B cell adrenoceptors and sulphonylurea-induced insulin release in mouse islets

Summary Interactions of tolbutamide and glibenclamide with B cell adrenoceptors have been reported. This study evaluated the possible role of such interactions in the stimulation of insulin release. Mouse islets were incubated in the presence of 10 mmol/l glucose alone or with tolbutamide (10 mol/l) or glibenclamide (0.02 mol/l). At 0.01–10 mol/l, blockers of ₂-adrenoceptors (yohimbine, idazoxan) or ₁-adrenoceptors (prazosin) had practically no effect on glucose-induced insulin release and did not affect its potentiation by sulphonylureas, except for a slight increase by 10 mol/l prazosin and idazoxan. Nonspecific -blockers (phentolamine, dihydroergotamine) increased control release at 10 mol/l, but only the latter amplified the response to tolbutamide. Blockers of -adrenoceptors were tested at 0.1–100 mol/l: propranolol (₁, ₂), metoprolol (₁) and compound ICI 118-551 (₂). They increased glucose-induced insulin release at 100 mol/l but variably altered the effect of sulphonylureas. Blockers of adrenoceptors have, thus, no effect on insulin release in vitro at therapeutic concentrations. At high concentrations, they non-specifically affect the action of sulphonylureas. We conclude that an interaction with B cell adrenoceptors is not involved in the insulinotropic action of sulphonylureas.     

Effect of insulin on GLUT-4 mRNA and protein concentrations in skeletal muscle of patients with NIDDM and their first-degree relatives

Summary We examined whether insulin resistance, i. e. impaired insulin stimulated glucose uptake in NIDDM patients and their first-degree relatives is associated with alterations in the effect of insulin on the expression of the GLUT-4 gene in skeletal muscle in vivo. Levels of GLUT-4 mRNA and protein were measured in muscle biopsies taken before and after a euglycaemic insulin clamp from 14 NIDDM patients, 13 of their first-degree relatives and 17 control subjects. Insulin stimulated glucose uptake was decreased in the diabetic subjects (19.8±3.0 mol · kg LBM^–1 · min^–1, both p<0.001) compared with control subjects (44.1±2.5 mol · kg LBM^–1 · min^–1) and relatives (39.9±3.3 mol · kg LBM^–1 · min^–1). Basal GLUT-4 mRNA levels were significantly higher in diabetic subjects and relatives compared to control subjects (99±8 and 108±9 pg/g RNA vs 68±5 pg/g RNA; both p<0.01). Insulin increased GLUT-4 mRNA levels in all control subjects (from 68±5 to 92±6 pg/ug RNA; p<0.0001), but not in the diabetic patients (from 99±8 to 90±8 pg/g RNA, NS), or their relatives (from 94±9 to 101±11 pg/g RNA, NS). In the relatives, individual basal GLUT-4 mRNA concentrations varied between 55 and 137 pg/g RNA. Insulin-resistant (n=6, mean glucose uptake rate=30.6±3.4 mol · kg LBM^–1 · min^–1) but not insulin-sensitive relatives (n=7, mean glucose uptake rate=47.4±3.2 mol · kg LBM^–1 · min^–1) had higher basal GLUT-4 mRNA concentrations compared to control subjects (108±9 vs 68±5 pg/ug RNA, p<0.01). GLUT-4 protein content in muscle did not differ between the groups in the basal state and remained unchanged in all groups after insulin infusion. Neither insulin-stimulated GLUT-4 mRNA nor protein concentrations correlated with insulin-stimulated glucose uptake in any of the groups studied. We conclude, that impaired glucose uptake in NIDDM is not related to insulin-stimulated GLUT-4 mRNA or protein concentrations. Acute stimulation of GLUT-4 mRNA by insulin is altered in skeletal muscle of NIDDM patients and their first-degree relatives. This might be a consequence of chronic hyperinsulinaemia elevating basal GLUT-4 mRNA concentrations rather than the cause of insulin resistance.Abbreviations NIDDM Non-insulin dependent diabetes mellitus - GLUT-4 glucose transporter 4 - LBM lean body mass - IDDM insulin-dependent diabetes mellitus     

Iron stores in 1359, 30- to 60-year-old Danish women: Evaluation by serum ferritin and hemoglobin

