Preserved GLP-I effects on glycogen synthase a activity and glucose metabolism in isolated hepatocytes and skeletal muscle from diabetic rats.

To search if biological effects of GLP-I on glucose metabolism in extrapancreatic tissue are present in diabetic states, we have studied the action of GLP-I and insulin on glycogen-enzyme activity, glycogen synthesis, and glucose metabolism in isolated hepatocytes and soleus muscle from adult streptozotocin (STZ)- and neonatal STZ-treated diabetic rats. This work confirms the previously reported insulin-like effects of GLP-I on glucose metabolism in both muscle and liver tissue from normal rats (control). The present study extends those observations to the muscle and liver tissue of diabetic animals. In both muscle and liver tissue, the metabolism of D-glucose, in the absence of added peptides, was more severely affected in adult STZ (IDDM model) than in neonatal STZ (nSTZ; NIDDM model) rats, and the magnitude of hormonal effect on metabolic variables was lower in diabetic rats than in control rats, as a rule. Nevertheless, in liver and muscle tissue of diabetic rats, GLP-I was able to increase glycogen synthase activity, augment the net rate of D-[U-14C]glucose incorporation into glycogen, and increase D-[5-3H]glucose utilization, D-[U-14C]glucose oxidation, and lactate production. In conclusion, GLP-I exerts insulin-like effects on D-glucose metabolism in both muscle and liver tissue in IDDM or NIDDM animal models, and present observations reinforce the view that GLP-I may represent a most promising tool in the treatment of diabetic patients.

Failure of D-fructose to stimulate protein biosynthesis in pancreatic islets.

Rat pancreatic islets were incubated for 90 min with L-[4-3H]phenylalanine in either the absence or presence of D-glucose and D-fructose. Relative to basal value, D-glucose (16.7 mM) increased the incorporation of the tritiated amino acid into trichloroacetic acid-precipitable material. However, when D-fructose was tested in the 80 to 240 mM concentration range, it failed to stimulate islet biosynthetic activity. Since D-fructose causes, in the same concentration range, a dose-related stimulation of insulin release, the dissociation between the biosynthetic and secretory responses to D-fructose supports the view that the insulinotropic action of the ketohexose does not entail the same metabolic determinants as those operative in glucose-stimulated islets.

GLP-1 (7-36) amide: effects on glucose transport and metabolism in rat adipose tissue.

In rat adipocytes, GLP-1 (7-36) amide induced an increment in 2-deoxy-glucose uptake, which was additive to that of insulin. Furthermore, in rat fat, GLP-1 (7-36) amide provoked a rise in glycogen synthesis, glucose oxidation and utilization and lipogenesis, the increments being lower than those obtained with insulin. These data support the idea that GLP-1 exerts insulin-like effects on glucose metabolism in rat adipose tissue, as it does in rat hepatocytes and skeletal muscle, although with a lower potency than that of insulin.

Repaglinide preserves nutrient-stimulated biosynthetic activity in rat pancreatic islets.

The meglitinide analogue repaglinide is a novel non-sulphonylurea insulinotropic agent which, like hypoglycaemic sulphonylureas, causes the closing of ATP-sensitive K+ channels in islet cells. We have now explored the effect of repaglinide upon proinsulin biosynthesis in rat pancreatic islets. Groups of eight islets each were incubated for 90 min in the presence of L-[4-(3)H]phenylalanine (4 microM) and glucose (2.8 or 16.7 mM), in the absence or presence of repaglinide (10 microM). A rise in glucose concentration caused a four-fold increase of the incorporation of L-[4-(3)H]phenylalanine into TCA-precipitable material. Repaglinide failed to adversely affect protein biosynthesis, whether at low or high glucose concentrations. Further characterization of the biosynthetic response was achieved by separation of the tritiated peptides by gel filtration. In the absence of repaglinide, the (pro)insulin/total ratio of tritiated peptides averaged 33.3 +/- 10.2 and 58.7 +/- 1.7% (n = 6 in both cases) at 2.8 and 16.7 mM D-glucose, respectively. Repaglinide again failed to significantly affect such ratios. In conclusion, repaglinide may offer the advantage over hypoglycaemic sulphonylureas of preserving nutrient-stimulated biosynthetic activity in pancreatic islet cells.

