The stimulus-secretion coupling of amino acid-induced insulin release metabolic interaction of L-asparagine and L-leucine in pancreatic islets

Because L-asparagine augments insulin release evoked by L-leucine, the metabolism of these two amino acids was investigated in rat pancreatic islets. L-Leucine inhibited the uptake and deamidation of L-asparagine, but failed to exert any obvious primary effect upon the further catabolism of aspartate derived from exogenous asparagine. L-Asparagine augmented the oxidation of L-leucine, an effect possibly attributable to activation of 2-ketoisocaproate dehydrogenase. The association of L-asparagine and L-leucine exerted a sparing action on the utilization of endogenous amino acids, so that the integrated rate of nutrients oxidation was virtually identical in the sole presence of L-leucine and simultaneous presence of L-asparagine and L-leucine, respectively. It is proposed that the enhancing action of L-asparagine upon insulin release evoked by L-leucine is attributable to an increased generation rate of cytosolic NADPH rather than any increase in nutrients oxidation.     

The stimulus-secretion coupling of amino acid-induced insulin release

Summary Aminooxyacetate, an inhibitor of cytosolic transamination reactions, inhibited insulin release evoked by either 2-ketoisocaproate or L-leucine in rat pancreatic islets incubated in the presence of L-glutamine or L-asparagine. As a rule, aminooxyacetate also inhibited the oxidation of these nutrient secretagogues and impaired the respiratory response of the islets to the combinations of nutrients. However, the oxidative and secretory response to the combination of L-leucine and L-glutamine was less severely affected by aminooxyacetate than that evoked by the three other combinations of exogenous nutrients. These findings reinforce the view that the stimulus-secretion coupling of insulin release in response to L-leucine and 2-ketoisocaproate in association with either L-glutamine or L-asparagine tightly depends on the oxidation of these nutrient secretagogues, on their effect upon O₂ uptake and, within limits, on the intracellular site of generation of reducing equivalents in the pancreatic islet cells.This paper is the 16th in a series.     

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.     

The stimulus-secretion coupling of amino acid-induced insulin release synergistic effects of L-glutamine and 2-keto acids upon insulin secretion

1. L-Glutamine markedly enhances insulin release evoked in rat pancreatic islets by 2-ketoisocaproate or 2-ketocaproate. L-Glutamine exerts a lesser enhancing action in the presence of 2-ketovalerate or 2-ketoisovalerate, which are themselves poor insulin secretagogues. L-Glutamine fails to affect insulin release in the presence of 2- ketobutyrate, pyruvate and β-hydroxybutyrate. 2. The relase of insulin evoked by the combination of L-glutamine and 2-ketoisocaproate represents a sustained phenomenon. It coincides with a stimulation of ⁴⁵Ca net uptake by the islets, and is inhibited in the absence of extracellular Ca²⁺ and presence of either menadione or epinephrine. 3. L-Valine inhibits insulin releaseevoked by either 2-ketoisocaproate or 2-ketocaproate, whether in the presence or absence of L-glutamine, but does not abolish the enhancing action of L-glutamine. L-Valine fails to affect insulin release evoked by the combination of L-leucine and L-glutamine. 4. L-Isoleucine also inhibits 2-keto acid-induced insulin release. However, in contrast to L-valine, L-isoleucine fails to affect or slightly augments insulin release in the simultaneous presence of L-glutamine and either 2-ketoisocaproate or 2-ketocaproate. 5. L-Leucine causes a dose-related enhancement of insulin release evoked by the combination of 2-ketoisocaproate and L-glutamine. Likewise, in the presence of L-glutamine, L-leucine and 2-ketocaproate act synergistically upon insulin release. 6. The hypothesis is advances that the enhancing action of L-glutamine upon 2-keto acid-stimulated insulin release depends on the availability of the 2-keto acid to act as a partner in the conversion of L-glutamate derived from exogenous L-glutamine to 2-ketoglutarate by transamination reaction, rather than being attributable to activation of glutamate dehydrogenase as observed in islets exposed to both L-glutamine and L-leucine.     

Participation of endogenous fatty acids in the secretory activity of the pancreatic B-cell.

