Effect of exogenous ATP upon inositol phosphate production, cationic fluxes and insulin release in pancreatic islet cells

Endogenous ATP is thought to play a key regulatory role in nutrient-stimulated insulin release. The present study deals with the effect of exogenous ATP and its stable analog alpha, beta-methylene ATP upon pancreatic islet function. Both alpha, beta-methylene ATP (5.0 microM to 0.2 mM) and ATP (0.3-3.0 mM) caused a rapid and concentration-related increase in insulin output by rat islets incubated or perfused at an intermediate concentration of D-glucose (8.3 mM). The effect of the ATP analog faded out at both lower and higher D-glucose concentrations. In the presence of 8.3 mM D-glucose, ATP also increased both 86Rb and 45Ca outflow from prelabelled islets. The cationic response to ATP persisted in the absence of extracellular Ca2+ and, hence, was reminiscent of that evoked by cholinergic agents. Like carbamylcholine, ATP caused a dose-related increase in the production of [3H]inositol phosphates from prelabelled islets or tumoral islet cells (RINm5F line). The latter effect was duplicated by alpha, beta-methylene ATP and unaffected by atropine. It is speculated that ATP, liberated together with insulin at the exocytotic site, might participate in a positive feedback control of insulin release.     

Ultrastructural changes of the pancreatic beta-cell and the insulin secretion by islets from lactating and non-lactating rats

Islets isolated from lactating rats, as compared to islets from non-lactating rats, release less insulin when incubated in the absence of exogenous nutrient or presence of either D-glucose (11.1 mM) or the association of L-leucine and L-glutamine (10.0 mM each). The insulin content of the islets is not different in lactating and non-lactating rats. The volume density of the dark granules in the beta-cells is not at variance in both groups. However the volume density of the light (pale) granules is significantly lower in the lactating rats. The reduced amount of light granules is in keeping with the reduced secretory capacity of the beta-cells from lactating rats.     

Inhibition of glucose-induced insulin release by 3-O-methyl-D-glucose: Enzymatic,metabolic and cationic determinants

The analog of D-glucose, 3-O-methyl-D-glucose, is thought to delay the equilibration of D-glucose concentration across the plasma membrane of pancreatic islet B-cells, but not to exert any marked inhibitory action upon the late phase of glucose-stimulated insulin release. In this study, however, 3-O-methyl-D-glucose, when tested in high concentrations (30-80 mM) was found to cause a rapid, sustained and not rapidly reversible inhibition of glucose-induced insulin release in rat pancreatic islets. In relative terms, the inhibitory action of 3-O-methyl-D-glucose was more marked at low than high concentrations of D-glucose. It could not be attributed to hyperosmolarity and appeared specific for the insulinotropic action of D-glucose, as distinct from non-glucidic nutrient secretagogues. Although 3-O-methyl-D-glucose and D-glucose failed to exert any reciprocal effect upon the steady-state value for the net uptake of these monosaccharides by the islets, the glucose analog inhibited D-[5-3H]glucose utilization and D-[U-14C]glucose oxidation. This coincided with increased 86Rb outflow and decreased 45Ca outflow from prelabelled islets, as well as decreased 45Ca net uptake. A preferential effect of 3-O-methyl-D-glucose upon the first phase of glucose-stimulated insulin release was judged compatible with an altered initial rate of D-glucose entry into islet B-cells. The long-term inhibitory action of the glucose analog upon the metabolic and secretory response to D-glucose, however, may be due, in part at least, to an impaired rate of D-glucose phosphorylation. The phosphorylation of the hexose by beef heart hexokinase and human B-cell glucokinase, as well as by parotid and islet homogenates, was indeed inhibited by 3-O-methyl-D-glucose. The relationship between insulin release and D-glucose utilization or oxidation in the presence of 3-O-methyl-D-glucose was not different from that otherwise observed at increasing concentrations of either D-glucose or D-mannoheptulose. It is concluded, therefore, that 3-O-methyl-D-glucose adversely affects the metabolism and insulinotropic action of D-glucose by a mechanism largely unrelated to changes in the intracellular concentration of the latter hexose.     

