Beta-cell tropin- and glucose-induced hypersecretion of insulin and amylin from perfused fatty rat pancreas.

The neurointermediate pituitary peptide beta-cell tropin (BCT) has potent insulin-releasing and lipogenic properties and is elevated in obesity and type-2 diabetes. The effects of BCT and glucose on the release of insulin and amylin from the perfused pancreas of obese 'fatty' (fa/fa) rats and lean (Fa/?) controls were measured. Pancreata were perfused, sequentially, with buffer containing: 5.6 mmol glucose/l (basal); basal glucose +/- 0.5 nmol BCT/l; 16.7 mmol glucose/l (high). Insulin and amylin release during basal glucose treatment was eight to nine times greater from pancreata from fatty than from lean rats. BCT induced a fivefold greater monophasic insulin and amylin release from fatty compared with lean pancreata. When not preceded by BCT there was a twofold greater high glucose-induced amylin release from fatty pancreata but no difference in insulin secretion. When preceded by BCT stimulation, high glucose induced twofold greater insulin and fourfold larger amylin release from fatty compared with lean pancreata. Molar secretion ratios of insulin:amylin varied between 30:1 and 50:1. In view of the elevated levels of BCT found in the fatty rat and in the light of the above findings, it is concluded that the peptide may have a role in the development of hyperinsulinaemia, hyperamylinaemia and insulin resistance in this animal model of obesity and diabetes.

Elevated plasma concentrations of beta-cell tropin (ACTH22-39) in diet-treated type II diabetic patients.

beta-Cell tropin, the pituitary peptide ACTH22-39, is a potent insulin secretagogue and stimulates lipogenesis in adipose tissue in rodents. Plasma beta-cell tropin was measured fasting and after glucose infusion (5 mg.kg glucose ideal body weight-1.min0-1 for 90 min) in 10 mild diet-treated non-insulin-dependent (type II) diabetic subjects and 10 control subjects (body mass index) (BMI): 26.4 +/- 3.2 and 24.1 +/- 2.0 kg/m-2, NS, fasting plasma glucose 7.8 +/- 2.7 mM and 4.7 +/- 0.3 mM, respectively). The diabetic subjects had raised fasting plasma beta-cell tropin compared with the normal subjects (geometric mean (1 SD range): 0.49 (0.25-0.96) nM and 0.17 (0.10-0.28) nM, respectively, P = 0.007). In response to the glucose infusion, plasma glucose rose higher in the diabetic subjects (mean +/- 1 SD: 13.7 +/- 3.1 and 9.6 +/- 0.9 mM, P = 0.007) and plasma insulin was impaired in the diabetic compared with the nondiabetic subjects (geometric mean (1 SD range): 14 (8-26) and 34 (18-63), P less than 0.01). beta-Cell tropin concentrations in the diabetic subjects rose to 1.31 (0.74-2.30) nM (P = 0.007), whereas beta-cell tropin did not change in the normal subjects at 0.19 (0.11-0.91) nM. There was no overlap between glucose-stimulated plasma beta-cell tropin in the two groups (P = 0.0002). Pituitary-adrenal function, as assessed by plasma cortisol, did not differ between the two groups when fasting and did not change after the glucose infusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Correlation between plasma beta-cell tropin concentrations and body weight in obese rhesus monkeys.

The fasting plasma concentration of the pituitary peptide beta-cell tropin [beta-CT, adrenocorticotropic hormone-(22-39)] was measured in 17 rhesus monkeys from a colony known to develop spontaneous obesity. The weight of the animals was 9.4-23.9 kg (12-46% body fat). Plasma beta-CT concentrations were 0.03-0.84 nmol/l and were strongly correlated with body weight (P = 0.014, r = 0.584). Plasma beta-CT was also correlated with plasma insulin concentration as a power function (P = 0.011, r = 0.600) and with percent body fat up to 40% (P = 0.003, r = 0.0804). Plasma insulin is also correlated with body weight (P = 0.015, r = 0.578) but does not decline when body fat is in excess of 40%, supporting the hypothesis that beta-CT may be involved in a feed-back control mechanism, perhaps mediated by insulin. Because beta-CT has been shown in rodent studies to be a potent insulin secretagogue and lipogenic agent, it is possible that beta-CT is causally involved in the development of obesity and that there may be central determinants of obesity mediated through pituitary secretion of beta-CT.

The effects of insulin and the pituitary peptide beta-cell tropin on the incorporation of D-3-3H-glucose into lipid in brown adipocytes from lactating and non-lactating rats.

Lactating and non-lactating rat brown adipocytes were used to study the dose-dependent stimulation of lipogenesis by Beta-cell tropin (BCT) and insulin. In non-lactating animals BCT increased lipogenesis approximately 2-fold compared to a 3-fold stimulation with insulin; however BCT was effective at a substantially lower molar concentration than insulin. In lactating animals resistance was observed to both BCT and insulin action.

The effects of structural modifications on the insulin-releasing activity of beta-cell-tropin.

