Perifused Rat Pancreatic Islets Research Articles

Arylcyanoguanidines as activators of Kir6.2/SUR1K ATP channels and inhibitors of insulin release.

Phenylcyanoguanidines substituted with lipophilic electron-withdrawing functional groups, e.g. N-cyano-N'-[3,5-bis-(trifluoromethyl)phenyl]-N' '-(cyclopentyl)guanidine (10) and N-cyano-N'-(3,5-dichlorophenyl)-N' '-(3-methylbutyl)guanidine (12) were synthesized and investigated for their ability to inhibit insulin release from beta cells, to repolarize beta cell membrane potential, and to relax precontracted rat aorta rings. Structural modifications gave compounds, which selectively inhibit insulin release from betaTC6 cells (e.g. compound 10: IC(50) = 5.45 +/- 1.9 microM) and which repolarize betaTC3 beta cells (10: IC(50) = 4.7 +/- 0.5 microM) without relaxation of precontracted aorta rings (10: IC(50) > 300 microM). Inhibition of insulin release from rat islets was observed in the same concentration level as for betaTC6 cells (10: IC(50) = 1.24 +/- 0.1 microM, 12: IC(50) = 3.8 +/- 0.4 microM). Compound 10 (10 microM) inhibits calcium outflow and insulin release from perifused rat pancreatic islets. The mechanisms of action of 10 and 12 were further investigated. The compounds depolarize mitochondrial membrane from smooth muscle cells and beta cell and stimulate glucose utilization and mitochondrial respiration in isolated liver cells. Furthermore, 10 was studied in a patch clamp experiment and was found to activate Kir6.2/SUR1 and inhibit Kir6.2/SUR2B type of K(ATP) channels. These studies indicate that the observed effects of the compounds on beta cells result from activation of K(ATP) channels of the cell membrane in combination with a depolarization of mitochondrial membranes. It also highlights that small structural changes can dramatically shift the efficacy of the cyanoguanidine type of selective activators of Kir6.2/SUR2 potassium channels.

GAD65-mediated Glutamate Decarboxylation Reduces Glucose-stimulated Insulin Secretion in Pancreatic Beta Cells

Mitochondrial metabolism plays a pivotal role in the pancreatic beta cell by generating signals that couple glucose sensing to insulin secretion. We have demonstrated previously that mitochondrially derived glutamate participates directly in the stimulation of insulin exocytosis. The aim of the present study was to impose altered cellular glutamate levels by overexpression of glutamate decarboxylase (GAD) to repress elevation of cytosolic glutamate. INS-1E cells infected with a recombinant adenovirus vector encoding GAD65 showed efficient overexpression of the GAD protein with a parallel increase in enzyme activity. In control cells glutamate levels were slightly increased by 7.5 mm glucose (1.4-fold) compared with the effect at 15 mm (2.3-fold) versus basal 2.5 mm glucose. Upon GAD overexpression, glutamate concentrations were no longer elevated by 15 mm glucose as compared with controls (-40%). Insulin secretion was stimulated in control cells by glucose at 7.5 mm (2.5-fold) and more efficiently at 15 mm (5.2-fold). INS-1E cells overexpressing GAD exhibited impaired insulin secretion on stimulation with 15 mm glucose (-37%). The secretory response to 30 mm KCl, used to raise cytosolic Ca(2+) levels, was unaffected. Similar results were obtained in perifused rat pancreatic islets following adenovirus transduction. This GAD65-mediated glutamate decarboxylation correlating with impaired glucose-induced insulin secretion is compatible with a role for glutamate as a glucose-derived factor participating in insulin exocytosis.

A potent diazoxide analogue activating ATP-sensitive K+ channels and inhibiting insulin release.

