Hassan Jijakli

Stimulus-secretion coupling of arginine-induced insulin release: comparison between the cationic amino acid and its methyl ester.

The role currently ascribed to the accumulation of l-arginine in the pancreatic islet B-cell as a determinant of its insulinotropic action was reevaluated by comparing the uptake and the metabolic, ionic, electric, and secretory effects of the cationic amino acid with those of its more positively charged methyl ester in rat pancreatic islets. The response to l-arginine methyl ester differed from that evoked by the unesterified amino acid by a lower uptake and oxidation, lack of inhibitory action on d-glucose metabolism, more severe inhibition of the catabolism of endogenous l-glutamine, inhibition of 45Ca net uptake, decrease in both 86Rb outflow from prelabeled islets perifused at normal extracellular Ca2+ concentration and 45Ca efflux from prelabeled islets perifused in the absence of extracellular Ca2+, and delayed and lesser insulinotropic action. These findings reinforce the view that the carrier-meadiated entry of l-arginine into the islet B-cells, with resulting depolarization of the plasma membrane, represents the essential mechanism for stimulation of insulin release by this cationic amino acid.The role currently ascribed to the accumulation of L-arginine in the pancreatic islet B-cell as a determinant of its insulinotropic action was reevaluated by comparing the uptake and the metabolic, ionic, electric, and secretory effects of the cationic amino acid with those of its more positively charged methyl ester in rat pancreatic islets. The response to L-arginine methyl ester differed from that evoked by the unesterified amino acid by a lower uptake and oxidation, lack of inhibitory action on D-glucose metabolism, more severe inhibition of the catabolism of endogenous L-glutamine, inhibition of 45Ca net uptake, decrease in both 86Rb outflow from prelabeled islets perifused at normal extracellular Ca2+ concentration and 45Ca efflux from prelabeled islets perifused in the absence of extracellular Ca2+, and delayed and lesser insulinotropic action. These findings reinforce the view that the carrier-mediated entry of L-arginine into the islet B-cells, with resulting depolarization of the plasma membrane, represents the essential mechanism for stimulation of insulin release by this cationic amino acid.

Effects of artificial sweeteners on insulin release and cationic fluxes in rat pancreatic islets

Beta-L-glucose pentaacetate, but not alpha-D-galactose pentaacetate, was recently reported to taste bitter and to stimulate insulin release. This finding led, in the present study, to the investigation of the effects of both bitter and non-bitter artificial sweeteners on insulin release and cationic fluxes in isolated rat pancreatic islets. Sodium saccharin (1.0-10.0 mM), sodium cyclamate (5.0-10.0 mM), stevioside (1.0 mM) and acesulfame-K (1.0-15.0 mM), all of which display a bitter taste, augmented insulin release from islets incubated in the presence of 7.0 mM D-glucose. In contrast, aspartame (1.0-10.0 mM), which is devoid of bitter taste, failed to affect insulin secretion. A positive secretory response to acesulfame-K was still observed when the extracellular K+ concentration was adjusted to the same value as that in control media. No major changes in 86Rb and 45Ca outflow from pre-labelled perifused islets could be attributed to the saccharin, cyclamic or acesulfame anions. It is proposed that the insulinotropic action of some artificial sweeteners and, possibly, that of selected hexose pentaacetate esters may require G-protein-coupled receptors similar to those operative in the recognition of bitter compounds by taste buds.

Gut permeability and intestinal mucins, invertase, and peroxidase in control and diabetes-prone BB rats fed either a protective or a diabetogenic diet.

This study deals with the enteropathy recently identified in diabetes-prone BB rats (BBdp). Diabetes-resistant BB rats (BBc) and BBdp rats were fed from days 32–39 onward either a protective diabetes-retardant hydrolyzed casein diet (HC) or a plant-based diabetogenic (NTP) diet. The NTP diet decreased body weight and plasma insulin in BBc and BBdp rats. The BBdp rats displayed low intestinal invertase and increased intestinal peroxidase activity. In the BBdp rats fed the HC diet, the mucin content 30–35 cm below the pylorus was higher and the gut permeability lower than in the other three rat groups. There was a significant inverse correlation between gut permeability and the insulinogenic index in the BBdp rats fed the HC or NTP diet. Thus, in BBdp rats, the HC diet somehow prevents the increase in gut permeability and the decrease in the insulinogenic index otherwise found in some of these diabetes-prone animals.

Stimulation by D-glucose of 36Cl- efflux from prelabeled rat pancreatic islets.

Abstractd-glucose was previously reported to cause a concentration-related decrease in the 36Cl− content of prelabeled islets prepared from ob/ob mice, a current animal model of inherited obesity. From these findings, it was inferred that the hexose stimulates Cl− efflux from islet cells and that such an increase in Cl− permeability may partly mediate glucose-induced depolarization of insulin-producing cells. The aim of the present study was to investigate the possible extension of these findings to islets prepared from normal rats by measuring the changes evoked by increasing concentrations of d-glucose in 36Cl− outflow itself from prelabeled isolated islets. After 60 min preincubation at 37°C in the presence of 3 mMd-glucose and 36Cl(75 µCi/mL), the islets were incubated for 8–10 min at 37°C in the presence of increasing concentrations of the hexose (3–20 mM). The changes in 36Cl− outflow during incubation indicated that d-glucose, in excess of a threshold concentration close to 5 mM, indeed increases effluent radioactivity from the prelabeled islets. It is proposed, therefore, that a gating of volume-sensitive anion channels in glucosestimulated insulin-producing islet cells participates in the depolarization of the plasma membrane recorded in the range of insulinotropic concentrations of the hexose.

