Pancreatic hormones and plasma glucose: regulation mechanisms in the goose under physiological conditions. I. Pancreatectomy and replacement therapy

Measurements of plasma GLI and IRI in normal fasting geese, before and during constant I.V. infusion of saline, gave GLI/I ratios of 1.32 +/- .07 and 1.34 +/- .03 (w/w). As total pancreatectomy markedly reduces the pancreatic hormone level, leading to a mortal hypoglycaemia, we attempted to maintain plasma glucose within the normal range by constant I.V. infusion of glucagon and insulin into operated animals. The results as follows: 1. Blood glucose levels can be maintained within the normal range during experiments lasting 6 or more hours with a constant G/I ratio. 2. The G/I ratio obtained in operated animals (.96 +/- .12) is near to, but significantly lower (p less than .005) than, the GLI/I ratio measured in normal animals. This difference may be explained by the presence of a small amount of circulating gut GLI in the 2nd group.     

Pancreatic hormones disappearance after total pancreatectomy in the duck: correlation between plasma glucagon and glucose

The decline of 3 plasma pancreatic hormones, glucagon (G), insulin (I) and somatostatin (S) was studied in fasting ducks after total pancreatectomy. The results show that the decrease of plasma glucose is highly correlated with the disappearance of G, while no important variation of the G/I ratio occurs during the period of observations (80 minutes) the animal being kept fasted. No participation of pancreatic S in glucose metabolism could be detected, the origin of peripheral S in the fasting state seeming due to the intestine for about 50%.     

Glucose, insulin and glucagon in the diabetic goose.

The relationships between plasma glucose, insulin and glucagon were studied in geese made diabetic by subtotal pancreatectomy. As early as the first hours after the operation, the plasma glucose increases and a permanent diabetes develops. This diabetic state is characterized by two features: a very low plasma insulin level, which does not vary during the survival of the diabetic animals and a concentration of plasma glucagon (of pancreatic origin) which transiently diminishes then rises far above the normal level, and is correlated with the basal concentration of plasma glucose. The impaired glucose tolerance observed in diabetic animals is related to the suppression of the glucose-insulin and glucose-glucagon feedback mechanisms.     

Role of endocrine pancreas in temperature acclimation

Role of endocrine pancreas in temperature acclimation in rats was investigated. Plasma glucagon level increased and insulin level decreased in cold-acclimated rats (CA). The reverse was observed in heat-acclimated rats (HA). In the pancreas there were no changes in glucagon and insulin in CA, but a decrease in glucagon and an increase in insulin were found in HA. Plasma insulin/glucagon molar ratio (I/G) declined in CA and rose in HA. Pancreatic I/G rose in HA. Acute cold exposure elevated plasma glucagon, but did not affect plasma insulin. Pancreatic glucagon, insulin and I/G were not influenced by acute cold exposure, while plasma I/G decreased. Plasma I/G was inversely correlated with both blood free fatty acids and glucose levels. These results suggest that endocrine pancreas is closely associated with metabolic acclimation to cold and heat through its regulation of the metabolic direction to catabolic phase in cold acclimation and to anabolic phase in heat acclimation.     

Progressive loss of sensitivity of the A cell to insulin in geese made diabetic by subtotal pancreatectomy

The effects of insulin deficiency on pancreatic A cell responsiveness to glucose was studied in subtotally depancreatized geese. In geese operated for 3 to 5 days and receiving insulin therapy (I.M.: 0.5-1.0 U/kg/24 h), A cell response to glucose (I.V. injection: 0.5 g/kg) was abolished, but could be restored to normal range by insulin (I.V. injection: 0.025-0.2 U/kg) together with glucose. After 5 weeks of therapy, A cell sensitivity declined: the physiological amount of insulin (0.025 U/kg) was insufficient to suppress glucagon during the glucose load, whereas the large dose (0.2 U/kg) partially restored A cell response. In addition, daily insulin treatment prevented a severe increase of fasting plasma glucose and glucagon. Geese receiving no insulin therapy showed "total blindness" to glucose, even when given insulin at the time of the test. These data suggest a progressive loss of sensitivity of the A cell to insulin. Endocrine and/or panacrine insulin deficiency may play a role on the dysfuncion of the glucose-glucagon feedback mechanism.     