To characterise the effects of BPDZ 73 (7-chloro-3-isopropylamino-4H-1,2,4-benzothiadiazine 1,1-dioxide), a newly synthesised diazoxide analogue, on insulin secretory cells. Measurements of 86Rb, 45Ca outflow, membrane potential, [Ca2+]i, insulin release in secretory cells as well as measurements of smooth muscle contractile activity and glycaemia were carried out. The analogue BPDZ 73 induced a dose-dependent decrease in insulin output. The IC50 value averaged 0.73 +/- 0.05 mumol/l. The drug increased the rate of 86Rb (42K substitute) outflow from perifused rat pancreatic islets. This effect was inhibited by glibenclamide, a KATP channel blocker. Measurements of DiBAC4(3) fluorescence further indicated that BPDZ 73 hyperpolarised the insulin secreting cells. It also decreased 45Ca outflow from pancreatic islets perifused throughout in the presence of 16.7 mmol/l glucose and extracellular Ca2+. By contrast, the drug did not affect the increase in 45Ca outflow mediated by K+ depolarisation. In single beta cells, BPDZ 73 inhibited the glucose-induced but not the K(+)-induced rise in [Ca2+]i. Moreover, in Wistar rats, i.p. injection of BPDZ 73 provoked a considerable increase in blood glucose concentration whereas diazoxide induced a modest rise in glycaemia. Lastly, the vasorelaxant properties of BPDZ 73 were slightly less pronounced than those of diazoxide. The inhibitory effect of BPDZ 73 on the insulin-releasing process results from the activation of KATP channels with subsequent decrease in Ca2+ inflow and [Ca2+]i. The drug seems to be a KATP channel opener, more potent and more selective than diazoxide for insulin secreting cells.

Effect of D-mannoheptulose upon the cationic and secretory responses to alpha-D-glucose pentaacetate in perifused rat pancreatic islets.

The metabolism of alpha-D-glucose pentaacetate and its positive insulinotropic action in isolated rat pancreatic islets are both unexpectedly resistant to D-mannoheptulose, as judged from experiments conducted over 90-120 min incubation. In the present study, the possible effects of the heptose upon the immediate cationic and secretory response to the ester were investigated in perifused islets prelabeled with either 86Rb or 45Ca. At a 10 mM concentration, sufficient to abolish the inhibitory action of unesterified D-glucose upon 86Rb outflow, D-mannoheptulose failed to suppress the decrease in 86Rb outflow and increase in 45Ca efflux caused by alpha-D-glucose pentaacetate at normal extracellular Ca2+ concentration and also failed to prevent the decrease in both 45Ca and insulin release provoked by the ester in the absence of extracellular Ca2+. The sole obvious effect of the heptose was to change the early peak-shaped positive secretory response to alpha-D-glucose pentaacetate to a transient inhibition of insulin release. This change was observed in islets either deprived of any other exogenous nutrient or exposed to L-leucine throughout the experiments. These findings support the view that the islet functional response to alpha-D-glucose pentaacetate is largely resistant to D-mannoheptulose. They also reinforce the concept that the insulinotropic action of this and other monosaccharide esters involves a dual modality of B-cell activation, linked to both the catabolism of their carbohydrate moieties and a direct effect of the esters themselves upon a specific receptor system.

Effects of the polyacetate esters of nutrient and nonnutrient monosaccharides on 45calcium efflux and insulin release from perifused rat pancreatic islets.

The polyacetate esters of selected nonnutrient monosaccharides represent potential tools for either stimulation of insulin release in noninsulin-dependent diabetes or inhibition of insulin secretion in hyperinsulinemic syndromes. The polyacetate esters of several monosaccharides were recently shown to display greater nutritional value or biological efficiency than the corresponding unesterified carbohydrates. The effects of seven polyacetate esters of monosaccharides, all tested at a 1.7-mM concentration, on both 45Ca efflux and insulin release were investigated in prelabeled rat pancreatic islets perifused in the presence of 10.0 mM succinic acid dimethyl ester. Both alpha-D-glucose penta-acetate and, to a lesser extent, beta-L-glucose penta-acetate stimulated insulin release. Inversely, alpha-D-galactose penta-acetate, but not beta-D-galactose penta-acetate inhibited insulin secretion evoked by succinic acid dimethyl ester. Esters of carbohydrates which are inhibitors of D-glucose metabolism, such as D-mannoheptulose hexa-acetate and the two anomers of 2-deoxy-D-glucose tetra-acetate also enhanced insulin output, with a preference for the alpha-anomer of 2-deoxy-D-glucose tetra-acetate. Only those esters with positive insulinotropic action augmented 45Ca efflux from the prelabeled islets.