Fasting-induced dissociation of cationic and secretory events in pancreatic islets

In pancreatic islets removed from 48 h-fasted rats, as distinct from fed animals, the release of insulin evoked by D-glucose is more severely impaired than that evoked by 2-ketoisocaproate. This decreased secretory response to D-glucose contrasts with an unimpaired cationic response to the sugar in terms of the glucose-induced decrease in both 86Rb and 45Ca outflow from pre-labelled islets. Likewise, fasting only causes a modest decrease of the secondary rise in 45Ca outflow evoked by D-glucose in islets perifused at normal Ca2+ concentration. The latter decrease appears more marked, however, if the cationic response to glucose is expressed relative to that evoked by 2-ketoisocaproate in islets removed from rats in the same nutritional state. It is concluded that, in the process of nutrient-stimulated insulin release, neither the decrease in K+ conductance (inhibition of 86Rb outflow) nor the sequestration of Ca2+ by intracellular organelles and/or direct inhibition of Ca2+ outward transport (decrease in 45Ca outflow) represent the sole determinant(s) of the subsequent gating of Ca2+ channels (secondary rise in 45Ca efflux).     

Glucose-induced mobilisation of intracellular Ca2+ in depolarised pancreatic islets

Perifused rat pancreatic islets, prelabelled with ⁴⁵Ca, were exposed for 90 min to a medium containing 30 mM K⁺, 0.25 mM diazoxide and 0.5 mM EGTA, but deprived of CaCl₂. Either verapamil (0.05 mM) or Cd²⁺ (0.05 mM) were also present in the perifusate. Under these conditions a rise in D-glucose concentrations from either 2.8 to 16.7 mM or zero to 8.3 mM increased both ⁴⁵Ca outflow and insulin release, after an initial and transient decrease in effluent radioactivity. These findings suggest that, in islets depolarised by exposure to a high extracellular concentration of K⁺, D-glucose provokes an intracellular redistribution of Ca²⁺ ions and subsequent stimulation of insulin release. The functional response to D-glucose is apparently not attributable to either the closing of ATP-sensitive K⁺ channels, which were actually activated by diazoxide, or stimulation of Ca²⁺ influx, which was prevented by the absence of extracellular Ca²⁺. The present experimental design thus reveals a novel component of the glucose-induced remodelling of Ca²⁺ fluxes in islet cells. Such an effect might also be operative under physiological conditions, when the hexose leads to depolarisation of the islet B-cells.     

Stimulus-secretion coupling of arginine-induced insulin release. Functional response of islets to L-arginine and L-ornithine

L-Arginine and L-ornithine stimulate insulin release from pancreatic islets exposed to D-glucose. This coincides with an increased outflow of 86Rb and 45Ca from prelabelled islets and an increased net uptake of 45Ca by the islets. In the presence of D-glucose, L-lysine stimulates insulin secretion to the same extent as L-arginine or L-ornithine, but the hormonal release is not further enhanced by combinations of these cationic amino acids. L-Arginine or L-ornithine failed to enhance insulin release evoked by either L-leucine or 2-ketoisocaproate. The inhibitor of ornithine decarboxylase D,L-alpha-difluoromethyl ornithine failed to affect the metabolism and insulinotropic action of D-glucose in pancreatic islets, and only caused a partial inhibition of the secretory response to either L-arginine or L-ornithine. The latter amino acids inhibited modestly but significantly D-glucose utilization and oxidation by pancreatic islets. These and complementary findings suggest that the secretory response to L-arginine and L-ornithine is not attributable to any major change in the overall oxidative catabolism of nutrients, but involves mainly a biophysical component, such as the depolarization of the plasma membrane by these cationic amino acids.     

Stimulus-secretion coupling of arginine-induced insulin release: significance of changes in extracellular and intracellular pH.

The possible relevance of changes in extracellular and/or intracellular pH to the insulinotropic action of L-arginine and L-homoarginine was investigated in rat pancreatic islets. A rise in extracellular pH from 7.0 to 7.4 and 7.8 augmented the secretory response to these cationic amino acids whilst failing to affect the uptake of L-arginine by islet cells and whilst decreasing the release of insulin evoked by D-glucose. Under these conditions, a qualified dissociation was also observed between secretory data and 45Ca net uptake. Moreover, at high extracellular pH, the homoarginine-induced increase in 86Rb outflow from prelabelled islets rapidly faded out, despite sustained stimulation of insulin release. The cationic amino acids failed to affect the intracellular pH of islet cells, whether in the absence or presence of D-glucose and whether at normal or abnormal extracellular pH. These findings argue against the view that the secretory response to L-arginine would be related to either a change in cytosolic pH or the accumulation of this positively charged amino acid in the beta-cell. Nevertheless, they suggest that the yet unidentified target for L-arginine and its non-metabolized analogue in islet cells displays pH-dependency with optimal responsiveness at alkaline pH.     

