Chizuko Ohboshi

Effect of glucagon-like peptide-1 on insulin secretion

The insulinotropic actions of two forms of glucagon-like peptide 1 (GLP-1) containing 31 and 37 amino acid residues on perfused rat pancreas were compared with that of gastric inhibitory polypeptide (GIP), hitherto the most potent intestinal insulinotropic polypeptide known. The smaller form, C-terminally amidated GLP-1-(7-36), strongly enhanced insulin secretion stimulated by 11.1 mM D-glucose at a concentration as low as 0.1 nM. Comparable effects of GIP and GLP-1-(1-37) on insulin secretion were observed at concentrations of 1.0 nM and 10.0 nM, respectively. At the doses tested, neither GLP-1s nor GIP had any effect on insulin secretion induced by 3.3 mM D-glucose. At a concentration of 1.0 nM, GLP-1-(7-36 amide) also enhanced insulin secretion induced by 5 mM L-arginine whereas at concentrations of up to 10.0 nM, GLP-1-(1-37) did not. The results show that the smaller form of GLP-1 is more strongly insulinotropic than GIP. These findings suggest that the smaller GLP-1 may have a physiologically more important role as a modulator of insulin release.

Comparison of half-disappearance times, distribution volumes and metabolic clearance rates of exogenous glucagon-like peptide 1 and glucagon in rats

The pharmacokinetics of glucagon-like peptide-1 (GLP-1) in vivo after bolus and continuous i.v. administrations of the peptide were compared with those of glucagon in rats. The half-disappearance time (t1/2) distribution volume (Vd) and metabolic clearance rate (MCR) of GLP-1 given as a bolus injection and by constant infusion, were, respectively, as follows: t1/2 (min), 47.7 +/- 14.5 and 39.5 +/- 15.5 (mean +/- S.D.); Vd (ml), 903.8 +/- 62.4 and 516.3 +/- 92.1 and MCR (ml kg-1 min-1), 27.4 +/- 10.8 and 18.6 +/- 8.6. These values differed significantly from the respective values for glucagon (t1/2, 3.3 +/- 0.6 and 5.8 +/- 1.0; Vd, 206.5 +/- 25.9 and 240.0 +/- 76.1; and MCR, 83.1 +/- 8.2 and 46.7 +/- 13.3). These findings demonstrate that GLP-1 is degraded more slowly than glucagon in vivo.

Suppression of synthesis and release of glucagon by glucagon-like peptide-1 (7–36 amide) without affect on mRNA level in isolated rat islets

Glucagon-like peptide-1 (GLP-1) has been reported to inhibit glucagon release. To investigate the mechanism involved, we examined the effects of GLP-1 on the preproglucagon mRNA level and the content and release of glucagon in the isolated rat islets. Arginine significantly increased the content and release of glucagon after incubation for 1 h or 18 h. The preproglucagon mRNA level was not increased after incubation for 1 h but was increased after incubation for 18 h. GLP-1 (7-36 amide) significantly decreased the content and release of glucagon after incubation for 1 h or 18 h, but did not affect arginine-induced increase in the preproglucagon mRNA level after incubation for 18 h. These data suggest that GLP-1 suppresses post-transcriptional processes in the content and release of glucagon.

Identification of Glucagon-Like Peptide-1(7–36) Amide in Rat Brain

Acid-urea extract of rat brain was examined by glucagon-like peptide-1 (GLP-1) specific radioimmunoassay. A single peak was observed which co-eluted with GLP-1(7–36)amide on gel filtration and anion exchange chromatography. In contrast, GLP-1(1–37) was not detected under our experimental conditions. The fact that GLP-1 (7–36)amide, but not GLP-1(1–37), was present in rat brain suggests that preproglucagon was processed in the brain in the same manner as in the intestine and not as in the pancreas.