Evidence for a circadian rhythm of insulin release from perifused rat pancreatic islets.

This study aims to analyse a circadian rhythm of insulin secretion from isolated rat pancreatic islets in vitro and its potential modulation by melatonin, the concentrations of which change in vivo inversely to that of insulin. The circadian rhythm was evaluated in a perifusion system, adapted to the specific conditions of pancreatic islets. To determine rhythmicity of insulin secretion, 30-min fractions were collected continuously for investigative periods of 44 to 112 h. Insulin secretion in 10 experiments was analysed by using the MacAnova-program for period length (tau), the chi2-periodogram for test of significance (p < 0.001), and additionally the empirical cosine adaptation for amplitude and goodness-of-fit. Thereby a circadian pattern was observed with periods (tau) between 21.8 and 26.2 h. The period duration (mean +/- SEM) was 23.59 +/- 0.503 h, the overall mean insulin release 1038 +/- 13 pmol/l and the mean amplitude 88 +/- 17 pmol/l. Adding melatonin (10 nmol/l, t = 2 h) as a hormonal Zeitgeber during analysis of circadian insulin secretion phase-response studies show phase-shifts with approximately 9 h phase advance. Thereafter the circadian period was maintained, while the amplitude was enhanced. From this it is concluded that an endogenous circadian oscillator is located within the pancreatic islets of the rat that regulates circadian insulin secretion of the insulin-producing beta cells. The pacemaker is remarkably stable, because its periodicity is not affected by factors altering insulin secretion. In agreement with inhibitory influences of melatonin (range 0.5 nmol/l to 5 micromol/l) on the insulin response in vitro, the phase-responses support the contention that pancreatic beta cells may be targets for melatonin action.

Dynamic insulin secretion from perifused rat pancreatic islets.

Insulin secretion from isolated pancreatic islets of 8- to 12-day-old rats was investigated in a dynamic in vitro (perifusion) system. The aims of the study were (i) to describe a carefully controlled in vitro method to study the mechanism of insulin secretion and to analyse the effects and dynamic interactions of bioactive compounds on isolated rat pancreatic islets, (ii) to validate the method by comparing fundamental data on the functions of the islets obtained with this method to those collected with other techniques; and (iii) to find novel features of the control of insulin secretion. The method was carefully designed to maintain the functional capacity of the explanted cells. A functional standardization system was elaborated consisting of (i) analysis of the changes in the basal hormone secretion of the cells; (ii) evaluating responses to a standard, specific stimuli (50 mM glucose for 3 min); (iii) determining the alteration of the momentary size of the hormone pool with responses to KCl; and (iv) direct determination of the total intracellular hormone content from the extract of the column. The technique provides accurate quantitative data on the dynamic responses to biologically active compounds that act directly on the pancreatic islets. The islets maintained their full responsiveness for up to 7 days, and responses as close as in 1-min intervals could be distinguished. A linear dose-response relationship was found on the glucose-induced insulin release in case of 3-min stimulation with 4 and 500 mM of glucose (lin-log graph). Utilizing this method, we showed that no desensitization to glucose-induced insulin release can be observed if the responsiveness of the cells is properly maintained and the parameters of the stimulation are carefully designed. Exposure of the explanted islets to 10 microM acetylcholine or 30 mM arginine (Arg) induced a transitory elevation of insulin release similar in shape to that experienced after glucose stimulation. Norepinephrine (NE), dopamine (DA) and somatostatin (SS) did not induce any detectable alteration on the basal insulin secretion of the islets. However, 100 nM SS given together with 50 mM glucose, 30 mM Arg or 10 microM acetylcholine significantly reduced the insulin-releasing effect of these substances (by 75.5, 71.5 and 72.5%, respectively). At the same time, SS did not alter the insulin response of the islets to 100 mM elevation of K+ concentration. SS also inhibited glucose-induced insulin release in a dose-dependent way (ED50 = 22 nM). A similar dose-dependent inhibitory effect on glucose-induced insulin release was found with NE (ED50 = 89 nM) and DA (ED50 = 2.2 microM). gamma-Aminobutyric acid (GABA) did not influence insulin release under similar circumstances.