Dynamics of the cationic and secretory responses to A-4166 in perifused pancreatic islets

Summary— The dynamics of the cationic and secretory response to A‐4166, a hypoglycemic meglitinide analogue, was investigated in rat islets prelabelled with either 86Rb or 45Ca and placed in a perifusion system. In the absence of D‐glucose, A‐4166 (10 μM) provoked an immediate, sustained and rapidly reversible inhibition of 86Rb outflow, this contrasting with a short‐lived stimulation of insulin release. In the presence of 6 mM D‐glucose, A‐4166 provoked a rapid, sustained and rapidly reversible stimulation of both insulin release and 45Ca efflux. The latter cationic response was suppressed in the absence of extracellular Ca2+, in which case A‐4166 even caused a modest decrease in effluent radioactivity.

CATIONIC EVENTS STIMULATED BY D-GLUCOSE IN DEPOLARIZED ISLET CELLS

Pancreatic islets prelabelled with either 86Rb or 45Ca were exposed to a rise in D-glucose concentration from 2.8 to 16.7 mM whilst perifused in the presence of 2 microM glibenclamide, 30 mM extracellular K+ and both 30 mM K+ and 250 microM diazoxide. In all three situations, the rise in glucose concentration provoked a dramatic increase in insulin output, despite unchanged or even increased efflux of 86Rb from the prelabelled islets. Also in all three situations, glucose sharply decreased effluent radioactivity from islets prelabelled with 45Ca but perifused in the absence of extracellular Ca2+, while augmenting 45Ca efflux, to a variable extent, from islets perifused at normal extracellular Ca2+ concentration (1.0 mM). It is proposed, therefore, that the insulinotropic action of D-glucose in depolarized islets, and presumably also under normal conditions, may involve the gating of voltage-insensitive Ca2+ channels.

Insulinotropic action of α-d-glucose pentaacetate: functional aspects

The functional determinants of the insulinotropic action of α-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, inhibiting86Rb outflow, and augmenting45Ca 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, α-d-galactose pentaacetate, and β-d-galactose pentaacetate all failed to stimulate insulin release. The secretory response to α-d-glucose pentaacetate was reproduced by β-d-glucose pentaacetate and, to a lesser extent, by β-l-glucose pentaacetate. It differed from that evoked by unesterifiedd-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 α-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 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 β-l-glucose pentaacetate

The metabolism of β-l-glucose pentaacetate and its interference with the catabolism ofl-[U-14C]glutamine, [U-14C]palmitate,d-[U-14C]glucose, andd-[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 β-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.

Tritiated taurine handling by isolated rat pancreatic islets

A gating of volume-sensitive anion channels may participate in the depolarization of the plasma membrane caused by high concentration of d-glucose in insulin-producing B-cells of the endocrine pancreas. The efflux of tritiated taurine from prelabeled cells is currently used to assess changes in the activity of such channels. The handling of [1,2-3H]taurine by isolated rat pancreatic islets was therefore investigated. The net uptake of [1,2-3H]taurine was found to represent a concentration-, time-, and temperature-dependent process. It was progressively increased in the range of d-glucose concentrations between 2.8 and 8.3 mM, but no further increase was observed at 16.7 mMd-glucose. Over 15 min incubation, the efflux of radioactivity from prelabeled islets was inhibited by MK571 (1.0 mM). It was increased in response to hypoosmolarity both in the presence and absence of extracellular Na+. Whether in salt-balanced or Na+-deprived media, the efflux of radioactivity from prelabeled islets increased in response to a rise in d-glucose concentration from 2.8 to 5.6 or 8.3 mM, but decreased when the concentration of the hexose was further increased from 8.3 to 16.7 mM. In perifused islets, however, the radioactive efflux from prelabeled islets was inhibited, in a concentration-related manner, when islets first deprived of d-glucose for 45 min were then exposed to 2.8, 5.6, or 16.7 mMd-glucose. Likewise, in prelabeled and perifused islets first exposed for 45 min to 4.0 mMd-glucose, a later rise in hexose concentration to 8.3 mM failed to affect significantly effluent radioactivity, while an increase in hexose concentration from 4.0 to 16.7 mM inhibited the radioactive outflow. In these perifusion experiments, the rise in d-glucose concentration provoked the expected changes in insulin output. The findings obtained in islets examined immediately after preincubation in the presence of [1,2-3H]taurine are consistent with the presence of volume-sensitive anion channels in islet cells and with a gating of such channels in response to a rise in d-glucose concentration from 2.8 to 5.6–8.3 mM. However, the radioactive fractional outflow rate from prelabeled islets seems to reach its highest value at about 8.3 mMd-glucose, being unexpectedly decreased at a higher concentration (16.7 mM) of the hexose. In conclusion, the pleiotropic effects of d-glucose upon tritiated taurine outflow from prelabeled rat islets, which could conceivably be ascribed to differences in the handling of this amino sulfonic acid by distinct islet cell types, indicates that the present approach is far from optimal to characterize unambiguously the regulation by the hexose of volume-sensitive anion channel activity in insulin-producing islet cells.