Insulinotropic action of 1,2,3-Tri(methylsuccinyl)glycerol ester.

A novel ester of succinic acid, 1,2,3-tri(methylsuccinyl)glycerol ester (3SMG), was found to stimulate insulin release from rat pancreatic islets. In the presence of 7 mM d-glucose, a 10 microM concentration of 3SMG was sufficient to cause a significant increase in insulin output. The ester mimicked the effect of other nutrient secretagogues in enhancing the synthesis of islet peptides, with a preferential action on proinsulin as distinct from nonhormonal peptides, in decreasing 86Rb outflow from prelabeled islets, and in stimulating Ca2+ inflow into the islet cells. It is proposed, therefore, that 3SMG displays the attributes suitable for stimulation or potentiation of insulin release in noninsulin-dependent diabetes, without requiring administration in large amounts and, hence, without the risk of excessive hepatic gluconeogenesis.

Insulinotropic action of methyl pyruvate: secretory, cationic, and biosynthetic aspects.

Methyl pyruvate was found to exert a dual effect on insulin release from isolated rat pancreatic islets. A positive insulinotropic action prevailed at low concentrations of D-glucose, in the 2.8 to 8.3 mM range, and at concentrations of the ester not exceeding 10.0 mM. It displayed features typical of a process of nutrient-stimulated insulin release, such as decreased K+ conductance, enhanced Ca2+ influx, and stimulation of proinsulin biosynthesis. A negative insulinotropic action of methyl pyruvate was also observed, however, at a high concentration of D-glucose (16.7 mM) and/or at a high concentration of the methyl ester (20.0 mM). It was apparently not attributable to any adverse effect of methyl pyruvate on ATP generation, but might be due to hyperpolarization of the plasma membrane. The ionic determinant(s) of the latter change was not identified. The dual effect of methyl pyruvate probably accounts for an unusual time course of the secretory response, including a dramatic and paradoxical stimulation of insulin release upon removal of the ester.

The riddle of formycin A insulinotropic action.

Formycin A augments insulin release evoked by glucose (5.6 mm or more), this effect not being rapidly reversible. The mechanism responsible for the insulinotropic action of formycin A was investigated in isolated pancreatic islets. It could not be ascribed to facilitation of glucose metabolism. On the contrary, formycin A inhibited glucose oxidation, lowered ATP content, and impaired glucose-stimulated protein biosynthesis. The insulinotropic action of formycin A was apparently attributable to its conversion to formycin A 5'-triphosphate, both this process and the secretory response to formycin A being abolished by the inhibitor of adenosine kinase 5-iodotubercidin. In agreement with the latter view, adenosine receptor antagonists such as 8-cyclopentyl-1, 3-dipropylxanthine and 3,7-dimethyl-1-propargylxanthine failed to suppress and, instead, augmented the insulinotropic action of formycin A. Unexpectedly, however, formycin A failed to decrease 86Rb efflux, this coinciding with a low efficiency of formycin A 5'-triphosphate to inhibit KATP-channel activity in excised membranes and with the fact that formycin A increased gliben-clamide-stimulated insulin release. The secretory response to formycin A represented a Ca2+-dependent process suppressed in the absence of extracellular Ca2+ or presence of verapamil and associated with an increased net uptake of 45Ca. Nevertheless, the view that formycin A exerts any major effect upon intracellular Ca2+ redistribution, protein kinase C activity, or cyclic AMP net production also met with objections such as the minor secretory effect of formycin A in islets exposed to a high concentration of K+ in the presence of a diazoxide analog, the resistance of formycin A insulinotropic action to bisindolylmaleimide, the poor increase of cyclic AMP content in formycin A-stimulated islets, and the pronounced enhancement by forskolin or theophylline of insulin release from islets exposed to formycin A. It is concluded, therefore, that the mechanism of action of formycin A in the pancreatic beta-cell remains to be elucidated.