Effects of ghrelin injection on plasma concentrations of glucose, pancreatic hormones and cortisol in Holstein dairy cattle

Ghrelin affects not only growth hormone secretion but also nutrient utilization and metabolic hormone secretion in humans and experimental animals. The effects of ghrelin on plasma metabolic hormone and metabolite levels in domestic herbivores remain unclear despite the fact that the physiological characteristics of nutrient digestion and absorption imply specific responses to ghrelin. Therefore, the effects of ghrelin on plasma glucose, pancreatic hormones and cortisol concentrations were investigated in Holstein dairy cattle in various physiological states. Ghrelin (0.3 nmol/kg) or placebo (2% bovine serum albumin in saline) was intravenously injected in pre-ruminant calves (pre-rumen function), adult non-lactating (functional rumen) and lactating cows (functional rumen and lactation), and plasma glucose, insulin, glucagon and cortisol concentrations were then determined. Ghrelin injection increased plasma glucose concentrations in adult cows, especially in lactating cows. No hyperglycemic response was observed in pre-ruminant calves. A transient rise of insulin and glucagon levels was distinctively found in lactating cows in response to the ghrelin administration. Ghrelin injection decreased the insulin level in pre-ruminant calves. Ghrelin increased cortisol secretion independently of the physiological state. The results of the present study suggest that the effects of ghrelin on plasma glucose and pancreatic hormone levels may reflect differences in the physiological states of dairy cattle.     

Contribution to glucose tolerance of insulin-independent vs. insulin-dependent mechanisms in mice

To study the contributions of insulin-dependent vs. insulin-independent mechanisms to intravenous glucose tolerance (K(G)), 475 experiments in mice were performed. An intravenous glucose bolus was given either alone or with exogenous insulin or with substances modulating insulin secretion and sensitivity. Seven samples were taken over 50 min. Insulin [suprabasal area under the curve (DeltaAUC(ins))] ranged from 0 to 100 mU. ml(-1). 50 min. After validation against the euglycemic hyperinsulinemic clamp, the minimal model of net glucose disappearance was exploited to analyze glucose and insulin concentrations to measure the action of glucose per se independent of dynamic insulin (S(G)) and the combined effect of insulin sensitivity (S(I)) and secretion. Sensitivity analysis showed that insulin [through disposition index (DI)] contributed to glucose tolerance by 29 +/- 4% in normal conditions. In conditions of elevated hyperinsulinemia, contribution by insulin increased on average to 69%. K(G) correlated with DI but was saturated for DeltaAUC(ins) above 15 mU. ml(-1). 50 min. Insulin sensitivity related to DeltaAUC(ins) in a hyperbolic manner, whereas S(G) did not correlate with the insulin peak in the physiological range. Thus glucose tolerance in vivo is largely mediated by mechanisms unrelated to dynamic insulin and saturates with high insulin.     

Role of insulin and glucagon in oxytocin effects on glucose metabolism.

Oxytocin (OT) infusion in normal dogs increases plasma insulin and glucagon levels and increases rates of glucose production and uptake. The purpose of this study was to determine whether the effects of OT on glucose metabolism were direct or indirect. The studies were carried out in normal, unanesthetized dogs in which OT infusion was superimposed on infusion of either somatostatin, which suppresses insulin and glucagon secretion, or clonidine, which suppresses insulin secretion only. Infusion of 0.2 microgram/kg/min of somatostatin suppressed basal levels of plasma insulin and glucagon and inhibited the OT-induced rise of these hormones by about 60-80% of that seen with OT alone. The rates of glucose production and uptake by tissues, measured with [6-3H] glucose, were significantly lower than those seen with OT alone, and the rise in glucose clearance was completely inhibited. Clonidine (30 micrograms/kg, sc), given along with an insulin infusion to replace basal levels of insulin, completely prevented the OT-induced rise in plasma insulin and markedly reduced the glucose uptake seen with OT alone, but did not reduce the usual increase in plasma glucose and glucagon levels or glucose production. To determine whether the OT-induced rise in plasma insulin was in response to the concomitant increase in plasma glucose, similar plasma glucose levels were established in normal dogs by a continuous infusion of glucose and an OT infusion was superimposed. OT did not raise plasma glucose levels further, but plasma insulin levels were increased, indicating that OT can stimulate insulin secretion independently of the plasma glucose changes. Studies by others have shown that the addition of OT to pancreatic islets or intact pancreas can stimulate insulin and glucagon secretion, indicating a direct effect. Our studies agree with that and suggest that in vivo, OT raises plasma insulin levels, at least in part, through a direct action on the pancreas. These studies also show that OT increases glucose production by increasing glucagon secretion and, in addition, a direct effect of OT on glucose production is likely. The OT-induced increase in glucose uptake is mediated largely by increased insulin secretion.     