Glucose-induced mobilisation of intracellular Ca 2+ 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 CaCl 2. Either verapamil (0.05 mM) or Cd 2+ (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 Ca 2+ 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 Ca 2+ influx, which was prevented by the absence of extracellular Ca 2+. The present experimental design thus reveals a novel component of the glucose-induced remodelling of Ca 2+ 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.

Verapamil, a phenylalkylamine Ca2+ channel blocker, inhibits ATP-sensitive K+ channels in insulin-secreting cells from rats.

Radioisotopic and electrophysiological techniques were used to assess the effects of verapamil, a phenylalkylamine Ca2+ channel blocker, on K+ permeability of insulin-secreting cells. Verapamil provoked a concentration-dependent inhibition of 86Rb (42K substitute) outflow from prelabelled and perifused rat pancreatic islets. This property appears to be inherent to the phenylalkylamine Ca2+ channel blockers since gallopamil, a methoxyderivative of verapamil, but not nifedipine, a 1,4-dihydropyridine Ca2+ channel blocker, inhibited 86Rb outflow. The experimental data further revealed that verapamil interacted with a Ca2+-independent, glucose- and glibenclamide-sensitive modality of 86Rb extrusion. Moreover, verapamil prevented the increase in 86Rb outflow brought about by BPDZ 44; a potent activator of the ATP-sensitive K+ channel. Single-channel current recordings by the patch clamp technique confirmed that verapamil elicited a dose-dependent inhibition of the ATP-dependent K+ channel. Lastly, under experimental conditions in which verapamil clearly inhibited the ATP-sensitive K+ channels, the drug did not affect 45Ca outflow, the cytosolic free Ca2+ concentration or insulin release. It is concluded that the Ca2+ entry blocker verapamil inhibits ATP-sensitive K+ channels in pancreatic beta cells. This effect was not associated with stimulation of insulin release.

BTS 67 582 stimulates insulin secretion from perifused rat pancreatic islets.

The novel antidiabetic agent BTS 67 582 (1,1-dimethyl-2-[2-(4-morpholinophenyl)]guanidine monofumarate) demonstrated a concentration-dependent stimulation of insulin release in perifused rat pancreatic islets. EC50 values of 7.7 microM and 6.3 microM were obtained for BTS 67 582 in the presence of 8 mM glucose, after islets were pre-equilibrated with 4 and 8 mM glucose respectively. In contrast, there was little or no stimulation of insulin release at substimulatory (4 mM) or maximal stimulatory (15 mM) glucose concentrations. The plasma EC50 value for the glucose lowering effect of BTS 67 582 in fasted normal rats was 3.9 microM indicating a similar potency in vivo. In islets, BTS 67 582 completely antagonised (EC50 value of 13.2 microM) the actions of the selective ATP-dependent K+ channel opener diazoxide indicating K+ channel blocking activity. BTS 67 582 only weakly reversed the alpha2-adrenoceptor mediated inhibition of insulin release in islets (EC50 of 83 microM). BTS 67 582, like other imidazoline/guanidine insulin releasing agents, appears to promote insulin release via an effect on the islet ATP-dependent K+ channel which is not mediated by binding to the sulphonylurea receptor.