Insulinotropic action of the monoethyl ester of succinic acid.

1. Selected esters of succinic acid are currently under investigation as potential insulinotropic tools in the treatment of non-insulin-dependent diabetes. At variance with the methyl esters of succinic acid used in most of the work so far conducted from this perspective, the monoethyl ester of succinic acid (EMS) offers the advantage of avoiding the undesirable generation of methanol by intracellular hydrolysis. In the present study, the metabolism and functional effects of EMS were investigated, therefore, in rat pancreatic islets. 2. At a 10 mM concentration, EMS enhanced insulin release from islets stimulated by 7-17 mM D-glucose but failed to do so at lower concentrations of the hexose. EMS was efficiently metabolized, as judged from the generation of 14CO2 by islets exposed to the monoethyl ester of either [1,4-14C] or [2, 3-14C]succinic acid. D-Glucose (6 mM) failed to affect the metabolism of EMS (10 mM), which itself failed to affect the metabolism of D-[5-3H]glucose or D-[U-14C]glucose. EMS also stimulated biosynthetic activity in the islets. It inhibited 86Rb and 45Ca outflow from prelabeled islets perfused in the absence of D-glucose but enhanced the efflux of the two cationic tracers in the presence of the hexose (7 mM). 3. It is concluded that the insulinotropic action of EMS is attributable, to a large extent, to its capacity to act as a nutrient in islet cells.

Insulinotropic action of alpha-D-glucose pentaacetate: functional aspects.

The functional determinants of the insulinotropic action of alpha-D-glucose pentaacetate were investigated in rat pancreatic islets. The ester mimicked the effect of nutrient secretagogues by recruiting individual B cells into an active secretory state, stimulating proinsulin biosynthesis, inhibiting 86Rb outflow, and augmenting 45Ca efflux from prelabeled islets. The secretory response to the ester was suppressed in the absence of Ca2+ and potentiated by theophylline or cytochalasin B. The generation of acetate from the ester apparently played a small role in its insulinotropic action. Thus acetate, methyl acetate, ethyl acetate, alpha-D-galactose pentaacetate, and beta-D-galactose pentaacetate all failed to stimulate insulin release. The secretory response to alpha-D-glucose pentaacetate was reproduced by beta-D-glucose pentaacetate and, to a lesser extent, by beta-L-glucose pentaacetate. It differed from that evoked by unesterified D-glucose by its resistance to 3-O-methyl-D-glucose, D-mannoheptulose, and 2-deoxy-D-glucose. It is concluded that the insulinotropic action of alpha-D-glucose pentaacetate, although linked to the generation of the hexose from its ester, entails a coupling mechanism that is not identical to that currently implied in the process of glucose-induced insulin release.

Insulinotropic action of beta-L-glucose pentaacetate.

The metabolism of beta-L-glucose pentaacetate and its interference with the catabolism of L-[U-14C]glutamine, [U-14C]palmitate, D-[U-14C]glucose, and D-[5-3H]glucose were examined in rat pancreatic islets. Likewise, attention was paid to the effects of this ester on the biosynthesis of islet peptides, the release of insulin from incubated or perifused islets, the functional behavior of individual B cells examined in a reverse hemolytic plaque assay of insulin secretion, adenylate cyclase activity in a membrane-enriched islet subcellular fraction, cAMP production by intact islets, tritiated inositol phosphate production by islets preincubated with myo-[2-3H]inositol, islet cell intracellular pH, 86Rb and 45Ca efflux from prelabeled perifused islets, and electrical activity in single isolated B cells. The results of these experiments were interpreted to indicate that the insulinotropic action of beta-L-glucose pentaacetate is not attributable to any nutritional value of the ester but, instead, appears to result from a direct effect of the ester itself on a yet unidentified receptor system, resulting in a decrease in K+ conductance, plasma membrane depolarization, and induction of electrical activity.