Summary Iron status, including serum (S-)ferritin and hemoglobin (Hb), was assessed in a population survey comprising 1359 nonpregnant Danish women in age cohorts of 30, 40, 50, and 60 years. S-ferritin levels were similar in 30- and 40-year-old women; they displayed a significant increase in 50-year-old women and a further significant increase in 60-year-old women. In the 30- and 40-year-old women, median S-ferritin was 38g/l, 5–95 percentile 6–135g/l; 17.2% had values < 15,g/l (i.e., depleted iron stores), 22.7% values from 15 to 30g/l (i.e., small iron stores), and 60.1% values > 30g/l (i.e., replete iron stores). In the 50-year-old women, median S-ferritin was 54g/l, 5–95 percentile 10–164g/l; 10.3% had values < 15g/l, 16.5% values from 15 to 30g/l, and 73.2% values > 30g/l. For the 60-year-old women, median S-ferritin was 84g/l, 5–95 percentile 25–249g/l; 1.6% had values < 15g/l, 8.6% values from 15 to 30g/l, and 89.8% values > 30g/l. Blood donors (n=180) had lower S-ferritin than nondonors in all age-groups (p<0.001). In the entire series, Hb levels were similar in 30- and 40-year-old women, median 137 g/l (8.5 mmol/l), 5–95 percentile 121–152 g/1 (7.5–9.4 mmol/l), and higher in 50- and 60-year-old women, median 140 g/l (8.7 mmol/l), 5–95 percentile 123-158 g/l (7.6–9.8 mmol/l) (p<0.0001). Hb values < 121 g/l (7.5 mmol/l) were observed in 3.8% of the women. Women with S-ferritin < 15 g/l (n=161) had lower Hb, median 134 g/l (8.3 mmol/l), than those with S-ferritin > 15 g/l, median 139 g/l (8.6 mmol/l) (p<0.001). Iron deficiency anemia (S-ferritin < 15 g/l and Hb < 121 g/l) was seen in 2.3% of 30- and 40-year-old women, and in 1.1% of 50- and 60-year-old women.     

Modulation of hepatic glucose production by non-esterified fatty acids in Type 2 (non-insulin-dependent) diabetes mellitus

Summary To study the effect of changes in plasma non-esterified fatty acid concentration on suppression of hepatic glucose production by insulin eight Type 2 (non-insulin-dependent) diabetic patients participated in three euglycaemic, hyperinsulinaemic (108pmol · m^2–1 · min^–1) clamp studies combined with indirect calorimetry and infusion of [3-³H]-glucose and [1-¹⁴C]palmitate; (1) a control experiment with infusion of NaCl 154 mmol/l, (2) heparin was infused together with insulin, and (3) an antilipolytic agent, Acipimox, was administered at the beginning of the experiment. Six healthy volunteers participated in the control experiment. Plasma non-esterified fatty acid concentrations during the insulin clamp were in diabetic patients: (1) 151±36 mol/1, (2) 949±178 mol/l, and (3) 65±9 mol/l; in healthy control subjects 93±13 mol/l. Non-esterified fatty acid transport rate, oxidation and non-oxidative metabolism were significantly higher during the heparin than during the Acipimox experiment (p<0.001). Suppression of hepatic glucose production by insulin was impaired in the diabetic compared to control subjects (255±42 vs 51±29 mol/min, p<0.01). Infusion of heparin did not affect the suppression of hepatic glucose production by insulin (231±49 mol/min), whereas Acipimox significantly enhanced the suppression (21±53 mol/min, p<0.001 vs 154 mmol/l NaCl experiment). We conclude that insulin-mediated suppression of hepatic glucose production is not affected by increased non-esterified fatty acid availability. In contrast, decreased non-esterified fatty acid availability enhances the suppression of hepatic glucose production by insulin.     