Cationic and secretory effects of 6-O-acetyl-D-glucose in rat pancreatic islets.

Selected esters of D-glucose were recently proposed as tools to supply the sugar to cells affected by a defect in the carrier-mediated process of hexose transport across the plasma membrane. The present study extends this knowledge to 6-O-acetyl-D-glucose. At a 2.0 mM concentration, the ester was indeed found to stimulate insulin release from perifused rat pancreatic islets. This coincided with stimulation of calcium influx into the islet cells, as judged from comparison of the ester effects on 45Ca outflow from prelabelled islets perifused in either the absence or presence of extracellular Ca2+. The facilitated inflow of Ca2+ did not appear attributable to a decrease in K+ conductance, 6-O-acetyl-D-glucose augmenting 86Rb efflux from islets prelabelled with this radioactive cation. Both the latter phenomenon and the insulinotropic action of 6-O-acetyl-D-glucose failed to be antagonized by D-mannoheptulose. These findings indicate that the cationic events coupling the recognition of D-glucose to the stimulation of insulin release are not identical in islets exposed to the hexose or its ester.

Influence of melatonin and serotonin on glucose-stimulated insulin release from perifused rat pancreatic islets in vitro.

Insulin plays a key role in the control of glucose homeostasis in mammals. Insulin secretion is regulated by a coordinated interplay of several factors. The role of the indoleamines in the control of insulin secretion has not been fully elucidated yet. The present study was addressed to investigate the function of melatonin and serotonin in the direct control of insulin secretion from the pancreatic islets. Explanted rat Langerhans' islets were treated with melatonin or serotonin while also being exposed to specific (glucose) or non-specific (KCl) stimulus either in a pulsatile or long-term manner in a perifusion system. Insulin content from the effluent tissue culture media was analyzed with RIA. Pulsatile administration of melatonin and serotonin alone did not alter the basal insulin secretion from the explanted islets even at pharmacological (5 microM) level. However, insulin response to specific (glucose) or non-specific (KCl) stimulus was significantly reduced while the islets were treated with melatonin (3 to 12 hr, 10 nM to 5 microM). This effect was reversible and repeatable. Both the start and end of the effect was rapid, evolving and disappearing within 10 min. On the other hand, under similar experimental protocol, serotonin (at 5 microM concentration) significantly enhanced both glucose and KCl stimulated insulin release. Since the effect of the non-specific stimulation (with KCl) was also altered, melatonin and serotonin seem to alter not only the release but also the synthesis of the insulin. Our data show that melatonin and serotonin have a direct effect on the insulin secretion from the pancreatic islets.

Glucose-induced oscillatory insulin secretion in perifused rat pancreatic islets and clonal beta-cells (HIT).

Normal insulin secretion is oscillatory in vivo and from groups of perifused islets. Stimulation of rat islets with different glucose concentrations gave insulin oscillations of similar period (5-8 min) but increasing amplitude. It has been assumed that oscillatory secretion is due to oscillations in intracellular free Ca2+, as seen in single islets and single pancreatic beta-cells. However, when islets were perifused with diazoxide and high KCl to maintain high intracellular free Ca2+, insulin oscillations of similar amplitude and period still occurred on glucose stimulation, although superimposed on elevated basal secretion. Several likely possibilities for a diffusible synchronizing factor were tested, including pyruvate, lactate, ATP, and insulin itself; nevertheless, perifusion with high concentrations of these did not prevent insulin oscillations. Clonal pancreatic beta-cells (HIT) and dissociated islets also exhibited oscillatory insulin secretion, but with the 5- to 8-min period oscillations superimposed on 15- to 20-min period oscillations. These results indicate that the mechanisms for generating and synchronizing insulin oscillations reside in the beta-cell, although the structure of the islet may modulate the oscillation pattern.