32P, 86Rb+ and 45Ca2+ handling by tumoral insulin-secreting cells (RINm5F line)

In perifused tumoral islet cells (RINm5F line), which were prelabelled with either [32P]orthophosphate, 86Rb+ or 45Ca2+, the administration of D-glucose (1.4, 2.8 or 16.7 mM) increased the efflux of 32P, decreased the outflow of 86Rb, increased slightly the efflux of 45Ca from cells perifused in the presence of Ca2+, and decreased modestly the outflow of 45Ca from cells perifused in the absence of Ca2+. D-glucose also stimulated the net uptake of 45Ca2+. When Ba2+ (2 mM) was used, in the absence of Ca2+, instead of D-glucose as an insulin secretagogue, the efflux of 32P was little affected, but the outflow of 45Ca was dramatically increased. These changes are qualitatively similar to those occurring in normal islet cells. Nevertheless, the ionic response to D-glucose appeared, as a rule, less marked in tumoral than normal islet cells. Moreover, the concentration-response relationship was shifted to a lower range of hexose concentrations in the RINm5F cells.     

Studies on the role of inositol trisphosphate in the regulation of insulin secretion from isolated rat islets of Langerhans.

Glucose (20 mM) and carbachol (1 mM) produced a rapid increase in [3H]inositol trisphosphate (InsP3) formation in isolated rat islets of Langerhans prelabelled with myo-[3H]inositol. The magnitude of the increase in InsP3 formation was similar when either agent was used alone and was additive when they were used together. In islets prelabelled with 45Ca2+ and treated with carbachol (1 mM), the rise in InsP3 correlated with a rapid, transient, release of 45Ca2+ from the cells, consistent with mobilization of 45Ca2+ from an intracellular pool. Under these conditions, however, insulin secretion was not increased. In contrast, islets prelabelled with 45Ca2+ and exposed to 20mM-glucose exhibited a delayed and decreased 45Ca2+ efflux, but released 7-8-fold more insulin than did those exposed to carbachol. Depletion of extracellular Ca2+ failed to modify the increase in InsP3 elicited by either glucose or carbachol, whereas it selectively inhibited the efflux of 45Ca2+ induced by glucose in preloaded islets. Under these conditions, however, glucose was still able to induce a small stimulation of the first phase of insulin secretion. These results demonstrate that polyphosphoinositide metabolism, Ca2+ mobilization and insulin release can all be dissociated in islet cells, and suggest that glucose and carbachol regulate these parameters by different mechanisms.     

The stimulus-secretion coupling of glucose-induced insulin release. Cationic and secretory effects of menadione in the endocrine pancreas.

1. Menadione (2-methyl-1,4-naphthoquinone) inhibits insulin release evoked in the rat endocrine pancreas by glucose or glyceraldehyde, but fails to affect the secretory response to Ca2+, Ba2+, theophylline or gliclazide. The inhibitory effect of menadione upon glucose-induced insulin release is a dose-related, rapid and reversible phenomenon, menadione and glucose acting apparently as competitive antagonists. Menadione affects both the early and late phase of the secretory response to glucose. Menadione also antagonizes in a dose-related fashion the ability of glucose to reduce 86Rb efflux, to provoke 86Rb accumulation, to cause biphasic changes in 45 Ca efflux and to stimulate 45 Ca net uptake in pancreatic islets. 2. It is concluded that menadione impairs the insulinotropic action of glucose and other nutrients by impeding the remodelling of cationic fluxes normally provoked by these secretagogues in islet cells. Menadione, however, does not affect the capacity of divalent cations to activate the effector system which controls the release of secretory granules. Menadione may therefore represent a valuable tool to elucidate the mechanism by which glucose normally modifies the movement of cations in the pancreatic B-cell.     