Experimental streptozotocin-induced diabetes and intestinal glucose metabolism in the rat,in vivo and in vitro

The intestine has a high glycolytic activity, but its metabolic role could be altered in diabetes mellitus. The aim of the present work was to investigate in vivo the glucose retained and the lactate produced by the intestine of normal and diabetic rats and in vitro the effect of different arterial glucose concentrations on glucose utilization and lactate, alanine, and pyruvate production in normal and diabetic rats when the glucose is supplied to the intestine exclusively via the vascular route. In vivo, the normal and diabetic rats retained similar percentages of the arterially supplied glucose (14.7±3.2 and 12.6±2.4, respectively). In vitro, when the preparations were perfused under hyperglycemic conditions, the glucose consumed, as a fraction of the quantity infused, was significantly lower (P<0.05) in the diabetic (247.0±22.8 mol/mmol infused glucose) than in normal (315.0±16.3 mol/mmol infused glucose) rats. The lactate produced was significantly higher in diabetic than in normal rats whether the preparations were perfused under isoglycemic (P<0.01; 1916.4±124.0 vs 1284±67.7 mol/mmol consumed glucose) or hyperglycemic (P<0.05; 1356.4±199.7 vs 898.0±87.3 mol/mmol consumed glucose) conditions. There was significantly (P<0.05) greater alanine release from the diabetic (123.7±21.8 mol/mmol consumed glucose) than from the normal (40.7±10.3 mol/mmol consumed glucose) rat preparations perfused under isoglycemic conditions.     

Catecholamines and tumour promoting phorbolesters inhibit insulin receptor kinase and induce insulin resistance in isolated human adipocytes

Summary The effect of the catecholamine isoprenaline (10^–5mol/l) and of the tumour promoting phorbolester tetradecanoyl--phorbol acetate (10^–9mol/l) on insulin stimulated 3-O-methyl-glucose transport was studied in freshly isolated human adipocytes. Both substances reduced the maximal responsiveness of the glucose transport system to insulin by approximately 50%. To test if this is caused by inhibition of the insulin receptor kinase the receptor from phorbolester and isoprenaline treated cells was solubilized, partially purified and its kinase activity studied in vitro. Insulin stimulated ³²P-incorporation into the -subunit of the insulin receptor of phorbolester or isoprenaline treated cells was reduced to 20–60% of the values found with receptor from control cells at insulin concentrations between 10^–10mol/l and 10^–7mol/l. This inhibition of kinase activity of receptor from phorbolester and isoprenaline treated cells was observed at nonsaturating adenosine triphosphate levels (5 mol/l), and it could be overcome with higher concentrations of -³²P-adenosine triphosphate in the phosphorylation assay. A Lineweaver Burk analysis of the insulin stimulated receptor phosphorylation revealed that the Michaelis constant for adenosine triphosphate of the receptor kinase from phorbolester and isoprenaline treated cells was increased to >100 mol/l compared with <50 mol/l for receptor from control cells. We conclude from the data that catecholamine and phorbolester treatment of human adipocytes modulates the kinase activity of the insulin receptor by increasing its Michaelis constant for adenosine-triphosphate, and propose that this modulation of receptor kinase is a mechanism that can contribute to the pathogenesis of insulin resistance in human fat cells.     

Pharmacological evidence for beta3 adrenoceptors in the control of rat gastric acid secretion

The effect of the ₃-adrenoceptor agonist BRL37344 on gastric acid secretion evoked by different secretory stimuli was investigated in anaesthetized rats with lumen-perfused stomachs in comparison with the ₂-adrenoceptor agonist clenbuterol. Intravenous injections of BRL37344 (1–10 mol/kg) and clenbuterol (0.01–1 mol/kg) dose-dependently reduced 2-deoxy-D-glucose-induced acid secretion, with BRL37344 about forty times less potent than clenbuterol. BRL37344 (0.1–3 mol/kg) inhibited pentagastrin-induced acid output, whereas clenbuterol was effective only at high doses (10–100 mol/kg). The inhibitory effect of BRL37344 on pentagastrin-induced acid secretion was not modified by the nonselective –adrenoceptor antagonist propranolol, but it was prevented by bupranolol, a ₃-adrenoceptor antagonist. Furthermore, neither BRL37344 (10 mol/kg) nor clenbuterol (100 mol/kg) modified the acid secretion induced by histamine. These data suggest that ₃ adrenoceptors have an inhibitory role in the control of rat gastric acid secretion induced by indirect stimuli.     