The pancreatic B-cell may represent a fuel-sensor organ, the release of insulin evoked by nutrient secretagogues being attributable to an increased oxidation of exogenous and/or endogenous substrates. The participation of endogenous fatty acids in the secretory response of isolated rat pancreatic islets was investigated. Methyl palmoxirate (McN-3716, 0.1 mM), an inhibitor of long-chain-fatty-acid oxidation, suppressed the oxidation of exogenous [U-14C]palmitate and inhibited 14CO2 output from islets prelabelled with [U-14C]palmitate. Methyl palmoxirate failed to affect the oxidation of exogenous D-[U-14C]glucose or L-[U-14C]glutamine, the production of NH4+ and the output of 14CO2 from islets prelabelled with L-[U-14C]glutamine. In the absence of exogenous nutrient and after a lag period of about 60 min, methyl palmoxirate decreased O2 uptake to 69% of the control value. Methyl palmoxirate inhibited insulin release evoked by D-glucose, D-glyceraldehyde, 2-oxoisohexanoate, L-leucine, 2-aminobicyclo[2.2.1]heptane-2-carboxylate or 3-phenylpyruvate. However, methyl palmoxirate failed to affect insulin release when the oxidation of endogenous fatty acids was already suppressed, e.g. in the presence of pyruvate or L-glutamine. These findings support the view that insulin release evoked by nutrient secretagogues tightly depends on the overall rate of nutrient oxidation, including that of endogenous fatty acids.     

The coupling of metabolic to secretory events in pancreatic islets. The possible role of glutathione reductase

The participation of glutathione reductase in the process of nutrient-stimulated insulin release was investigated in rat pancreatic islets exposed to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). BCNU caused a time-and dose-related, irreversible inhibition of glutathione reductase activity. This coincided with a fall in both GSH/GSSG ratio and the thiol content of the islets. Pretreatment of the islets with BCNU inhibited the oxidation of glucose and its stimulant action upon both 45Ca net uptake and insulin release. Although BCNU (up to 0.5 mM) failed to affect the oxidation of L-leucine and L-glutamine, it also caused a dose-related inhibition of insulin release evoked by the combination of these two amino acids. The latter inhibition was apparently not fully accounted for by the modest to negligible effects of BCNU upon 45Ca uptake, 45Ca efflux, 86Rb efflux and cyclic AMP production. Since BCNU failed to inhibit insulin release evoked by the association of Ba2+ and theophylline, these results support the view that glutathione reductase participates in the coupling of metabolic to secretory events in the process of nutrient-stimulated insulin release. However, the precise modality of such a participation, for example the control of intracellular Ca2+ distribution, remains to be elucidated.     

Mechanism of 3-phenylpyruvate-induced insulin release. Secretory, ionic and oxidative aspects.

1. 3-Phenylpyruvate caused a dose-related stimulation of insulin release from rat pancreatic islets deprived of exogenous nutrient or incubated in the presence of 5.6 or 8.3 mM-D-glucose. 2. 3-Phenylpyruvate inhibited insulin release evoked by high concentrations of D-glucose (16.7 or 27.8 mM) or 4-methyl-2-oxopentanoate (10.0 mM). This inhibitory effect appeared to be attributable to impairment of 2-oxo-acid transport into the mitochondria, with resulting inhibition of D-glucose, pyruvate or 4-methyl-2-oxopentanoate oxidation. 3. 3-Phenylpyruvate failed to affect the oxidation of, and secretory response to, L-leucine, and did not augment insulin release evoked by a non-metabolized analogue of the latter amino acid. 4. L-Glutamine augmented 3-phenylpyruvate-induced insulin release. The release of insulin evoked by the combination of 3-phenylpyruvate and L-glutamine represented a sustained phenomenon, abolished in the absence of extracellular Ca2+ or the presence of menadione and potentiated by theophylline. 5. Whether in the presence or in the absence of L-glutamine, the secretory response to 3-phenylpyruvate coincided with an increase in O2 uptake, a decrease in K+ conductance, a stimulation of both Ca2+ inflow and 45Ca2+ net uptake and an increase in cyclic AMP content. 6. It is concluded that the release of insulin induced by 3-phenylpyruvate displays features classically encountered when the B-cell is stimulated by nutrient secretagogues, and is indeed attributable to an increase in nutrient catabolism.     