The stimulus-secretion coupling of amino acid-induced insulin release. Insulinotropic action of L-alanine

Available information on the fate and insulinotropic action of L-alanine in isolated pancreatic islets is restricted to data collected in obese hyperglycemic mice. Recent data, however, collected mostly in tumoral islet cells of either the RINm5F line or BRIN-BD11 line, have drawn attention to the possible role of Na(+) co-transport in the insulinotropic action of L-alanine. In the present study conducted in islets prepared from normal adult rats, L-alanine was found (i) to inhibit pyruvate kinase in islet homogenates, (ii) not to affect the oxidation of endogenous fatty acids in islets prelabelled with [U-14C]palmitate, (iii) to stimulate 45Ca uptake in islets deprived of any other exogenous nutrient, and (iv) to augment insulin release evoked by either 2-ketoisocaproate or L-leucine, whilst failing to significantly affect glucose-induced insulin secretion. The oxidation of L-[U-14C]alanine was unaffected by D-glucose, but inhibited by L-leucine. Inversely, L-alanine decreased the oxidation of D-[U-14C]glucose, but failed to affect L-[U-14C]leucine oxidation. It is concluded that the occurrence of a positive insulinotropic action of L-alanine is restricted to selected experimental conditions, the secretory data being compatible with the view that stimulation of insulin secretion by the tested nutrient(s) reflects, as a rule, their capacity to augment ATP generation in the islet B cells. However, the possible role of Na(+) co-transport in the secretory response to L-alanine cannot be ignored.     

Pancreatic islet function in omega-3 fatty acid-depleted rats: alteration of calcium fluxes and calcium-dependent insulin release

Considering the insufficient supply of long-chain polyunsaturated omega-3 fatty acids often prevailing in Western populations, this report deals mainly with alterations of Ca(2+) fluxes and Ca(2+)-dependent insulin secretory events in isolated pancreatic islets from omega-3-depleted rats. In terms of (45)Ca(2+) handling, the islets from omega-3-depleted rats, compared with those from normal animals, displayed an unaltered responsiveness to an increase in extracellular K(+) concentration, a lower inflow rate and lower fractional outflow rate of the divalent cation, and higher (45)Ca(2+)-labeled cellular pool(s) at isotopic equilibrium. The latter anomaly was corrected 120 min after intravenous injection of a novel medium-chain triglyceride-fish oil (MCT:FO) emulsion, distinct from a control omega-3-poor MCT-olive oil (MCT:OO) emulsion. At 8.3 mM D-glucose, insulin release was higher in islets from omega-3-depleted rats vs. control animals, coinciding with a higher cytosolic Ca(2+) concentration. The relative magnitude of the increase in insulin output attributable to a rise in D-glucose as well as extracellular Ca(2+) or K(+) concentration, to the absence vs. presence of verapamil and to the presence vs. absence of extracellular Ca(2+), theophylline, phorbol 12-myristate 13-acetate, or Ba(2+), was always more pronounced in islets from omega-3-depleted rats injected with the MCT:OO compared with the MCT:FO emulsion. A comparable situation prevailed when comparing islets from noninjected omega-3-depleted and normal rats. In light of these and previous findings, we propose that an impairment of Na(+),K(+)-ATPase activity plays a major, although not an exclusive, role in the perturbation of Ca(2+) fluxes and Ca(2+)-dependent secretory events in the islets from omega-3-depleted rats.     

Changes in the properties of cytosolic acetyl-CoA carboxylase studied in cold-clamped liver samples from fed, starved and starved-refed rats.

The mechanism responsible for the insulin resistance described in vivo in brown adipose tissue (BAT) of lactating rats was investigated. The effect of insulin on glucose metabolism was studied on isolated brown adipocytes of non-lactating and lactating rats. Insulin stimulation of total glucose metabolism is 50% less in brown adipocytes from lactating than from non-lactating rats. This reflects a decreased effect of insulin on glucose oxidation and lipogenesis. However, the effect of noradrenaline (8 microM) on glucose metabolism was preserved in brown adipocytes from lactating rats as compared with non-lactating rats. The number of insulin receptors is similar in BAT of lactating and non-lactating rats. The insulin-receptor tyrosine kinase activity is not altered during lactation, for receptor autophosphorylation as well as tyrosine kinase activity towards the synthetic peptide poly(Glu4-Tyr1). The defect in the action of insulin is thus localized at a post-receptor level. The insulin stimulation of pyruvate dehydrogenase activity during euglycaemic/hyperinsulinaemic clamps is 2-fold lower in BAT from lactating than from non-lactating rats. However, the percentage of active form of pyruvate dehydrogenase is similar in non-lactating and lactating rats (8.6% versus 8.9% in the basal state, and 37.0% versus 32.3% during the clamp). A decrease in the amount of pyruvate dehydrogenase is likely to be involved in the insulin resistance described in BAT during lactation.     

Is glucokinase responsible for the anomeric specificity of glycolysis in pancreatic islets?