Modulation of pacemaker activity in sheep cardiac Purkinje fibers by stimulation of β-adrenoceptor subtypes

The electrophysiological effects mediated by ₁- and ₂- in spontaneously active sheep cardiac Purkinje fibers were investigated using the non-selective agonist (–)-isoproterenol (IPN) and the selective agonists (–)-noradrenaline (₁) and procaterol (₂) in the absence and presence of the selective antagonists bisoprolol (₁) and ICI 118,551 (₂).IPN (0.01 mol/l) increased the spontaneous rate by 54% and the slope of diastolic depolarization by 68% of the respective control values. Further, IPN increased the action potential duration at –20 mV (APD –20 mV) from 96 to 154 ms, reduced the APD –70 mV by 17% and the duration of the diastole by 39% and slightly hyperpolarized the maximum diastolic potential. These effects were partially inhibited by ICI 118,551 (0.03 mol/l), diminished by bisoprolol (0.1 mol/l) and almost completely blocked by the combination of both antagonists. Concentration response curves of IPN were influenced by the selective antagonists as follows: ICI 118,551 (0.03 mol/l) shifted the curves to the right by 0.2–0.4 log units and increased the slope factor. Bisoprolol (0.1 mol/l) induced a greater shift to the right by 1.1–1.5 log units. Combination of bisoprolol with ICI 118,551 shifted the curves to the right by 1.5–1.7 log units.Noradrenaline (0.3 mol/l) elicited similar actions as IPN. Bisoprolol (0.1 mol/l) shifted the concentration response curves of noradrenaline to the right by 1.1–1.9 log units. Actions of procaterol (0.1 mol/l) were weak, attained only 15–35% of the maximal effects of IPN and could be blocked by ICI 118,551 (0.03 mol/l).These results show that the increase of pacemaker activity induced by catecholamines in sheep cardiac Purkinje fibers is predominantly mediated by stimulation of ₁. However, contribution of ₂ mediated effects could be demonstrated.Supported by Ministerium für Wissenschaft und Forschung, Nordrhein-Westfalen, Projekt-Nr, 40008786.     

The circulating molecular weight forms of infused recombinant insulin-like growth factor-I and effects on glucose and fat metabolism in lambs

Summary We have investigated the relationship between the plasma distribution of infused recominant insulin-like growth factor-I across the insulin-like growth factor binding proteins and the resultant effects on glucose and fat metabolism. The studies were performed in 24-h fasted ram lambs which received primed constant infusions of ³H labelled glucose tracer. When isotopic equilibrium had been reached, the animals received 90-min infusions of human insulin-like growth factor-I at various doses (2.5, 20, 40 and 120 g· kg^–1·h^–1, n=3 for each dose). Total plasma insulin-like growth factor-I was significantly elevated by infusion at a rate of 40 g·kg^–1·h^–1 (from 185±14 g/l to 442±41 g/l, p<0.05) and 120g·kg^–1h^–1 (from 181±2 g/l to 953±39 g/1, p<0.005). The plasma concentrations of insulin-like growth factor-I not associated with binding proteins remained undetectable (<15 g/l) at the end of the 2.5 and 20 g·kg^–1·h^–1 doses, but were significantly elevated at the end of the 40 and 120 g·kg^–1·h^–1 infusions (to 71±14 g/l, p<0.05 and 176±55 g/l, p<0.01 respectively). The infused insulin-like growth factor-I associated primarily with 35–60 kilodalton binding proteins. Glucose kinetics were significantly altered only by the highest dose infusion, during which there was a fall in plasma glucose concentration from 3.5±0.2 mmol/l to 1.9±0.2 mmol/l (p<0.05). This was due to a 51% increase in the rate of glucose clearance. There was no significant change in the rate of glucose production. The plasma concentrations of glycerol and non-esterified fatty acid were not changed by any of the doses infused. We conclude that the hypoglycaemic action of infused recombinant insulin-like growth factor-I relates to a marked elevation of free insulin-like growth factor-I in the plasma, but that a threshold concentration of free insulin-like growth factor-I must be exceeded before this action is observed. The hypoglycaemic action of recominant insulin-like growth factor-I results primarily from an increase in glucose clearance while glucose metabolism was more sensitive than fat metabolism to infused recominant insulin-like growth factor-I. Both these actions contrast with those of insulin, and suggest that the acute metabolic effects of recombinant insulin-like growth factor-I are not mediated simply by cross-reaction with insulin receptors.     