Iterative stimulation of pancreatic islets by glipizide.

Iterative administrations of glipizide (2 mumol/l) for 10 min each to perifused rat pancreatic islets provoked at low glucose concentration (2.8 mmol/l) rapid and rapidly reversible decreases in 86Rb outflow, rapid and rapidly reversible increases in 45Ca outflow and modest stimulations of insulin release. In the presence of Ca2+, the reascension in 86Rb outflow was preceded by a transient fall in effluent radioactivity upon withdrawal of the sulfonylurea. At a higher concentration of D-glucose (8.3 mmol/l), glipizide provoked, each time, biphasic and rapidly reversible increases in both 86Rb and 45Ca outflow, as well as in insulin release. These results are compatible with the view that glipizide decreases K+ conductance, leading to depolarization of the B-cell plasma membrane and gating of voltage-sensitive Ca2+ channels. In the presence of 8.3 mmol/l D-glucose, however, the inhibitory effect of glipizide on effluent K+ permeability may be masked by activation of Ca(2+)-sensitive K+ channels. The present data also indicate that glipizide is able to evoke, at the occasion of iterative administrations, biphasic ionic and secretory responses, without evidence of tachyphylaxis.

Comparative effects of LN 5330 and diazoxide on insulin release and 86Rb + permeability in perifused rat islets of Langerhans

The benzothiadiazine derivative LN 5330 (chloro-7 trifluoromethyl-6 benzothiadiazine-1,2,4 dioxide-1,1) has been shown to inhibit insulin secretion and calcium uptake. The present study was carried out to investigate the effects of LN 5330 on insulin release and 86Rb + efflux from perifused rat pancreatic islets; a comparison was made with the structural analogue diazoxide. In the presence of 8.3 mM glucose, LN 5330 (100 μM) accelerated 86Rb + efflux while reducing insulin output from the islets. LN 5330 induced a dose-dependent acceleration of 86Rb + efflux and appeared to be a more potent activator of 86Rb + efflux than diazoxide. The stimulatory effect of LN 5330 on 86Rb + efflux persisted in the absence of extracellular calcium. In the absence of glucose, 86Rb + permeability also increased, LN 5330 being again significantly more efficient than diazoxide at an equimolar concentration. These data indicate that the benzothiadiazine derivative LN 5330 inhibits insulin secretion by increasing the potassium permeability of the plasma membrane. It is suggested that, like diazoxide, this drug could activate the ATP-sensitive K + channel.

Content of CoA-Esters in Perifused Rat Islets Stimulated by Glucose and Other Fuels

Fluorometry and high-performance liquid chromatography were used to measure the content of free CoA and the esters of acetate, malonate, succinate, and long-chain fatty acids in isolated perifused rat pancreatic islets exposed to 25 mM glucose or a mixture of fuels (25 mM glucose plus 10 mM glutamine, 10 mM lactate, and 1 mM pyruvate) to assess the role of intermediates of lipid metabolism as candidate metabolic coupling factors in the mechanism of fuel-induced insulin secretion. Insulin secretion was stimulated in a biphasic manner with the fuel mixture, showing twice the potency compared with high glucose alone. Islets perifused for 3 min with high glucose alone or the fuel mixture compared with 2.5 mM glucose showed a significant increase in malonyl-CoA and succinyl-CoA and a decrease in acetyl-CoA. Free CoA and long-chain acyl-CoA levels were unaltered. Perifused islets stimulated with 25 mM glucose for 30 min showed a significant increase in succinyl-CoA and long-chain acyl-CoA and decrease in acetyl-CoA, whereas malonyl-CoA was not affected. However, when islets were stimulated by the fuel mixture for 30 min, malonyl-CoA was maintained at a high level, and the change in succinyl-CoA and long-chain acyl-CoA was similar to that observed in islets stimulated with 25 mM glucose alone. The acetyl-CoA concentration in the islets stimulated with the fuel mixture decreased slightly. These results confirm the viability of the hypothesis that malonyl-CoA and long-chain acyl-CoA serve as metabolic coupling factors in signal transduction when islets are stimulated by high glucose or glucose combined with other fuels.