At a low concentration of D-glucose (3.3 mM), the phosphorylation rate of this hexose in rat pancreatic islet homogenates incubated at 8 degrees C is higher with the beta- than with the alpha-anomer, as expected from the anomeric specificity of hexokinase. In the presence of a high concentration of glucose 6-phosphate (3.0 mM), which inhibits hexokinase but not glucokinase, the phosphorylation rates of the two anomers are not significantly different from one another. Nevertheless, in intact islets exposed at 8 degrees C to the same low concentration of D-glucose, the alpha-anomer augments, more than the beta-anomer, the production of lactic acid and net uptake of 45Ca. At the same concentration (3.3 mM), the alpha-anomer is also more potent than the beta-anomer in enhancing insulin release from perfused pancreases stimulated at 37 degrees C by L-leucine or by the combination of Ba2+ and theophylline. It is concluded that the participation of glucokinase is not essential for the anomeric specificity of glycolysis and insulin release in rat pancreatic islets.     

The coupling of metabolic to secretory events in pancreatic islets: does hexose transport affect cationic fluxes?

Poorly metabolized hexoses, such as 3-O-methyl-D-glucose, 2-deoxy-D-glucose and D-galactose failed to reproduce the inhibition of 86Rb outflow, the early inhibition and secondary rise in 45Ca efflux and the stimulation of insulin release evoked by D-glucose in perifused rat islets. Insulin release induced by either D-glucose or 2-ketoisocaproate was also unaffected by 3-O-methyl-D-glucose. It is concluded that hexose transport in islet cells does not represent in itself a significant determinant of the cationic and secretory response to D-glucose.     

Stimulation of the insulin secretory mechanism following barium accumulation in pancreatic beta-cells

Electrothermal atomic absorption spectroscopy was employed for measuring barium in beta-cell-rich pancreatic islets microdissected from ob/ob-mice. Both the uptake and efflux of barium displayed two distinct phases. There was a 4-fold accumulation of barium into intracellular stores when its extracellular concentration was 0.26 mM. Unlike divalent cations with more extensive intracellular accumulation, the washout of Ba2+ was not inhibited by D-glucose. Ba2+ served as a substitute for Ca2+ both in maintaining the glucose metabolism after removal of extracellular Ca2+ and making it possible for glucose to stimulate insulin release. Furthermore, Ba2+ elicited insulin release in the absence of glucose and other secretagogues. The latter effect was reversible and was markedly potentiated under conditions known to increase the beta-cell content of cyclic AMP. It is likely that the observed actions of Ba2+ are mediated by Ca2+, since Ca2+ -dependent regulatory proteins, such as calmodulin, apparently cannot bind Ba2+ specifically.     

Regulation of calcium fluxes in rat pancreatic islets: Quinine mimics the dual effect of glucose on calcium movements

The effects of quinine and 9-aminoacridine, two blockers of potassium conductance in islet cells, on ⁴⁵Ca efflux and insulin release from perifused islets were investigated in order to elucidate the mechanisms by which glucose initially reduces ⁴⁵Ca efflux and later stimulates calcium inflow in islet cells. In the absence of glucose, 100 μM quinine stimulated ⁴⁵Ca net uptake, ⁴⁵Ca outflow rate and insulin release. Quinine also dramatically enhanced the cationic and the secretory response to intermediate concentrations of glucose, but had little effect on ⁴⁵Ca net uptake, ⁴⁵Ca fractional outflow rate and insulin release at a high glucose concentration (16.7 mM). The ability of quinine to stimulate ⁴⁵Ca efflux depended on the presence of extracellular calcium, suggesting that it reflects a stimulation of calcium entry in the islet cells. In the absence of extracellular calcium, quinine provoked a sustained decrease in ⁴⁵Ca efflux. Such an inhibitory effect was not additive to that of glucose, and was reduced at low extracellular Na⁺ concentration. At a low concentration (5 μM), quinine, although reducing ⁸⁶Rb efflux from the islets to the same extent as a non-insulinotropic glucose concentration (4.4 mM), failed to inhibit ⁴⁵Ca efflux. In the presence of extracellular calcium, 9-aminoacridine produced an important but transient increase in ⁴⁵Ca outflow rate and insulin release from islets perifused in the absence of glucose. In the absence of extracellular calcium, 9-aminoacridine, however, failed to reduced ⁴⁵Ca efflux from perifused islets. It is concluded that quinine, by reducing K⁺ conductance, reproduces the effect of glucose to activate voltage-sensitive calcium channels and to stimulate the entry of calcium into the B-cell. However, the glucose-induced inhibition of calcium outflow rate, which may also participate in the intracellular accumulation of calcium, does not appear to be mediated by changes in K⁺ conductance.     