Developmental changes in the fatty (fafa) rat: Evidence for defective thermogenesis preceding the hyperlipogenesis and hyperinsulinaemia

Summary Preobese fatty rats have been identified by their lower rectal temperature. Of 51 pups born from matings of heterozygote (Fafa) parents, 16 had low rectal temperatures from day 16 onward (34.6±0.2° C v 35.4±0.3° C) and all subsequently became obese. No animal with the higher normal rectal temperature developed obesity. Hepatic fatty acid synthesis (preobese 0.6±0.1; lean 0.6±0.1 mol/ g/h), hepatic glucose-6-phosphate dehydrogenase activity (G6PDH) (preobese 0.68±0.07; lean 0.71 ±0.03 mol/g/min) and serum insulin (preobese 64 ±2; lean 58±4 U/ml) were unchanged in 18 day preobese, suckling fafa rats. 3 days after weaning hepatic lipogenesis (preobese 25.3±2.0; lean 5.4±0.7 mol/g/h) and G6PDH activity (preobese 4.5±0.5; lean 0.90±0.05 mol/g/min) had increased in both lean and preobese rats although the values attained in preobese rats were significantly greater than in lean rats. When weaning was delayed there was no enhancement in lipogenesis, G6PDH or serum insulin in the preobese rat. The results suggest that the primary genetic defect in fatty rats is not related to the increase in lipogenesis or serum insulin but may reflect a defective thermogenic process.     

Cellular ionic effects of insulin in normal human erythrocytes: a nuclear magnetic resonance study

Summary Elevated erythrocyte cytosolic free calcium, and suppressed free magnesium and pH values are associated with the hyperinsulinaemia and insulin resistance of hypertension, obesity, and Type 2 (non-insulin-dependent) diabetes mellitus. To determine the role of insulin in this process, we utilized ¹⁹F- and ³¹P-nuclear magnetic resonance spectroscopy to study the cellular ionic effects of insulin in vitro on normal human erythrocytes. Insulin elevated cytosolic free calcium levels in a dose- and time-dependent manner. The effect began at 10 U/ml, peaked at 200 U/ml, and continued at both the 500 U/ml and 1000 U/ml doses. At 200 U/ml, free calcium levels rose from 24.6±2.5 nmol/l to a peak value at 120 min of 66.4±11 nmol/l (p<0.05 vs basal), levels remaining elevated throughout the incubation (45.7±5.6 nmol/l at 60 min, and 47.9±9.1 nmol/l at 180 min, p<0.05 vs basal, respectively). Similarly, insulin also increased intracellular free magnesium at all time points (basal: 177± 11 mol/l; 60 min: 209±19 mol/l; 120 min: 206±22 mol/l; and 180 min: 202±12 mol/l; p<0.05 vs basal at all times). No insulin-induced changes in pH were observed. We conclude (i) that insulin in physiological concentrations may participate in regulating divalent cations in the mature human erythrocyte, (ii) that insulin per se cannot account for the previously described cellular ionic lesions of hypertension and diabetes, and (iii) that future clinical studies of cell ion metabolism should be conducted in the fasting state, be controlled for ambient circulating insulin levels, or both.     

Protective effect of vitamin E supplementation on increased thermal stability of collagen in diabetic rats

Summary Products similar to non-enzymatic glycation end products are known to arise from the interactions between proteins and lipid peroxides in vitro. In this study, we assessed the effect of vitamin E, which possibly modifies lipid peroxide, on advanced glycation end products or similar products in vivo by measuring the fluorescence and thermal rupture time of tail tendon collagen in streptozotocin-induced diabetic rats. The diabetic rats and non-diabetic rats were fed a vitamin E supplemented diet, and a control diet starting 3 days after the streptozotocin injection. After the 4-week treatment, the serum lipid peroxide levels expressed as thiobarbituric acid reactants in the diabetic rats on control diet (15.9 ± 2.6 mol/l [SEM]) were significantly (p <0.05) higher than in the non-diabetic rats (7.9 ± 1.3 mol/l on control diet and 3.3 ± 0.4 mol/l on supplemented diet), but the levels in the diabetic rats on supplemented diet (7.9 + 2.3 mol/l) were reduced to the normal levels. No significant differences were found in serum glucose and glycated haemoglobin levels within the diabetic rats on control and supplemented diets. Both the fluorescence and thermal rupture time of collagen were significantly (p <0.05) increased in the diabetic rats on both diets compared with those in the corresponding non-diabetic rats. Although there was no significant difference in the collagen-linked fluorescence within the diabetic rats on control and supplemented diets, the thermal rupture time was significantly (p <0.01) shortened with supplemented diet (10.8 ± 0.7 min on supplemented diet vs 15.0 ± 0.7 min on control diet). The effect of vitamin E on the thermal rupture time was not observed in non-diabetic rats (6.6 ± 0.5 min on supplemented diet vs 6.2 ± 0.5 min on control diet). These results indicate that the formation of advanced glycation end products or similar products seen in hyperglycaemia can be partially inhibited by vitamin E supplementation by lowering lipid peroxide levels or oxidative stress. This study is thought to be the first to show vitamin E as an anti-oxidant agent limiting the formation of advanced glycation end products or similar products in vivo.     