Maintenance of the postabsorptive plasma glucose concentration: insulin or insulin plus glucagon?

The prevalent view is that the postabsorptive plasma glucose concentration is maintained within the physiological range by the interplay of the glucose-lowering action of insulin and the glucose-raising action of glucagon. It is supported by a body of evidence derived from studies of suppression of glucagon (and insulin, among other effects) with somatostatin in animals and humans, immunoneutralization of glucagon, defective glucagon synthesis, diverse mutations, and absent or reduced glucagon receptors in animals and glucagon antagonists in cells, animals, and humans. Many of these studies are open to alternative interpretations, and some lead to seemingly contradictory conclusions. For example, immunoneutralization of glucagon lowered plasma glucose concentrations in rabbits, but administration of a glucagon antagonist did not lower plasma glucose concentrations in healthy humans. Evidence that the glycemic threshold for glucagon secretion, unlike that for insulin secretion, lies below the physiological range, and the finding that selective suppression of insulin secretion without stimulation of glucagon secretion raises fasting plasma glucose concentrations in humans underscore the primacy of insulin in the regulation of the postabsorptive plasma glucose concentration and challenge the prevalent view. The alternative view is that the postabsorptive plasma glucose concentration is maintained within the physiological range by insulin alone, specifically regulated increments and decrements in insulin, and the resulting decrements and increments in endogenous glucose production, respectively, and glucagon becomes relevant only when glucose levels drift below the physiological range. Although the balance of evidence suggests that glucagon is involved in the maintenance of euglycemia, more definitive evidence is needed, particularly in humans.     

Effect of prolonged fasting upon insulin and glucagon secretion from isolated rat pancreatic islets.

To study insulin-glucagon interrelationships in the regulation of pancreatic islet functions, glucose-mediated insulin and glucagon secretion have been studied in isolated pancreatic islets from fed and from 4 and 8-day fasted rats. At low glucose levels (50 mg %) a continuous decrease of insulin and increase of glucagon secretion were observed during prolonged fasting. High glucose concentrations 300 mg %) stimulated insulin and inhibited glucagon secretion until 4 days, but did not cause any effect after 8 days fasting. These results suggest that the secretory mechanisms of the two hormones may have a common basis.     

Glucagon secretion in chronic pancreatitis.

Plasma insulin, pancreatic glucagon and immunoreactive glucagon-like polypeptide of intestinal origin (enteroglucagon) have been measured in 10 patients with chronic pancreatitis and 5 normal subjects. Basal levels and changes following oral glucose (50 g) and an intravenous infusion of arginine (25 g in 30 min) have been studied. In patients with chronic pancreatitis the plasma insulin response to oral glucose and intravenous arginine was reduced. Basal pancreatic glucagon was increased in the patients and increased further with oral glucose. During an arginine infusion the pancreatic glucagon showed a brisk early increase greater than that seen in the normal subjects. Basal enteroglucagon levels were significantly increased in chronic pancreatitis but response to orla glucose and arginine infusion were little different from those seen in the normal subjects.     

Effect of low-dose dopamine infusion on insulin and glucagon release in fasting normal man

The effect of a low-dose infusion of dopamine on basal circulating concentrations of insulin, glucagon and glucose in six healthy male subjects is reported. Dopamine (0.1 microgram/kg/min) or placebo was given intravenously for 60 minutes. During infusion of the catecholamine, circulating plasma dopamine was 3.46 +/- 1 ng/ml. No change in circulating concentrations of insulin, glucagon and glucose were seen during infusion of dopamine when compared with placebo infusion. It is concluded that dopamine acting at a D2 receptor is unlikely to be of physiological importance in regulation of basal pancreatic islet cell function in man.     

Glucagon secretion during the development of insulin-secreting tumors induced by streptozotocin and nicotinamide.

Serial oral glucose tolerance tests in rats treated with streptozotocin and nicotinamide showed that blood glucose levels after glucose loading were suppressed significantly 7 months after treatment as compared to those of earlier stages. Post-glucose plasma insulin levels were significantly elevated at the 9th to 12th month and concomitantly fasting plasma glucagon levels rose significantly. At that time pancreatic islet cell tumors were demonstrated in all of the rats in this experiment. Post-glucose plasma glucagon levels, however, did not show remarkable changes throughout the observation. In spite of hyperinsulinemia, post-glucose plasma glucagon levels of tumor-bearing rats were significantly lower than those of body weight adjusted controls. It is inferred from the study that secretory activity of pancreatic A-cells of tumor-bearing rats is restrained by excess insulin released from islet cell tumors.     