Islet-activating protein. Enhanced insulin secretion and cyclic AMP accumulation in pancreatic islets due to activation of native calcium ionophores.

The mechanism whereby "islet-activating protein" (IAP) purified from the culture medium of Bordetella pertussis potentiates insulin secretion was studied by experiments in vitro with islets of rats once injected with IAP (0.5 micrograms/100 g body weight, 3 days before killing) or with islets that had been exposed to IAP (0.1 to 100 ng/ml) for 24 h. The IAP treatment markedly enhanced insulin secretory responses and cAMP accumulation in islets, facilitated the efflux of 45Ca through the cell membrane, and abolished the alpha-adrenergic action of epinephrine (and somatostatin) to inhibit glucose-induced insulin release, cAMP accumulation, and 45Ca uptake. These effects of the IAP treatment were reduced when islets were incubated in a low calcium medium. Based on these results, it was concluded that IAP interacts directly but slowly with the islet B cell in such a manner as to render more calcium available to the stimulus-secretion coupling mechanism as a result of sustained activation of native calcium ionophores on the cell membrane.     

Reduced insulin secretion in response to nutrients in islets from malnourished young rats is associated with a diminished calcium uptake

Changes in (45)Ca uptake and insulin secretion in response to glucose, leucine, and arginine were measured in isolated islets derived from 4-week-old rats born of mothers maintained with normal protein (NP, 17%) or low protein (LP, 6%) diet during pregnancy and lactation. Glucose provoked a dose-dependent stimulation of insulin secretion in both groups of islets, with basal (2.8 mmol/L glucose) and maximal release (27.7 mmol/L glucose) significantly reduced in LP compared with NP islets. In the LP group the concentration-response curve to glucose was shifted to the right compared with the NP group, with the half-maximal response occurring at 16.9 and 13.3 mmol/L glucose, respectively. In LP islets, glucose-induced first and second phases of insulin secretions were drastically reduced. In addition, insulin response to individual amino acids, or in association with glucose, was also significantly reduced in the LP group compared with NP islets. Finally, in LP islets the (45)Ca uptake after 5 minutes or 90 minutes of incubation (which reflect mainly the entry and retention, respectively, of Ca(2+)), was lower than in NP islets. These data indicate that in malnourished rats both initial and sustained phases of insulin secretion in response to glucose were reduced. This poor secretory response to nutrients seems to be the consequence of an altered Ca(2+) handling by malnourished islet cells.     

Arachidonic acid metabolism in isolated pancreatic islets. VI. Carbohydrate insulin secretagogues must be metabolized to induce eicosanoid release.

Pancreatic islets stimulated with D-glucose are known to liberate arachidonic acid from membrane phospholipids and release prostaglandin E2 (PGE2). A component of the eicosanoid release induced by D-glucose has been demonstrated to occur without calcium influx and must be triggered by other coupling mechanisms. In this study, we have attempted to identify mechanisms other than calcium influx which might couple D-glucose stimulation to hydrolysis of arachidonate from membrane phospholipids in islet cells. We have found that occupancy of the beta cell plasma membrane D-glucose transporter is insufficient and that D-glucose metabolism is required to induce islet PGE2 release because 3-O-methylglucose fails to induce and mannoheptulose prevents PGE2 release otherwise induced by 17 mM D-glucose. The carbohydrate insulin secretagogues mannose and D-glyceraldehyde have also been found to induce islet PGE2 release, but the non-secretagogue carbohydrates L-glucose and lactate do not. Carbohydrate secretagogues are known to be metabolized to yield ATP and induce depolarization of the beta cell plasma membrane. We have found that depolarization by 40 mM KCl induces PGE2 release only in the presence and not in the absence of extracellular calcium, but exogenous ATP induces islet PGE2 release with or without extracellular calcium. Carbachol is demonstrated here to interact synergistically with increasing concentrations of glucose to amplify PGE2 release and insulin secretion. Pertussis toxin treatment is shown here not to prevent PGE2 release induced by glucose or carbachol but to increase the basal rate of PGE2 release and the islet cyclic AMP content. Theophylline (10 mM) exerts similar effects. Eicosanoid release in pancreatic islets can thus be activated by multiple pathways including muscarinic receptor occupancy, calcium influx, increasing cAMP content, and a metabolic signal derived from nutrient secretagogues, such as ATP.