Alterations in cardiac oxygen consumption under chronic pressure overload

Summary Hypertension and resulting left ventricular hypertrophy was induced in young male Wistar rats (60 to 70 days old) by narrowing of one renal artery (Goldblatt II). 8 and 24 weeks after operation, myocrdial oxygen consumption was measured on a modified in situ heart-lung preparation with nearly isovolumetric left ventricular contractions. Measured myocardial oxygen consumption was related to left ventricular wall stress. The myosin isoenzyme pattern of each heart was determined with pyrophosphate gel electrophoresis.Oxygen consumption related to wall stress averaged over the entire heart cycle amounted to 15 moles O₂/g·min 8 weeks after operation, and 24.4 moles O₂/g·min in age-matched controls ( 38%, p<0.0005). When wall stress was averaged over systole, oxygen consumption of the hypertrophied hearts amounted to 0.112 moles O₂/g·beat, and 0.149 moles O₂/g·beat in the controls ( 25%, p<0.05). The proportion of VM-3 (the cardiac myosin isoenzyme of lowest ATPase activity) increased from 26.3% in the controls to 30.1% in the Goldblatt hearts ( 14%, n.s.).24 weeks after operation, oxygen consumption related to wall stress averaged over the entire heart cycle amounted to 16.1 moles O₂/g·min, in age-matched controls 20.5 moles O₂/g·min ( 21%, p<0.05). When wall stress was averaged over systole, oxygen consumption of the Goldblatt hearts amounted to 0.080 moles O₂/g·beat, and in the controls 0.107 moles O₂/g·beat ( 25%, p<0.005). The proportion of VM-3 increased from 33.5% in the controls to 43.2% in the hypertrophied hearts ( 29%, p<0.05).The present findings indicate that the reduced oxygen consumption of the pressure-loaded heart should be attributed to a redistribution of myosin isoenzymes. The transformation of myocardium into a slower, but more efficiently working muscle due to an increase in VM-3 can be interpreted as an adaptational process.Supported by the Deutsche Forschungsgemeinschaft     

Influence of phorbol esters on contractile force,action potential and calcium current of isolated guinea-pig heart tissues

Phorbol-12,13-dibutyrate (PDB) reduced concentration-dependently the contractile force of guineapig papillary muscles (EC₅₀ 1.07 mol/l) while phorbol-12-myristate-13-acetate (PMA) was ineffective. The protein kinase C inhibitors staurosporine (0.1 mol/l) and polymyxin B (70 mol/l) did not antagonize the negative inotropic effect of PDB. Neither PMA nor PDB, in concentrations up to 30 mol/l caused significant changes of the membrane resting potential, the maximum depolarization velocity, the action potential duration or the functional refractory period in intact papillary muscles. In isolated ventricular cardiomyocytes the inward calcium current was halved by either 1 mol/l PDB or 10 mol/l PMA. PKC inhibitors attenuated, but could not completely abolish this effect of the phorboles. It is concluded that the negative inotropic effect of PDB is caused by a reduction of the slow inward calcium current and that this inhibition is, for the greater part, not mediated by an activation of protein kinase C.     

