Citations

Although sulfonylureas enhance insulin secretion, it is unknown whether these hypoglycemic chemicals stimulate insulin secretion through the augmentation of the pulsatile or basal modes of insulin release. Enhanced pulsatile insulin could occur in turn through amplification of the burst mass or an increase in burst frequency. To address the mechanism of sulfonylurea action, we employed a recently validated canine model with a portal vein sampling catheter and flow probe to measure pulsatile insulin secretion in vivo directly in response to tolbutamide infusion or ingestion. After a 16-h fast, seven dogs were studied in the postabsorptive basal state and during a tolbutamide (0.2 mg/min) infusion when their plasma glucose concentrations were clamped at euglycemia. Insulin concentrations in the carotid artery (basal vs. tolbutamide, 85 ± 12 vs. 325 ± 66 pmol/l; P < 0.01) and portal vein (basal vs. tolbutamide, 345 ± 55 vs. 1,288 ± 230 pmol/l; P < 0.01) increased during tolbutamide infusion, but the portal vein plasma flow did not change. Increased plasma insulin concentrations were achieved by a fourfold increase in the total insulin secretion rate (2.3 ± 0.2 to 9.4 ± 1.9 pmol · kg−1 · min−1; basal vs. tolbutamide, P < 0.01). The augmented total insulin secretion was achieved mechanistically via a marked and selective increase in the insulin secretory burst mass (basal vs. tolbutamide, 266 ± 64 vs. 817 ± 144 pmol/pulse; P < 0.01), with no change in portal-vein insulin pulse frequency (basal vs. tolbutamide, 10.1 ± 0.6 vs. 11.1 ± 0.8 pulses/h; P = 0.3). Oral (250 mg) tolbutamide also magnified the endogenous insulin secretion rate by the preferential amplification of the secretory pulse mass (basal vs. tolbutamide, 167 ± 37 vs. 362 ± 50 pmol/pulse; P < 0.01). Neither the infusion nor the ingestion of tolbutamide changed the calculated clearance rates of endogenously secreted insulin. We conclude that sulfonylurea (tolbutamide) induced insulin secretion in vivo is achieved by the highly selective amplification of insulin secretory burst mass with no change in basal insulin release or the frequency of the β-cell-network pacemaker.

In the present studies, we used a recently validated canine model to determine 1) if glucose ingestion stimulates insulin secretion by amplifying the pulsatile component of insulin release, and if so, 2) whether this effect is achieved preferentially through burst mass or frequency modulation, and 3) if the mechanism of incretin effect of insulin secretion is mediated via the pulsatile mode of secretion. We report that 30 g of glucose ingestion stimulates an ∼ 550% increase in the overall rate of insulin secretion (1.8 ± 0.2 to 11.6 ± 1.5 pmol.kg−1 · min−1), which is achieved via an ∼ 400% increase in the mass of insulin secreted per burst (202 ± 38 to 1,003 ± 147 pmol/pulse, P < 0.001) and a ∼ 40% increase in burst frequency (8.7 ± 0.5 to 12.3 ± 0.6 pulse/h, P < 0.001). Of the insulin secreted after glucose ingestion, 68% (±4) was released in discrete secretory bursts. Further analyses showed that the incretin effect of ingested (GPO) versus infused glucose (GIV) is achieved through regulation of pulsatile insulin secretion. Glucose ingestion led to an ∼ 70% greater rate of insulin secretion than intravenous glucose delivery (10.0 ± 1.6 vs. 5.9 ± 0.9 pmol.kg−1 · min−1, P < 0.005, GPO vs. GIV). This incretin effect was achieved by the specific mechanism of an ∼ 70% greater pulse mass (930 ± 196 vs. 558 ± 97 pmol/pulse, P < 0.02, GPO vs. GIV) but with a comparable pulse frequency (13.1 ± 0.9 vs. 12.0 ± 0.5 pulses/h, P = 0.14, n = 9 dogs, GPO vs. GIV). We conclude that in vivo glucose regulates overall insulin secretion almost exclusively by amplification of the pulsatile mode of insulin secretion, and that the incretin effect is achieved by preferential enhancement of insulin secretory burst mass.