The minimal effective concentration of the pituitary insulin secretagogue beta-cell-tropin (beta-CT) on the in vitro perfused pancreas was established and the effects of various modifications of the peptide on its potency were tested: iodination with 127I and acetylation reduced the insulin-releasing activity of beta-cell-tropin, and the C-terminal fragments beta-CT-(2-18), beta-CT-(3-18) and beta-CT-(6-18) were all less potent than the intact molecule; beta-CT-(1-6) was not active and did not inhibit beta-CT-induced insulin release.

Studies on insulin secretion and the pituitary insulin secretagogue beta-cell-tropin in the sand-rat (Psammomys obesus).

The sand-rat (Psammomys obesus) is an animal model for the study of human maturity onset diabetes which appears to be controlled by caloric intake. In the present investigations, these animals have been studied in relation to the influence of low- and high-energy diets on body weight, plasma insulin and blood glucose levels, and on insulin secretion from the perfused pancreas and the secretion of corticotropin-like intermediate lobe peptide (CLIP, ACTH18-39) and the insulin secretagogue beta-cell-tropin (beta-CT, ACTH22-39) from the pituitary neurointermediate lobe. The sand-rats maintained on the high-energy diet all became obese. Insulin secretion from the perfused pancreas of the obese sand-rat in the presence of 5.6 mM glucose was significantly higher than in the lean controls maintained on low-energy diets. Increasing the glucose concentration to 16.7 mM only produced a small stimulation of insulin secretion in the obese animals, and the difference between the two groups was not significant. Stimulation of insulin secretion by beta-CT was variable, but the obese animals appeared to be more responsive. Pituitary neurointermediate lobes were incubated for 4 h to measure the secretion of the ACTH related peptide. These were separated by gel filtration and the concentrations measured by radioimmunoassay with a CLIP antiserum and a CLIP standard. In all experiments beta-CT was 4-6 per cent of the total CLIP immunoreactive material. In these experiments the obese animals maintained on a high-energy diet were divided into two groups, those with plasma insulin levels less than 500 mu u/ml and those with insulin levels greater than 500 mu u/ml. The latter group had a significantly higher blood glucose level, presumably due to the insulin resistance resulting from the severe hyperinsulinaemia. It was also observed that CLIP-IRM and beta-CT secretion was lower in this group than in the animals maintained on low-energy diets or those on high-energy diets with moderate hyperinsulinaemia. This suggests a possible feedback inhibition by insulin on the secretion of beta-CT.

Evidence for the presence of the pituitary insulin secretagogue beta-cell trophin in human plasma.

It has been demonstrated that the insulin secretagogue beta-cell-trophin, ACTH(22-39), is present in human plasma. The hormone, separated from plasma by affinity chromatography on a corticotrophin-like intermediate-lobe peptide antibody column, behaves similarly to synthetic beta-cell-trophin on a gel filtration column and on reverse-phase high-performance liquid chromatography. Sufficient amounts of the hormone were isolated from the plasma of two patients with Nelson's syndrome to demonstrate its biological activity on the perfused rat pancreas.

The insulin-like action of beta-cell-tropin on glucose and lipid metabolism in adipocytes.

The insulin-like activity of the pituitary pars-intermedia insulin secretagogue beta-cell-tropin, ACTH22-39, has been studied on rat adipocytes. The peptide was prepared by tryptic digestion of synthetic human CLIP, ACTH18-39. beta-Cell-tropin stimulated the incorporation of 3H2O into total lipids. The 50% maximal activity concentration was 5 X 10(-2) ng/ml-1 about 2.5 X 10(-11) M. Iodination of tyrosine, the penultimate amino-acid of the N-terminal, eliminated lipogenic activity, and acetylation of the N-terminal valine reduced activity. ACTH and CLIP (ACTH18-39) had no lipogenic action on the adipocyte system studied. beta-Cell-tropin stimulated the oxidation of glucose and the conversion of glucose into saponified fatty acids and glyceride glycerol. The influence of beta-cell-tropin and insulin on the incorporation of glucose into total lipids, saponified fatty acids and glyceride glycerol was not additive. The results suggest that beta-cell-tropin is either a potent lipogenic hormone, stimulating the conversion of glucose into lipids or that it activates endogenous insulin. The biological activity is associated with the N-terminal amino-acids of the peptide. The possible significance of beta-cell-tropin in obesity is discussed.

Beta-cell tropin: synthesis and biological activity.

The structure of beta-cell tropin, an insulin secretagogue released by the neuro-intermediate lobe of the obese (ob/ob) mouse, has recently been determined as the 22-39 moiety of ACTH. A method for the preparation of this octadecapeptide using mild solid-phase procedures followed by preparative high pressure liquid chromatography is described. The molecular weight of the synthetic peptide has been confirmed by Fast Atom Bombardment mass spectrometry. Synthetic beta-cell tropin is indistinguishable in its chromatographic, antigenic and biological properties from natural beta-cell tropin.

Evidence that the insulin secretagogue, beta-cell-tropin, is ACTH22-39.