Interference of a nonmetabolized analogue of L-leucine with lipid metabolism in tumoral pancreatic islet cells

The nonmetabolized analogue of L-leucine, 2-aminobicyclo[2,2,1]heptane-2-carboxylic acid (BCH), was recently found to inhibit O2 uptake and insulin release from tumoral islet cells of the RINm5F line. BCH inhibited lipogenesis, stimulated lipolysis, and severely decreased the oxidation of endogenous [U-14C]palmitate in prelabelled RINm5F cells. D-Glucose exerted metabolic effects which were sometimes opposite to those caused by BCH and, within limits, protected the islet cells against the inhibitor action of BCH. Since BCH augments NH4+ production and facilitated the catabolism of 14C-labelled amino acids in the prelabelled cells, it is proposed that the unexpected inhibition of O2 uptake by BCH is mainly attributable to a decrease in the oxidation of endogenous fatty acids.     

Insulin release: Reconciliation of the receptor and metabolic hypotheses

Summary Nutrients which stimulate insulin secretion are currently thought to initiate the series of cellular events eventually leading to insulin release either by interacting with a stereospecific receptor system (the regulatory site hypothesis) or by acting as a fuel (the substrate site hypothesis) in the pancreaticB-cell. The latter hypothesis is supported by a number of observations indicating that the capacity of nutrients to stimulate insulin release is indeed highly dependent on their capacity to increase catabolic fluxes in isolated pancreatic islets. However, these observations do not rule out the existence of nutrient receptors in islet cells. For instance, a nonmetabolized analog of L-leucine stimulates insulin release by causing allosteric activation of glutamate dehydrogenase, which should be considered, therefore, as a receptor for certain amino acids. Likewise, the increase in glycolytic flux, which is associated with the process of glucose-stimulated insulin release, is attributable not solely to a mass action phenomenon but also to the activation of phosphofructokinase by fructose 2.6-bisphosphate. The biosynthesis of this activator may involve a glucose receptor system. The fact that certain nutrient secretagogues (e.g D-glucose and L-leucine) act in the B-cell both as substrates and enzyme activators permits reconciliation of the substrate site and regulatory site hypotheses for insulin release.     

Effects of amino acids on the isolated chicken retina, and on its response to glutamate stimulation

Abstract— The effect of a number of amino acids on the transparency and on the release of of [¹⁴C]glutamate from isolated chicken retinae charged with this compound was investigated. Also the effect of various amino acids on the response of the retina to stimulation with unlabelled glutamate, which causes an increase in transparency and a release of the label, was examined. In parallel experiments the effect of these same amino acids on the transparency and spreading depression (SD) was investigated in preparations consisting of the posterior part of the eye. A number of amino acids such as L-leucine, L-phenylalanine, L-tryptophan, L-lysine, L-histidine, L-arginine and others had little or no effect on these preparations. DL-valine and DL-homoserine caused an increase in transparency but no release of the label and did not affect the response to glutamate. Another group of amino acids comprising DL-a-alanine, L-serine, L-threonine, L-proline and glycine also caused an increase in the transparency of the retina without a release of labelled glutamate, but prevented the increase in transparency resulting from glutamate stimulation without affecting the release of the label. A final group of amino acids which included L-glutamic acid diethyl ester, DL-a-methyl glutamate, L-glutamine, L-asparagine, DL-homocysteate and L-cysteine caused a change in transparency of the retina accompanied by a release of the label; they prevented the change in transparency as well as the release of the label during stimulation by glutamate. Some amino acids, L-serine, L-threonine, DL-a-methyl glutamate, L-asparagine, DL-homocysteate and L-cysteine, caused wrinkling and folding of the retinae which furthermore became opaque. Of the amino acids investigated, proline gave promise of being a practical antagonist to the action of glutamate on the retina.     

Does leucine- and norleucine-induced insulin release depend on amino acid aminotransferase activity?

In the absence of another exogenous nutrient, L-leucine but not L-norleucine stimulates insulin release from rat pancreatic islets, although the corresponding keto acids, 2-ketoisocaproate and 2-ketocaproate, are equally potent secretagogues. Such a situation cannot be ascribed to the preferential transamination of L-leucine as compared to L-norleucine in islet homogenates. Indeed, in the presence of a suitable activator of glutamate dehydrogenase, L-leucine and L-norleucine stimulate secretion to the same extent. It is concluded that the rate of transamination of these amino acids in intact islet cells depends on the availability of a 2-keto acid partner rather than on the assayed amino acid aminotransferase activity.     