Insulin has important effects to increase skeletal muscle (leg) blood flow under euglycemic hyperinsulinemic clamp conditions and after oral glucose tolerance testing. The present studies examined the effects of mixed meal consumption on the components of leg glucose uptake (LGU) in lean, healthy adults. Seventeen men and women underwent measures of leg plasma flow and arteriovenous (AV) glucose difference before and for 6 h after a mixed meal providing one-third of daily energy expenditure. Another eight men and women underwent the same studies before and during the consumption of the same-size meal administered in small frequent feedings over 6 h. After the bolus meal, peak leg AV glucose gradient increased ∼ fivefold (P < 0.001), whereas the peak increase in leg plasma flow was 20% (NS). No significant contribution of increased leg blood flow to the increase in postprandial LGU was apparent. Over the last 100 min of the frequent-feedings meal, the leg AV difference increased ∼ fourfold (P < 0.001 vs. basal), whereas leg blood flow increased only by 16% (NS vs. basal). We conclude that after a mixed meal, leg (primarily skeletal muscle) blood flow does not increase enough for blood flow to be a major contributor to glucose uptake. These findings raise questions regarding the relative importance of insulin's hemodynamic effects in modulating glucose tolerance under more usual conditions.

Insulin and insulin-like growth factor I (IGF-I) are structurally related peptides capable of stimulating a variety of metabolic and mitogenic processes. In this study, we investigated the interaction between these peptides and their receptor-mediated pathways in an untransformed cell line. Cultured bovine fibroblasts specifically bound IGF-I and insulin, and each peptide could stimulate DNA synthesis and cell replication through its own receptor. Preincubation of bovine fibroblasts with concentrations of insulin that did not bind to the IGF-I receptor resulted in complete but reversible cellular desensitization to IGF-I-stimulated mitogenesis. Preincubation with as little as 0.1 nM insulin was sufficient to inhibit subsequent IGF-I action. Various insulin analogs produced desensitization in direct relation to the affinity of the insulin for the insulin receptor. Desensitization required > 4 h of cell exposure to insulin and was blocked in the presence of cycloheximide. Neither serum-stimulated mitogenesis nor IGF-I-stimulated glucose uptake were affected by insulin pretreatment. 125I-labeled IGF-I affinity cross-linking experiments indicated that preincubation with insulin did not affect labeling of the 130,000-Mr α-subunit of the IGF-I receptor, but was associated with the loss of IGF-I– and insulin-inhibitive bands at Mr = 100,000, 85,000, 58,000, and 34,000. These studies suggest that insulin, via interaction with insulin receptors on bovine fibroblasts, regulates IGF-I action at a step distal to IGF-I receptor binding and are consistent with desensitization occurring at an intracellular step in the mitogenic pathway shared by insulin and IGF-I.