Glucose dependent stimulation by prostaglandin D2 of glucagon and insulin in perfused rat pancreas

Effects of prostaglandin D2 on pancreatic islet function in perfused rat pancreas were examined in comparison with those of prostaglandin E2, which has hitherto been suggested to be a modifier of pancreatic hormone release. In the presence of 2.8 mM glucose, only glucagon release was strongly stimulated by 14 microM of prostaglandin D2, while release of both glucagon and insulin was augmented by 14 microM of prostaglandin E2. When the glucose concentration was elevated to 11.2 mM, insulin release was accelerated by 14 microM of prostaglandin D2 but there was no effect upon glucagon release. Again, release of both glucagon and insulin was augmented by 14 microM of prostaglandin E2 in the presence of 11.2 mM of glucose. The regulation of glucagon and insulin release through prostaglandin D2 is apparently adapted to glycemic changes, and may be a physiological modulator of pancreatic islet function.     

Hepatotoxicity induced by diethylnitrosamine causes no significant disturbances of systemic glucose homeostasis in rats

In rats, a moderately hepatotoxic single dose of diethylnitrosamine (DEN) 100 mg/kg causing depletion of liver glycogen, elevation of aspartate aminotransferase and decreased liver uptake of 3-O-methylglucose, resulted in substantial changes in insulin and glucagon balance. Two days after DEN, insulin binding to liver membranes and insulin removal by the liver were sharply reduced whereas its binding to muscle and adipocyte membranes remained unaltered. Serum insulin (random and after an overnight fast) remained normal. Intravenous (I.V.) insulin (10 U/kg) caused the usual degree of hypoglycemia that, however, lasted longer than in the control animals. Removal of glucagon by liver was also depressed in spite of its normal binding to hepatocytes, and peripheral serum glucagon was increased three-fold. I.V. glucagon (40 micrograms/kg) resulted in a blunted response of plasma glucose. I.V. glucose tolerance test (1 g/kg) remained normal in spite of the insulin increase to a level twice as high as in the controls, and in spite of nonsuppressed glucagon. These changes were still present after 1-3 months, but disappeared by 6 months. The results demonstrate remarkable ability of homeostatic mechanisms to preserve normal plasma glucose and glucose tolerance in spite of dramatic changes in insulin and glucagon.     

Contribution of free fatty acids to impairment of insulin secretion and action: mechanism of beta-cell lipotoxicity

Type 2 diabetes is characterized by two major defects: a dysregulation of pancreatic hormone secretion (quantitative and qualitative--early phase, pulsatility--decrease of insulin secretion, increase in glucagon secretion), and a decrease in insulin action on target tissues (insulin resistance). The defects in insulin action on target tissues are characterized by a decreased in muscle glucose uptake and by an increased hepatic glucose production. These abnomalities are linked to several defects in insulin signaling mechanisms and in several steps regulating glucose metabolism (transport, key enzymes of glycogen synthesis or of mitochondrial oxidation). These postreceptors defects are amplified by the presence of high circulating concentrations of free fatty acids. The mechanisms involved in the of long-chain fatty acids are reviewed in this paper. Indeed, elevated plasma free fatty acids contribute to decrease muscle glucose uptake (mainly by reducing insulin signaling) and to increase hepatic glucose production (stimulation of gluconeogenesis by providing cofactors such as acetyl-CoA, ATP and NADH). Chronic exposure to high levels of plasma free fatty acids induces accumulation of long-chain acyl-CoA into pancreatic beta-cells and to the death of 50 % of beta-cell by apoptosis (lipotoxicity).     

Beta- and alpha-cell dysfunction in type 2 diabetes.