A large molecular form of glucagon-like peptide-1(GLP-1) immunoreactivity is co-released with glucagon from pancreas by arginine in normal subjects

Plasma immunoreactivities of glucagon-like peptide-1 (GLP-1IR) in normal subjects were measured with a specific radioimmunoassay during the arginine tolerance test. Plasma GLP-1IR after arginine infusion showed a 3-fold increase in parallel to plasma glucagon immunoreactivity and plasma glucagon-like immunoreactivity, measured with a glucagon C-terminal specific antiserum (OAL 123) and an N-terminal and/or central region specific glucagon antiserum (OAL 196), respectively. This finding suggested that the increased immunoreactivities of GLP-1 as well as that of glucagon were of pancreatic origin. Upon gel chromatography, plasma at the basal state showed three GLP-1 immunoreactive peaks, eluted in the position of void volume, synthetic GLP-1(72-108), and a smaller molecular fraction. Gel chromatography of plasma after an arginine load showed an additional peak (Mr 13,000-15,000) with little change in other GLP-1 immunoreactive peaks. This large molecular form of GLP-1IR was also shown to exist in the human pancreatic extract. Moreover, the free GLP-1 concentrations in plasma before and after an arginine load were shown to be about equal by reverse phase HPLC. These data suggested that in normal subjects arginine stimulation co-releases GLP-1IR, predominantly large molecular form, with glucogen from the pancreas.

Alterations of plasma immunoreactive glucagon-like peptide-1 behavior in non-insulin-dependent diabetics

The basal level of plasma immunoreactive glucagon-like peptide-1 (IR GLP-1) was significantly elevated in non-insulin-dependent diabetics (NIDD), and this elevation of IR GLP-1 was mainly due to an increase in the large component of IR GLP-1, corresponding to the pancreatic form. During the oral glucose-tolerance test (OGTT), the total plasma IR GLP-1 decreased in normal subjects but increased significantly in diabetic patients. Chromatographic analysis showed that IR GLP-1 consisted of several different molecular forms. OGTT caused a decrease in the pancreatic form but increased the intestinal form in normal subject, resulting into a net decrease in total plasma IR GLP-1. Whereas in NIDD the increase in the intestinal form was more prominent and the suppression of the pancreatic form was practically abolished to result in a net increase of total plasma IR GLP-1. This observation is consistent with the fact that in normal subjects the total change in IR GLP-1 was significantly correlated with both the total change of gut glucagon as well as that of pancreatic glucagon, but in diabetics the total change of GLP-1 only correlated to that of gut glucagon. The impaired suppression of pancreatic GLP-1 and enhanced release of intestinal GLP-1 could have some physiological importance in NIDD.

Specific radioimmunoassay of glucitol-lysine — application to lens proteins in streptozotocin-diabetic rats

SummaryA radioimmunoassay using antibody against glucitol-lysine was developed to quantitate glycated proteins in the lens of diabetic rats. The amount of glycated protein was expressed as molar equivalents of reduced glycated hippuryl lysine (GlcRED-Hip-Lysine). Significant differences (p<0.01) were found in the amounts of glycated protein in the lenses of rats with streptozotocin-induced diabetes (3.92±0.59 nmol/mg protein, n=5), those with streptozotocin-induced diabetes treated with insulin (2.94±0.36 nmol/mg protein, n=4) and normal rats (1.23±0.22 nmol/mg protein, n=5). There was a significant correlation between the concentration of glycated protein in the lens and the HbA1c level at the end of the 12 week experiment (r=0.957, p<0.001). These results indicate that glycation of lens protein is parallel with the severity of diabetes in rats.

Effect of the enteroinsular axis on both the A- and B-cell response to arginine after oral glucose in man

SummaryStudies were made on the effect of the enteroinsular axis on amino acid-induced insulin and glucagon secretion during hyperglycaemia in man. The responses of plasma immunoreactive insulin, C-peptide, and immunoreactive glucagon to arginine infusion were investigated in nine healthy subjects after induction of hyperglycaemia by an oral glucose load and by intravenous glucose infusion to produce similar glucose concentrations in the arterialised blood. The plasma immunoreactive insulin and C-peptide levels increased to higher levels after an oral glucose load than after an intravenous infusion of glucose. The incremental areas under the immunoreactive insulin and C-peptide curves during arginine infusion were significantly greater (p<0.01) after oral than after intravenous glucose administration. The plasma immunoreactive glucagon level was suppressed equally after oral and intravenous glucose loads. However, during subsequent arginine infusion, the plasma immunoreactive glucagon level rose more in the presence of hyperglycaemia induced by oral than intravenous glucose. The incremental area under the plasma immunoreactive glucagon curve during arginine infusion was 1.6-fold greater after glucose ingestion than after intravenous glucose infusion. These results suggest that the enteroinsular axis has a stimulatory effect on the responses of pancreatic A and B cells to arginine after oral glucose administration.