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.     

Failure of parathormone to affect insulin and glucagon release from the perfused rat pancreas.

Parathormone (0.15 U/ml) failed to affect the rate of glucagon and insulin release by the perfused rat pancreas exposed to glucose in either low (3.3 mM) or high (8.3 mM) concentration. Parathormone also failed to interfere with the suppressive effect of glucose (16.6mM) upon glucagon release and its stimulatory action upon insulin secretion. Likewise, the biphasic release of both glucagon and insulin evoked by arginine (10.0 mM) in the presence of glucose (8.3 mM) was unaffected by parathormone. These findings suggest that the endocrine pancreas may not be a target organ for any direct and immediate action of parathormone.     

The stimulus-secretion coupling of glucose-induced insulin release. Effect of exogenous pyruvate on islet function.

1. In isolated pancreatic islets, pyruvate causes a shift to the left of the sigmoidal curve relating the rate of insulin release to the ambient glucose concentration. The magnitude of this effect is related to the concentration of pyruvate (5--90 mM) and, at a 30 mM concentration, is equivalent to that evoked by 2 mM-glucose. Pyruvate also enhances insulin release in the presence of fructose, leucine and 4-methyl-2-oxopentanoate. 2. In the presence of glucose 8 mM), the secretory response to pyruvate is an immediate process, displaying a biphasic pattern. 3. The insulinotropic action of pyruvate coincides with an inhibition of 45Ca efflux and a stimulation of 45Ca net uptake. The relationship between 45Ca uptake and insulin release displays its usual pattern in the presence of pyruvate. 4. Exogenous pyruvate rapidly accumulates in the islets in amounts close to those derived from the metabolism of glucose. The oxidation of [2-14C]pyruvate represents 64% of the rate of [1-14C]pyruvate decarboxylation and, at a 30 mM concentration, is comparable with that of 8 mM-[U-14C]glucose. 5. When corrected for the conversion of pyruvate into lactate, the oxidation of 30 mM-pyruvate corresponds to a net generation of about 314 pmol of reducing equivalents/120 min per islet. 6. Pyruvate does not affect the rate of glycolysis, but inhibits the oxidation of glucose. Glucose does not affect pyruvate oxidation. 7. Pyruvate (30 mM) does not affect the concentration of ATP, ADP and AMP in the islet cells. 8. Pyruvate (30 mM) increases the concentration of reduced nicotinamide nucleotides in the presence but not in the absence of glucose. A close correlation is seen between the concentration of reduced nicotinamide nucleotides and the net uptake of 45Ca. Menadione inhibits the effect of pyruvate on insulin release, without altering its rate of oxidation. 9. Pyruvate, like glucose, modestly stimulates lipogenesis. 10. Pyruvate, in contrast with glucose, markedly inhibits the oxidation of endogenous nutrients. The latter effect accounts for the apparent discrepancy between the rate of pyruvate oxidation and the magnitude of its insulinotropic action. 11. Dichloroacetate fails to affect glucose oxidation and glucose-stimulated insulin release. 12. It is concluded that the effect of pyruvate to stimulate insulin release depends on its ability to increase the concentration of reduced nicotinamide nucleotides in the islet cells.     

The stimulus-secretion coupling of glucose-induced insulin release. LIII. Calcium-dependency of the cyclic AMP response to nutrient secretagogues

Above a threshold value in excess of 5.6 mM, D-glucose increases the amount of cyclic AMP measured by radioimmunoassay in pancreatic rat islets and their surrounding incubation medium. As judged from the cyclic AMP content of islets exposed to isobutylmethylxanthine (1.0 mM), the glucose-induced increment in the rate of cyclic AMP generation represents a rapid and sustained phenomenon. The stimulant action of glucose on cyclic AMP accumulation is mimicked by L-leucine, and L-glutamine, these amino acids acting synergistically of one another. Trifluoperazine slightly decreases but fails to abolish the effect of glucose. In the absence of extracellular Ca2+, however, the cyclic AMP response to D-glucose, L-leucine and/or L-glutamine is severely impaired. These findings are compatible with the view that an increase in the generation rate of cyclic AMP participates in the process of nutrient-stimulated insulin release. This increase could be secondary to the nutrient-induced accumulation of Ca2+ in the islet cells leading to activation of adenylate cyclase by calmodulin.     