Amyloid deposits that characteristically form in the pancreatic islets of patients with non-insulin-dependent diabetes mellitus (NIDDM) and in insulinomas are both derived from islet amyloid polypeptide (IAPP). Evidence from previous studies has suggested that deposition of IAPP-derived amyloid is related to inherent amyloidogenic sequences present within normal human IAPP, together with an increased production and local concentration of IAPP. However, whether the aggregation of IAPP to form amyloid fibrils is primarily an intra- or extracellular event is not clear. To address this question, we studied 20 human insulinomas by light and electron microscopy. By light microscopy, amyloid deposits were demonstrated in 13 of 20 (65%) human insulinomas. Furthermore, evaluation of Congo red-stained tumor sections showed small, globular or irregular, congophilic amyloid deposits within the cytoplasm of many tumor cells in 10 of 13 (77%) amyloid-containing insulinomas. Dense, punctate areas of IAPP immunoreactivity within tumor cells corresponded with the congophilic intracellular deposits. Ubiquitin immunoreactivity also was observed as punctate intracellular labeling and within large extracellular amyloid deposits. Among the 10 insulinomas available for electron microscopic evaluation, pathological IAPP-immunoreactive (immunogold) deposits were found in 3 of 5 insulinomas in which amyloid was demonstrated by light microscopy and in none of 5 tumors found negative for amyloid by light microscopy. Morphology of IAPP-immunoreactive deposits varied from those with the classical distinct 7- to 10-nm diameter nonbranching fibrils to those with distinct but faint fibrillarity to those without discernable fibrils. In each of the 3 tumors with ultrastructurally demonstrable amyloid deposits, intracellular aggregates of IAPP-immunoreactive amyloid fibrils were observed either free in the cytoplasmic matrix or in membrane-bound structures. Intracellular IAPP-immunoreactive deposits were up to 10 urn in the greatest dimension. Free intracellular aggregates of amyloid fibrils sometimes contained degenerating secretory vesicles and fragments of membranous organelles. These observations suggest that IAPP-derived amyloid deposits are initially formed intracellularly and are subsequently released to the extracellular space by exocytosis of the membrane-bound structures and/or after necrosis of the tumor cells. Our observations raise the possibility that IAPP-derived islet amyloid deposits (characteristically present in NIODM patients) also may be initially formed within the cytoplasm of β-cells, thus leading to β-cell necrosis and a reduction of β-cell mass as seen in NIDDM.

Both glucose and insulin are important regulators of glucose uptake and hepatic glucose release. Because insulin concentrations rarely if ever increase under daily living conditions, unless glucose concentrations also increase, we sought to determine whether hepatic and extrahepatic responses to changes in insulin and glucose concentration are impaired in patients with non-insulin-dependent diabetes mellitus (NIDDM). To address this question, glucose metabolism was measured in diabetic and nondiabetic subjects. A computer-driven infusion system was used to produce a nondiabetic postprandial insulin profile in both groups while sufficient exogenous glucose was infused to mimic nondiabetic postprandial glucose concentrations. Although NIDDM was associated with greater (P < 0.05) hepatic glucose release both before and during the prandial insulin infusion, suppression did not differ in the diabetic and nondiabetic subjects (−1.06 ± 0.20 vs. −0.86 ± 0.15 mmol/kg every 4 h). In contrast, stimulation of both glucose disappearance (0.77 ± 0.27 vs. 1.68 ± 0.27 mmol/kg every 4 h) and forearm glucose uptake (187 ± 81 vs. 550 ± 149 μmol/dl every 4 h) was lower (P < 0.05) in diabetic than in nondiabetic subjects. Thus, despite increased basal rates of glucose production, obese individuals with NIDDM had decreased stimulation of glucose disappearance but normal suppression of hepatic glucose release in response to nondiabetic prandial glucose and insulin concentrations. These data indicate that the increase in glucose that occurs with carbohydrate ingestion is likely to compensate for hepatic but not extrahepatic insulin resistance.

We previously reported the presence of endoneurial hypoxia, ischemia, impairment of the blood-nerve barrier, and reduction of norepinephrine and 6-ketoprostaglandin F1α in chronic streptozocin-induced diabetic neuropathy (SDN) and interpreted these findings as suggesting the involvement of oxygen free radicals (OFRs) but did not directly measure indices of OFR activity. In this study, we report on sciatic nerve conjugated dienes, hydroperoxides, norepinephrine, and malondialdehyde in SDN at 1, 4, and 12 mo in male Sprague-Dawley rats. Severe hyperglycemia was present throughout in diabetic rats. Conjugated dienes were consistently increased at all time points, hydroperoxides were consistently reduced, and malondialdehyde was not significantly different in diabetes compared with controls. These findings are consistent with increased OFR activity in experimental diabetes. It is necessary to monitor several indices of OFR activity in a metabolically active tissue such as the peripheral nerve.