The effect of monooleoylglycerol on insulin secretion from isolated perifused rat islets

Summary The effect of monooleoylglycerol on cholecystokinin-and tolbutamide-induced insulin secretion was examined in isolated perifused rat islets. In the presence of 5.5 mmol/l glucose, addition of 10 nmol/l cholecystokinin or 50 mol/l tolbutamide had practically no effect on insulin secretion. Combined tolbutamide and cholecystokinin led to a biphasic insulin secretory response which was significantly enhanced by addition of 50 mol/l monooleoylglycerol, an inhibitor of diacylglycerol kinase. Monooleoylglycerol (50 mol/l) alone had a minimal stimulatory effect on insulin release in the presence of 5.5 mmol/l glucose. Perifusion of islets with 1 mol/l forskolin had no significant effect on basal insulin secretion in the presence of 5.5 mmol/l glucose, but markedly enhanced the responses to both cholecystokinin plus tolbutamide, and to the combination of cholecystokinin, tolbutamide and monooleoylglycerol. Lowering the glucose level to 2.75 mmol/l abolished the profound stimulatory effect to these agonist combinations on insulin release. Finally, monooleoylglycerol also enhanced the first and second phase insulin secretory responses induced by 20 mmol/l glucose. These results are discussed in relationship to the possible role of protein kinase C in mediating insulin secretion.     

Inhibition of biliary phospholipid and cholesterol secretion by cefoperazone

The effect of cefoperazone, a third-generation cephalosporin, on biliary lipid secretion in rats was examined. Rats were anesthetized with ether and the mid-lumbar vein and common bile duct cannulated. Bile acid secretion was maintained by intravenous taurocholic acid infusion (28 mol/hr). A 1-hr control period was followed by intravenous cefoperazone infusion at either submaximal (20 mol/hr), or supramaximal (60 mol/hr) concentrations. At the cefoperazone infusion rate of 20 mol/hr (biliary secretion of 7.1±1.6 mol/hr) phospholipid secretion fell 19% and cholesterol secretion fell 31%; at a cefoperazone infusion rate of 60 mol/hr (biliary secretion rate of 27.1±5.1 mol/hr) phospholipid and cholesterol secretion were further reduced 40% and 56%, respectively, of controls. All changes were significant (P<0.01). Inhibition of both cholesterol and phospholipid secretion paralleled each other, was dose-dependent, and reversible. Cefoperazone's inhibitory action was abolished at a bile acid infusion rate of 108 mol/hr. Cefoperazone was not found to be associated with bile acid micelles or mixed micelles as determined by ultracentrifugation and gel filtration. Thus, the effect of cefoperazone on biliary lipid secretion is not due to the impairment of mixed micelle formation in the canalicular lumen but rather its inhibitory effect appears to be due to a presecretory event.     

Role of histamine H3 receptors in control of mouse intestinal motility in vivo and in vitro: comparison with alpha2-adrenoceptors

We tested drugs acting at histamine H₃ receptors in mice on the gastrointestinal transit of a charcoal meal in vivo and on neurogenic contractions of isolated ileal preparations. The agonist (R)--methylhistamine (100 mol/kg) caused a maximum 25% reduction of gastrointestinal transit, an effect mimicked by immepip (100 mol/kg) and antagonized by thioperamide (20 mol/kg) or clobenpropit (20 mol/kg). In the isolated ileum, (R)--methylhistamine (10–100 M) caused a slight, thioperamide-insensitive, reduction (maximum 15%) of electrically evoked cholinergic contractions. In comparison, the ₂-adrenoceptor agonist clonidine (0.1 mol/kg) caused a 35.2% inhibition of the gastrointestinal transit and almost completely reduced (maximum 82% at 1 M) the cholinergic contraction of the isolated ileum, both effects being antagonized by idazoxan (0.4 mol/kg and 1 M, respectively). These results suggest that histamine H₃ receptors, located outside the myenteric plexus, mediate an inhibition of the gastrointestinal transit in vivo. Conversely, the presence of ₂-adrenoceptors in the cholinergic nerve endings and their inhibitory role in the control of gastrointestinal propulsion is confirmed.     

Exogenous hyperglucagonaemia in insulin controlled diabetic rats increases urea excretion and nitrogen loss from organs