The pituitary neurointermediate lobe of genetically obese (ob/ob) mice contains a hormone which stimulates insulin release and which cross-reacts with a -COOH-terminal ACTH antiserum, suggesting that it is related to corticotropin-like intermediate lobe peptide (CLIP), the 18-39 fragment of ACTH. The hormone, which we have called beta-cell-tropin, has been shown to be present in the plasma of the ob/ob mouse and to potentiate glucose induced insulin secretion. We have now shown that ACTH22-39 prepared by tryptic digestion of human synthetic CLIP behaves similarly on Biogel chromatography and on reverse-phase HPLC to the naturally occurring beta-cell-tropin. Furthermore, beta-cell-tropin and ACTH22-39 have indistinguishable antigenic and insulin releasing properties.

Identification of beta-cell-trophin, a peptide of the pituitary pars intermedia which stimulates insulin secretion in plasma from genetically obese (ob/ob) mice.

The influence of plasma from genetically obese (ob/ob) and lean (+/+) mice on insulin secretion has been studied by perifusion of collagenase-prepared pancreatic islets maintained for 48 h in culture. Insulin secretion was measured at 2-min intervals and plasma from the ob/ob mice not from the +/+ mice rapidly stimulated insulin release, reaching a maximum in 2-4 min and falling to basal levels in about 10 min. Experimental evidence is given indicating that the plasma insulin secretagogue is identical to beta-cell-trophin, a peptide of the pituitary pars intermedia which stimulates insulin secretion. The evidence is based on (1) the antigenic properties of the peptides (both cross-react with a -COOH terminal ACTH antiserum raised to the 17-39 moiety of ACTH), (2) identical chromatographic separation on Biogel columns and on reverse-phase high pressure liquid chromatography and (3) the similarity of their insulin releasing action from perifused islets.

Insulin release from the perfused rat pancreas, stimulated by perifusates of the pituitary neurointermediate lobes of genetically obese and lean mice.

The perfused rat pancreas has been used to study the dynamics of insulin release during stimulation with a perifusate of the pituitary neurointermediate lobe (NIP) of genetically obese mice (ob/ob) and their lean litter-mates (+/+). It has been shown that the NIP of ob/ob mice is more active. The remainder of the study was carried out with NIP from ob/ob mice. The results showed that both phases of insulin release stimulated by 16.6 mM-glucose were increased by NIP. When the pancrease was first stimulated with NIP in the presence of 5.5 mM-glucose, followed by a high concentration of glucose, it responded to the latter with a normal biphasic response. If, however, the pancreas was stimulated first with glucose the response to NIP was refractory. The results are discussed in relation to the possible significance of an insulin secretagogue from the pituitary pars intermedia in the hyperinsulinaemia associated with obesity.

Effect of substrate interactions on in vitro fatty acid synthesis in adipose tissue of normal and genetically obese mice.

Fatty acid synthesis was measured in vitro in pieces of adipose tissue from lean and obese-hyperglycaemic (ob/ob) mice, using 14C-glucose or 14C-lactate and 3H2O to obtain absolute rates of total fatty acid synthesis. In the presence of lipoprotein-triglyceride (2.5 mumol/l) metabolic interaction occurred which decreased glucose incorporation into fatty acids by 30% in lean mouse tissue, but not in obese mouse tissue. In the absence of added insulin, the contribution of glucose to total fatty acid synthesis was 69% in obese mouse tissue, significantly lower than the value of 87% obtained in lean mouse tissue. Insulin increased the contribution of glucose to total synthesis in both lean and obese mouse tissues, although the value in obese mouse tissue (83%) remained lower than the value in lean mouse tissue (100%). Lactate was not a major precursor for fatty acid synthesis. When both lactate (2 mmol/l) and glucose (15 mmol/l) were present, the contribution of lactate to total fatty acid synthesis was not increased in obese mouse tissue, suggesting that even in the presence of insulin, about 30% of the carbon was provided by intracellular precursors.

Acute regulation of insulin release by the pituitary gland in relation to hyperinsulinaemia and obesity.

The pituitary glands from mice rendered obese by gold thioglucose treatment and by dietary manipulation, and pituitary glands from lean mice after a high food intake or a glucose load, were shown to stimulate insulin secretion from isolated pancreatic islets. The insulin releasing activity of pituitary glands from obese (ob/ob) mice was reduced by fasting for 24 and 48 h. Results obtained with pituitary glands from ob/ob and from lean ob/+ and +/+ mice suggest that the insulin releasing property manifests a gene dosage effect. Pituitary glands from 3-week-old (young) ob/ob mice stimulated insulin secretion to the same extent as pituitary glands from 3-month-old (adult) ob/ob mice. The pancreatic islets of young ob/ob mice were shown to be somewhat more responsive to stimulation by the pituitary factor than were lean ob/+ or +/+ islets from this age group. The concept that high insulin level, partly under pituitary control, and high caloric intake may be interlinked and may, in combination, be a major factor in producing obesity is discussed. Furthermore, it is suggested that the pituitary insulin releasing factor may play a role in the early development of obesity in the animal models studied.