The Riddle of Formycin A Insulinotropic Action

Formycin A augments insulin release evoked by glucose (5.6 mmor 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 decrease⁸⁶Rb efflux, this coinciding with a low efficiency of formycin A 5′-triphosphate to inhibit K_ATP-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 of⁴⁵Ca. 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 β-cell remains to be elucidated.     

Inhibition by a nonmetabolized analogue of L-leucine of protein biosynthesis in tumoral pancreatic islet cells

The effect of L-leucine, its deaminated metabolite 2-ketoisocaproate and its nonmetabolized analogue b(+/-)2-aminobicyclo[2,2,1]-heptane-2-carboxylic acid (BCH) upon protein labelling was examined in tumoral islet cells (RINm5F line) exposed to L-[4-3H]phenylalanine or L-[3-3H]serine. The interpretation of the results, in terms of changes in biosynthetic activity, was obscured by a possible interference of the tested nutrients with the uptake and further metabolism of the tracer tritiated amino acids. Nevertheless, when the cells were preincubated with the nutrient secretagogues and then incubated in the sole presence of L-[3-3H]serine, BCH, but not L-leucine or 2-ketoisocaproate, still inhibited protein labelling, this coinciding with a decrease in the ratio between TCA-precipitable and total radioactivity in the RINm5F cells. The inhibitory action of BCH was antagonized, to a limited extent, by D-glucose. It is proposed that BCH could be used as a tool to interfere with the function and growth of insulinoma cells.     

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.     

Transamination of neutral amino acids and 2-keto acids in pancreatic B-cell mitochondria

High aminotransferase activities catalyzing the reactions between L-glutamate and L-glutamine and the aliphatic ketomonocarboxylic acids 2-ketoisocaproate, 2-ketocaproate, and 2-ketoisovalerate were observed in pancreatic B-cell mitochondria. While maximal rates of transamination with L-glutamate were observed in the presence of micromolar concentrations of keto acid, maximal rates of transamination with L-glutamine were recorded only in the presence of millimolar concentrations of keto acid. The insulin secretagogue 2-ketoisocaproate was the most effective transamination partner for L-glutamate, while the insulin secretagogue 2-ketocaproate was the most effective transamination partner for L-glutamine. Since B-cell mitochondria are well supplied with L-glutamate and L-glutamine, 2-ketoglutarate generation in the presence of these two neutral 2-keto acids may be an important prerequisite for their insulin secretory potency. High rates of transamination of 2-ketoglutarate were observed in the pancreatic B-cell mitochondria with the branched-chain amino acids L-leucine and L-valine, but not with L-norleucine. In connection with the ability of L-leucine to activate glutamate dehydrogenase, this high activity of the branched-chain amino acid aminotransferase in pancreatic B-cell mitochondria may provide an explanation for the insulin secretory potency of this amino acid.     

Stimulation of insulin biosynthesis in isolated mouse islets by L-leucine, 2-aminonorbornane-2-carboxylic acid and α-ketoisocaproic acid

An immune binding technique was used for measuring the effects of certain amino acids on the rate of insulin biosynthesis. [³H]phenylalanine served as the radioactive precursor for insulin synthesized by isolated mouse pancreatic islets. L-Leucine was found to stimulate the insulin biosynthesis and this effect was observed already at a physiologic concentration in contrast to the much higher concentrations needed to stimulate insulin secretion in vitro. Furthermore, it was found that 2-aminonorbornane-2-carboxylic acid and α-ketoisocaproic acid shared with glucose and L-leucine the ability to stimulate insulin biosynthesis. In contrast, L-alanine, L-arginine and D-leucine had no stimulatory effect in the absence of glucose, while in the presence of 5 mM glucose L-arginine decreased and L-alanine increased the incorporation rate of tritiated phenylalanine. The fact that many of those compounds which stimulated insulin biosynthesis have also been shown elsewhere to be metabolized by the B-cells supports the view that the rate of insulin biosynthesis may be substrate dependent.