepresentative sampling of hepatic venous blood. However, for the whole group of animals, comparison of arterial insulin levels with the amount of insulin delivered to the liver suggested a transient increase in insulin extraction between 5 and 50 min after glucose administration. In no animal was there a decrease in the proportion of insulin extracted by the liver after glucose administration. The results indicate that the extraction process is not saturable at physiological insulin levels. Prior to glucose administration, net hepatic glucose output averaged between 30 and 40 mg/min. After glucose administration, the liver began to take up glucose and there was a significant correlation between hepatic glucose uptake and the amount of insulin reaching the liver. However, since the amount of glucose presented to the liver also increased, it is not established that the insulin was responsible for the change in hepatic carbohydrate metabolism. The data demonstrate an increase in the absolute amount of insulin extracted by the liver after glucose administration and an important role for the liver in regulating peripheral insulin concentrations.

/pdf/effect-of-intraduodenal-glucose-administration-on-hepatic-170xfcn3vl.pdf

Effect of Intraduodenal Glucose Administration on Hepatic Extraction of Insulin in the Anesthetized Dog

The effect of equal (1.1 +/- 0.1 g/kg body wt) amounts of glucose administered orally, or by peripheral intravenous or intraportal infusion on hepatic glucose uptake and fractional hepatic extraction of insulin and glucagon was studied in conscious dogs with chronically implanted Doppler flow probes on the portal vein and hepatic artery and catheters in the portal vein, hepatic vein, carotid artery, and superior mesenteric vein. Portal vein and hepatic vein plasma flow increased only after oral glucose administration. Arterial plasma glucose increased equally to 150-160 mg/100 ml after all three routes of glucose administration. Portal vein glucose was similar after oral (195 +/- 15 mg/100 ml) and intraportal glucose infusion (215 +/- 11 mg/100 ml) and significantly higher than after peripheral intravenous glucose. Hepatic glucose uptake after oral (68 +/- 4%) and intraportal glucose administration (65 +/- 7%) significantly exceeded that after peripheral intravenous glucose infusion (23 +/- 5%). The amount of insulin above basal presented to the liver during the 180 min after oral glucose was 7.6 +/- 1.3 U, 4.3 +/- 0.6 U after intraportal glucose, and 4.1 +/- 0.6 U after peripheral intravenous glucose. Hepatic extraction of insulin increased significantly after oral glucose (42 +/- 3 to 61 +/- 4%), but was unchanged after intraportal and peripheral intravenous glucose administration. When the portal vein glucose levels achieved during peripheral intravenous glucose infusion for 90 min were maintained by a subsequent 90-min intraportal glucose infusion, hepatic glucose uptake was significantly greater during the intraportal glucose infusion. Glucagon secretion was suppressed equally after oral glucose, intraportal glucose, and peripheral intravenous glucose administration; fractional hepatic extraction of that hormone, which was significantly less than that of insulin, was unchanged. These results indicate that hepatic glucose uptake is significantly greater after oral and intraportal glucose administration than after peripheral intravenous glucose infusion. This difference is not simply related to the amount of glucose or insulin presented to the liver and the increased hepatic glucose uptake did not depend solely upon the augmented fractional hepatic extraction of insulin. Hepatic extraction of insulin and hepatic glucose uptake appear to be regulated independently.

/pdf/differential-effects-of-oral-peripheral-intravenous-and-2b43xiokts.pdf

Differential effects of oral, peripheral intravenous, and intraportal glucose on hepatic glucose uptake and insulin and glucagon extraction in conscious dogs.

James B. Field

Papers

ADP-ribosylation of membrane components by pertussis and cholera toxin.

Studies on the mechanism of ethanol-induced hypoglycemia.

Impaired water excretion in myxedema

Effect of Intraduodenal Glucose Administration on Hepatic Extraction of Insulin in the Anesthetized Dog

Differential effects of oral, peripheral intravenous, and intraportal glucose on hepatic glucose uptake and insulin and glucagon extraction in conscious dogs.