Barbara J. Frankel

Abnormal Secretion of Insulin and Glucagon by the In Vitro Perfused Pancreas of the Genetically Diabetic Chinese Hamster

Hereditary insulin-deficient diabetes mellitus occurs in certain sublines of nonobese Chinese hamsters. Several characteristics of this syndrome are similar to those seen in insulin-deficient human diabetics. Therefore, to characterize pancreatic islet function, dynamic insulin and glucagon release from normal and nonketotic diabetic hamster pancreases in response to glucose (300 mg/100 ml) and theophylline (10 mM), infused singly and together, was studied in vitro.20-min glucose infusions of normal hamster pancreases caused biphasic insulin release, consisting of a rapid first peak and a gradually rising second phase, similar to that reported for man in vivo. Both phases were significantly reduced in the diabetic pancreases. Theophylline alone stimulated similar nonphasic insulin release in both the normal and the diabetic pancreases. Glucose and theophylline together caused greater insulin release than either stimulant alone in both normals and diabetics; however, the diabetic response was still subnormal. Glucose suppressed glucagon release from normal pancreases; suppression was significantly impaired in diabetics. Theophylline stimulated nonphasic glucagon release in both the normals and diabetics. Glucose partially suppressed the theophylline-stimulated release in both groups.Insulin/glucagon molar ratios of the diabetics were consistently subnormal, although individual hormone levels often overlapped into the normal range. IN SUMMARY, THE PANCREASES OF GENETICALLY DIABETIC CHINESE HAMSTERS PERFUSED IN VITRO SHOWED: (a) decreased first and second phase insulin release in response to glucose-containing stimuli-only partially ameliorated by theophylline-, and (b) impaired suppression of glucagon in response to glucose, resulting in (c) a decreased insulin/glucagon molar ratio. These data support the suggestion that both alpha and beta cells of diabetic pancreases may be insensitive to glucose.

The diabetic Chinese hamster: In vitro insulin and glucagon release; the “chemical diabetic”; and the effect of diet on ketonuria

SummaryInsulin and glucagon release (in response to glucose, theophylline, and arginine) were measured from thein vitro perfused pancreases of non-ketotic diabetic Chinese hamsters. These animals showed impaired alpha and beta cell sensitivity to glucose (i.e. excessive glucagon, and a reduction in both phases of insulin release) and abnormal alpha cell sensitivity to arginine (excessive glucagon), but normal responses to theophylline. Pancreatic insulin content was significantly decreased and glucagon content increased in unperfused diabetic pancreases. Abnormal responses to glucose and arginine were not directly related to pancreatic content, since responses to theophylline were normal. Insulin responses of normal animals from five normal sublines and non-ketotics from eight diabetic sublines were compared; certain diabetic sublines showed significantly less insulin release than did others, despite similar severities of glucosuria. — Fasting blood glucose during stress, glucose tolerance, andin vitro pancreatic insulin responses to glucose were measured in the normoglycemic, aglucosuric siblings of diabetics; the responses were diabetic-like, and therefore such animals are referred to here as “chemical diabetics”. — The incidence of glucosuria and ketonuria was studied in ketotic diabetic hamsters on high- and low-fat diets; glucosuria decreased and ketonuria disappeared on the low-fat diet.

Glucose-stimulated 45Ca Uptake in Isolated Rat Islets

Net 45Ca uptake (in excess of the extracellular 3H-sucrose space) and insulin release were measured under low (2 mM) and high (20 mM) glucose conditions in collagenase-isolated rat islets. 45Ca uptake curves were mathematically fitted and subjected to compartmental analysis. Within the first 60 seconds after addition of trace 45Ca, islets showed a similar rapid uptake of 45Ca regardless of the glucose concentration or length of time of prior exposure to glucose. Net 45Ca uptake continued to increase for 30-60 minutes, and the islets in high glucose showed approximately twofold greater maximum uptake than islets in low glucose. Islets preincubated in low glucose and then incubated in high glucose showed a 5-15-minute delay in net 45Ca uptake as compared with islets that had been preincubated in high glucose. Insulin release was detectable by 10 minutes of incubation with high glucose. Mathematical modeling of the low- and high-glucose net 45Ca uptake curves suggests that there are at least two calcium “compartments” within the β-cell. One compartment is small, rapidly filled, and insensitive to glucose, while the other, larger compartment, is slowly filled and fills to a much greater extent in the presence of high glucose. A major proportion of the glucose-stimulated uptake is at the level of influx.