Insulin resistance is a common pathogenetic feature of type 2 diabetes. However, hyperglycemia would not develop if a concomitant defect in insulin secretion were not present. Impaired insulin secretion results from functional and survival defects of the beta-cell. The functional defects can be demonstrated early in the natural history of diabetes and they are hallmarked by abnormal pulsatility of basal insulin secretion and loss of first-phase insulin release in response to a glucose challenge. Moreover, a significant reduction of the beta-cell mass is apparent at the time of the diagnosis of diabetes. The progressive increase in glucose levels, that seems to characterize the natural history of type 2 diabetes, has been claimed to be largely due to progressive reduction of function and mass of beta-cells. Although a genetic predisposition is likely to account for impaired insulin secretion, chronic exposure to hyperglycemia and high circulating FFA is likely to contribute to both functional and survival defects. The disturbance in the endocrine activity of the pancreas is not limited to insulin, since a concomitant increase in fasting plasma glucagon and impaired suppression after the ingestion of an oral glucose load are often observed. This alteration becomes prominent after the ingestion of a mixed meal, when plasma glucagon remains much higher in the diabetic patient as compared to normal individuals. The disproportionate changes in the plasma concentration of the two pancreatic hormones is clearly evident when the insulin:glucagon molar ratio is considered. It is the latter that mainly affects hepatic glucose production. Because of the reduction of the insulin:glucagon molar ratio basal endogenous glucose concentration will be higher causing fasting hyperglycemia, while the hepatic glucose output will not be efficiently suppressed after the ingestion of a meal, contributing to excessive post-prandial glucose rise. Correcting beta- and alpha-cell dysfunction becomes, therefore, an attractive and rational therapeutic approach, particularly in the light of new treatments that may directly act on these pathogenetic mechanisms of type 2 diabetes.     

Model for glucagon secretion by pancreatic α-cells

Glucagon hormone is synthesized and released by pancreatic α-cells, one of the islet-cell types. This hormone, along with insulin, maintains blood glucose levels within the physiological range. Glucose stimulates glucagon release at low concentrations (hypoglycemia). However, the mechanisms involved in this secretion are still not completely clear. Here, using experimental calcium time series obtained in mouse pancreatic islets at low and high glucose conditions, we propose a glucagon secretion model for α-cells. Our model takes into account that the resupply of releasable granules is not only controlled by cytoplasmic Ca2+, as in other neuroendocrine and endocrine cells, but also by the level of extracellular glucose. We found that, although calcium oscillations are highly variable, the average secretion rates predicted by the model fall into the range of values reported in the literature, for both stimulated and non-stimulated conditions. For low glucose levels, the model predicts that there would be a well-controlled number of releasable granules refilled slowly from a large reserve pool, probably to ensure a secretion rate that could last for several minutes. Studying the α-cell response to the addition of insulin at low glucose, we observe that the presence of insulin reduces glucagon release by decreasing the islet Ca2+ level. This observation is in line with previous work reporting that Ca2+ dynamics, mainly frequency, is altered by insulin. Thus, the present results emphasize the main role played by Ca2+ and glucose in the control of glucagon secretion by α-cells. Our modeling approach also shows that calcium oscillations potentiate glucagon secretion as compared to constant levels of this cellular messenger. Altogether, the model sheds new light on the subcellular mechanisms involved in α-cell exocytosis, and provides a quantitative predictive tool for studying glucagon secretion modulators in physiological and pathological conditions.     

Effects of biotin deficiency on pancreatic islet morphology, insulin sensitivity and glucose homeostasis

Several studies have revealed that physiological concentrations of biotin are required for the normal expression of critical carbohydrate metabolism genes and for glucose homeostasis. However, the different experimental models used in these studies make it difficult to integrate the effects of biotin deficiency on glucose metabolism. To further investigate the effects of biotin deficiency on glucose metabolism, we presently analyzed the effect of biotin deprivation on glucose homeostasis and on pancreatic islet morphology. Three-week-old male BALB/cAnN Hsd mice were fed a biotin-deficient or a biotin-control diet (0 or 7.2 μmol of free biotin/kg diet, respectively) over a period of 8 weeks. We found that biotin deprivation caused reduced concentrations of blood glucose and serum insulin concentrations, but increased plasma glucagon levels. Biotin-deficient mice also presented impaired glucose and insulin tolerance tests, indicating defects in insulin sensitivity. Altered insulin signaling was linked to a decrease in phosphorylated Akt/PKB but induced no change in insulin receptor abundance. Islet morphology studies revealed disruption of islet architecture due to biotin deficiency, and an increase in the number of α-cells in the islet core. Morphometric analyses found increased islet size, number of islets and glucagon-positive area, but a decreased insulin-positive area, in the biotin-deficient group. Glucagon secretion and gene expression increased in islets isolated from biotin-deficient mice. Our results suggest that biotin deficiency promotes hyperglycemic mechanisms such as increased glucagon concentration and decreased insulin secretion and sensitivity to compensate for reduced blood glucose concentrations. Variations in glucose homeostasis may participate in the changes observed in pancreatic islets.