Summary In order to study the effect of hyperglucagonaemia on nitrogen metabolism in diabetes, zinc protamin glucagon 60 g was injected subcutaneously 3 times daily for 4 weeks into streptozotocindiabetic rats (n=5), adequately treated with long acting insulin. This raised the plasma concentration of glucagon to 725±125 (mean±SEM), which is not different from that found in portal blood of uncontrolled diabetic rats: 400±75 ng/l. The controls were 5 diabetic rats treated with insulin alone and 5 non-diabetic rats.Compared with control rats the nitrogen balance was reduced (p<0.05) and the nitrogen contents of carcass, heart, intestines, and kidneys were reduced by 15–30% (p<0.05) in the glucagon treated rats. The hepatic capacity of urea synthesis and the alanine elimination rate were determined in the 3 above-mentioned groups, and confirmed in 3 identical groups followed for only 2 weeks; and in addition in a group of glucagon treated diabetic rats, where the long acting glucagon was substituted by neutral insulin the last two days before investigation. The capacity of urea-N synthesis and the alanine elimination rate were, respectively, in control rats: 9.6±0.8 and 5.9±0.3 mol/(min 100 g body weight), in insulin treated diabetic rats: 8.5±0.7 and 5.4±0.6 mol/(min 100g body weight), in glucagon treated rats: 6.3±0.4 (lower than controls, p<0.05) and 10.4±0.4 (higher than controls, p<0.05) (mol/(min 100 g body weight), and in glucagon treated rats given neutral insulin: 20.7±1.6 and 10.9±0.3 mol/(min 100 g body weight) (both higher than controls, p<0.05). Hyperglucagonaemia in itself leads to loss of nitrogen from organs, probably by an increased hepatic conversion of amino-nitrogen to urea-nitrogen, as evidenced by the increased urea excretion. This proceeds despite an insulin induced decrease in the capacity of urea synthesis and may thus rather be attributed to changes in the affinity of urea synthesis for amino-nitrogen.     

Noninsulin-like action of sodium orthovanadate in the isolated perfused liver of fed,non-diabetic rats

Summary Vanadium compounds exert insulin-like effects on isolated rat adipocytes and skeletal muscle and improve glucose homeostasis in diabetic rats and mice. However, reports on metabolic actions of vanadium in the liver are still contradictory. Thus, the acute effect of sodium orthovanadate infusion on net glucose production was measured in isolated perfused livers of non-fasting, non-diabetic rats. Continuous infusion (0.2 ml/min; 90 min) of vanadate (10–500 mol/l) rapidly increased hepatic glucose (p<0.001), but not cyclic AMP output, reaching peak values after 20 min. The cumulative glucose release displayed concentration dependence with a maximal net effect of 394.3 mol/100 g body weight and an apparent half-maximal effective vanadate concentration of 19.6 mol/l. The glycogenolytic response to vanadate was almost completely blocked by 100 mU/l insulin (p<0.005), by 0.1 mmol/l indomethacin (p<0.05) and in the absence of Ca²⁺ (p<0.001). These results indicate that sodium orthovanadate stimulates glycogenolysis in livers of fed, non-diabetic rats by a Ca²⁺-dependent mechanism, which may involve the release of prostaglandins.     

Granulocyte Elastase in Cirrhotic Patients with Spontaneous Bacterial Peritonitis

Granulocyte elastase (GE) is a powerfulproteolytic enzyme that is released by PMNs whendegranulated in infectious processes. The aim of thisstudy was to measure GE in ascites and plasma ofcirrhotic patients with spontaneous bacterial peritonitis(SBP). We studied 29 cirrhotic patients, 17 of themhaving SBP (group A). Twelve patients with noninfectedascites formed the control group (group B). At the time of diagnosis of SBP, GE levels inascites (183.17 ± 86.11 g/liter) and plasma(114.6 ± 35.99 g/liter) were higher in groupA than in group B (27.41 ± 11.54 g/liter, P< 0.00001 and 82.54 ± 20.52 g/liter, P = 0.01,respectively). Levels of GE in ascites had a high valuefor discriminating between patients with and withoutSBP. In the patients who responded to the initialantibiotic treatment, these values significantly decreasedin ascites (67.69 ± 54.22 g/liter, P = 0.003)and plasma (67 ± 22.39 g/liter, P = 0.01) 48hr after therapy was started, in parallel with thedecrease of PMN in ascites. In patients who did notrespond, the production of GE remained elevated.Patients who developed renal insufficiency following SBPhad more marked elevation of GE in plasma (144.8± 33.43 g/liter) than those with normal renalfunction (99.5 ± 27.53 g/liter, P = 0.02).These results suggest that the measurement of GE may behelpful for the diagnosis of SBP in patients withcirrhosis and for assessing the efficacy of therapy. Inaddition, the release of GE into plasma may contributeto the impairment of renal function that follows SBP insome patients.