Chronic exposure to high glucose and impairment of K(+)-channel function in perifused rat pancreatic islets

Chronic exposure to high glucose and impairment of K(+)-channel function in perifused rat pancreatic islets

Chronic Exposure to High Glucose and Impairment of K+-Channel Function in Perifused Rat Pancreatic Islets

Prolonged exposure to high glucose levels impairs the ability of pancreatic islets to secrete insulin as a response to that stimulus. Because glucose, like other insulin secretagogues, elicits insulin secretion by inhibiting the ATP-sensitive K+ channels, in this study, we investigated the effect of prolonged (24-h) exposure of rat pancreatic islets to high (16.7 mM) glucose concentration on 86Rb efflux (used as a tracer for K+). The data obtained indicate that islets exposed to high glucose concentration have impaired function of the glucose-sensitive K+ channel, this phenomenon is temporarily related to a defective response of glucose-induced insulin release, and these alterations are reversible.

Effect of Galanin on Arginine-Stimulated Pancreatic Hormone Release from Isolated Perifused Rat Islets

The effect of galanin on pancreatic hormone release was studied using isolated perifused rat pancreatic islets. In the presence of 100 mg/dl glucose, 10(-8) mol/L galanin significantly inhibited the basal somatostatin release compared with the perifusion without galanin, whereas there was no significant change in the basal insulin and glucagon release. However, under stimulation of 20 mmol/L arginine, 10(-8) mol/L galanin significantly enhanced glucagon release and suppressed insulin and somatostatin release. These effects disappeared immediately after cessation of galanin infusion. Additionally, 10(-8) mol/L galanin significantly enhanced the first and second phase of glucagon release stimulated by arginine, whereas arginine-stimulated insulin and somatostatin releases were significantly inhibited in both phases. In the cysteamine-treated rat islets, neither enhancement of glucagon release nor suppression of insulin release by galanin was reproducible. These findings indicate two possible explanations. First, it is suggested that the effects of galanin on insulin and glucagon release may be direct and reversed by non-specific effect of cycteamine. Secondly, it seems likely that galanin-enhanced glucagon release may be indirect and in part due to the concomitant somatostatin suppression. Galanin may have an important regulatory function on endocrine pancreas.

Effects of lactate on pancreatic islets. Lactate efflux as a possible determinant of islet-cell depolarization by glucose.

The secretion of insulin from perifused rat pancreatic islets was stimulated by raising the glucose concentration from 5.6 to 20 mM or by exposure to tolbutamide. The addition of sodium lactate (40 mM) to islets perifused in the presence of glucose (5.6 mM) resulted in a small, transient, rise in the rate of secretion. The subsequent removal of lactate, but not glucose or tolbutamide, from the perifusate produced a dramatic potentiation of insulin release. The rate of efflux of 45Ca2+ was also increased when islets were exposed to a high concentration of glucose or lactate or to tolbutamide, and again subsequently upon withdrawal of lactate. Efflux of 86Rb+ was modestly inhibited upon addition of lactate and markedly enhanced by the subsequent withdrawal of lactate from islets. The output of [14C]lactate from islets incubated in the presence of [U-14C]glucose increased linearly with increasing concentrations of glucose (1-25 mM). It is proposed that the activation of islets by the addition or withdrawal of lactate is not due to increased oxidative flux, but occurs as a result of the electrogenic passage of lactate ions across the plasma membrane, resulting in islet-cell depolarization, Ca2+ entry and insulin secretion. The production of lactate via the glycolytic pathway, and the subsequent efflux of lactate from the islet cells with concomitant exchange of H+ for Na+, could be a major determinant of depolarization and hence insulin secretion, in response to glucose.