Excessive amounts of glucose enter the systemic circulation when patients with non-insulin-dependent diabetes mellitus (NIDDM) eat a carbohydrate-containing meal. To determine the contribution of hepatic glucose cycling (defined as the net effect of glucose/glucose-6-phosphate cycling and uptake and release of glucose from hepatic glycogen) to postprandial hyperglycemia, diabetic, glucose-intolerant, and nondiabetic subjects were fed mixed meals. The meal contained both [2-3H]glucose (an isotope that is extensively detritiated during hepatic glucose cycling) and [6-3H]glucose (an isotope that is not detritiated during hepatic glucose cycling). Of the 50 g of carbohydrate contained in the meal, ∼4–8 g underwent hepatic glucose cycling. Although total cycling of ingested glucose did not differ between diabetic, glucose-intolerant, and nondiabetic subjects (361 ± 67 vs. 494 ± 106 vs. 322 ± 44 μmol · kg−1 · 5 h−1, respectively), the data suggested that hepatic cycling was increased in the diabetic and glucose-intolerant individuals but not in the nondiabetic subjects during the first 2 h after eating. Hepatic cycling during the first 2 h after eating was correlated with the prevailing glucagon concentration (r = 0.6, P < 0.01) and increased (P < 0.05) as hepatic glucose release increased. Hepatic glucose cycling had a marked effect on the measurement of so-called initial splanchnic glucose uptake. Nevertheless, however measured, initial splanchnic glucose uptake was not decreased and, if anything, was increased in diabetic and glucose-intolerant patients. Integrated postprandial hepatic glucose release increased (r < 0.01) with the severity of fasting hyperglycemia. These results indicate that postprandial hyperglycemia in diabetic and glucose-intolerant patients is not due to enhanced hepatic glucose cycling or a lack of uptake of ingested glucose by the splanchnic bed but rather to excessive release of glucose by the liver coupled with a failure of extrahepatic tissues to appropriately increase glucose disposal.

Human growth hormone (hGH) and prednisone cause insulin resistance and glucose intolerance. However, it is unknown whether hGH and prednisone antagonize insulin action on protein, fat, and carbohydrate metabolism by a common or independent mechanism. Therefore, protein, fat, and carbohydrate metabolism was assessed simultaneously in four groups of eight subjects each after 7 days of placebo, recombinant DNA hGH (rhGH; 0.1 mg · kg−1 · day−1), prednisone (0.8 mg · kg−1 · day−1), or rhGH and prednisone administration after an 18-h fast and during gut infusion of glucose and amino acids (fed state). Fasting plasma glucose concentrations were similar during placebo and rhGH but elevated (P < 0.001) during combined treatment, whereas plasma insulin concentrations were higher (237 ± 57 pmol/ml, P < 0.001) during combined than during placebo, rhGH, or prednisone treatment (34, 52, and 91 pM, respectively). In the fed state, plasma glucose concentrations were elevated only during combined treatment (11.3 ± 2.1 mM, P < 0.001). Plasma insulin concentrations were elevated during therapy with prednisone alone and rhGH alone (667 ± 72 and 564 ± 65 pmol/ml, respectively, P < 0.001) compared with placebo (226 ± 44 pmol/ml) but lower than with the combined rhGH and prednisone treatment (1249 ± 54 pmol/ml, P < 0.01). Protein oxidation ([14C]leucine) increased (P < 0.001) with prednisone therapy, decreased (P < 0.001) with rhGH treatment, and was normal during the combined treatment. By indirect calorimetry, glucose oxidation was similar in all groups; fasting fat oxidation was decreased by prednisone (P < 0.001) and increased by rhGH, although only significantly in the fed state (P < 0.01). In summary, insulin antagonism of rhGH and prednisone are probably caused by independent mechanisms in which rhGH and prednisone may reciprocally regulate the oxidation of protein and fat while decreasing the efficiency of glucose disposal.