Insulin and Glucagon Release in the Diabetic Chinese Hamster: Differences Among Inbred Sublines

SummaryRelease of insulin and glucagon from perfused pancreases in vitro of 40 normal male and female Chinese hamsters (from one inbred subline) and 110 male and female diabetic hamsters (from three inbred sublines) was measured in response to glucose plus arginine, theophylline alone, or potassium alone, in order to determine if differences in hormone secretion exist among different diabetic sublines. Glucose plus arginine and potassium produced subnormal insulin responses in all three diabetic sublines, whereas theophylline induced ‘normal’ or above normal insulin responses. Excessive glucagon release was consistently seen in only one diabetic subline. The female normal animals showed greater insulin release than the male normal hamsters in response to glucose plus arginine. This sex difference was not seen in the diabetic animals.

Abnormalities in Glucose-Stimulated Insulin Release, 45Ca Uptake, and 86Rb Efflux in Diabetic Chinese Hamster Islets

We loaded islets from normal and diabetic Chinese hamsters with 86Rb (an analogue for K+) and measured 86Rb efflux during stimulation with 20 mM D-glucose. Genetically diabetic Chinese hamsters were selected from a subline (L) known for subnormal pancreatic insulin release and excessive pancreatic glucagon release in vitro. 86Rb accumulation in 1 mM glucose was normal in the diabetic islets. Similar to the pattern of 86Rb efflux previously seen from normal rat and mouse islets, 20 mM glucose suppressed 86Rb efflux within 1-2 min, and efflux remained suppressed until return to 1 mM glucose in both normal and diabetic hamster islets. After the first 2 min of 20 mM glucose, suppression of 86Rb efflux was somewhat greater in the diabetic hamster islets than in the normals. In addition, glucose-stimulated insulin release and 45Ca uptake were significantly reduced in the diabetic islets. Therefore, in the diabetic hamster islets, there is at least no impairment in the initial suppression of 86Rb efflux by glucose. This suggests that the diabetic β-cells recognize glucose and carry out the initial steps in the stimulus-secretion coupling sequence normally. The later, excessive suppression of 86Rb efflux may be due to impaired Ca2+-induced changes in 86Rb efflux, suggesting that defective regulation of intracellular Ca2+ activity, rather than defective regulation of K+ permeability, may lead to the impaired insulin secretion.

Insulin, glucagon, and somatostatin release from the prediabetic Chinese hamster

SummaryDiabetes mellitus in the adult Chinese hamster is characterized by subnormal pancreatic insulin release in vitro, decreased insulin content, and lack of obesity. The cause of the islet B-cell failure is not clear. We measured insulin, glucagon, and somatostatin release from in vitro perfused pancreases of young (mean age 10 and 20 weeks), genetically diabetic animals (subline AC, mean plasma glucose 8.0 and 16.6mmol/l, respectively). Compared to age- and sex-matched normal hamsters (subline M, mean plasma glucose 5.3 mmol/l), the younger diabetic animals had a significantly elevated mean plasma glucose level, but net in vitro pancreatic release of insulin, glucagon, and somatostatin was normal. Pancreatic content of insulin and glucagon was also not significantly different from normal. At age 20 weeks, when the plasma glucose of the diabetic animals was even more elevated, pancreatic content and release of insulin were significantly subnormal, whereas glucagon and somatostatin release were normal, and pancreatic content of glucagon was normal. In a similar group of young (mean age 10 weeks) diabetic animals, non-fasting plasma insulin levels were within the normal range, but the corresponding glucose levels were excessive in most of the animals (13 out of 19). In conclusion, 10-week-old diabetic hamsters show mild hyperglycaemia which cannot be accounted for directly by decreased pancreatic release in response to a glucose plus arginine stimulus in vitro. Decreased ability of the B cell to respond in vivo to hyperglycaemia or peripheral resistance to insulin may contribute to later B-cell failure in the older diabetic hamster.

Effect of perchlorate on glucose-stimulated insulin release and 45Ca2+ uptake in pancreatic islets from diabetic Chinese hamsters

Insulin release and 45Ca2+ uptake were studied in isolated islets from Chinese hamsters of genetically diabetic and normal sublines. The calcium channel agonist, perchlorate (C104-, 12 mmol/L), augmented both 45Ca2+ uptake and insulin release from normal islets in the presence of 20 but not 1 mmol/L glucose. The agonist also amplified the glucose-stimulated 45Ca2+ uptake and insulin release from diabetic islets but did not normalize the insulin release despite normal insulin concentration in the diabetic Chinese hamster islets. The dry weight of the diabetic islets was subnormal (54%, p < 0.005) but the insulin concentration (insulin per dry weight of islet tissue) was not different from normal (122%). It appears that there are defective mechanisms in addition to the glucose-stimulated influx of Ca2+ in diabetic islet B cells.