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J C Hutton

Publications and source records attributed to J C Hutton.

At least 127 records · Page 7Linked to original sources
PubMed1981

Insulin secretion by a transplantable rat islet cell tumour.

Investigation of the subcellular and molecular components of insulin secretion has been made difficult by the small quantities of material available. The recent development of a transplantable rat islet cell tumour of high insulin content and state of differentiation suggested a system more amenable to analysis. To validate the tumour as a model of secretion we have studied its release of insulin. In acute experiments in vitro immunoreactive insulin release was increased by leucine, glucagon, theophylline and dibutyryl cyclic AMP, though not by glucose. Leucine (20 mmol/l) plus theophylline (5 mmol/l) caused an abrupt, sustained and rapidly reversible stimulation of two- to fivefold. The response was inhibited by antagonists of cellular oxidative phosphorylation (cyanide, 2,4-dinitrophenol, antimycin A), calcium flux (EGTA, verapamil, Mg2+), calmodulin (trifluoperazine), microtubules (vinblastine, colchicine) and by adrenaline and somatostatin. These findings suggest that the tumour secretes insulin by an exocytotic mechanism similar to that of normal islet tissue.

Adenoma, Islet Cell↗
PubMed1980

Interaction of branched chain amino acids and keto acids upon pancreatic islet metabolism and insulin secretion.

Insulin release evoked by 2-ketoisocaproate from isolated rat pancreatic islets was inhibited by L-valine, L-isoleucine, L-norvaline, and L-norleucine. The stereo-specificity of the inhibition and the finding that the decrease in secretory rate was correlated to the capacity of these amino acids to undergo transamination with the keto acid secretagogue suggested that a branched chain amino acid aminotransferase enzyme was the site of interaction of these substrates. The stimulation of transamination by these amino acids was accompanied by a decrease in the primary decarboxylation and oxidation of 2-ketoisocaproate in the tissue and a reduction in islet acetoacetate production. The oxidation of the amino acid substrate was slightly increased in the presence of 2-ketoisocaproate. Nevertheless, the oxidation rate of exogenous substrates and respiration were higher in the presence of 2-ketoisocaproate than when this substrate was combined with an amino acid. The action of 2-ketoisocaproate to increase the ratios of NADH/NAD+ and NADPH/NADP+, to enhance the uptake of 45Ca by islet tissue, and to promote proinsulin biosynthesis was also suppressed by the addition of branched chain amino acids to the incubation media. Radioisotopic uptake measurements suggested that many of these observations could be related to changes in the intracellular concentration of 2-ketoisocaproate in islet tissue.

Amino Acids, Branched-Chain↗
PubMed1980

Dynamics of O2 consumption in rat pancreatic islets.

The O2 consumption of rat pancreatic islets was determined by monitoring pO2 in the perifusate from groups of 200-300 islets. Basal respiration was maintained for up to 2 h. The insulin secretagogues, glucose and 4-methyl-2-oxopentanoate, provoked an immediate (< 5 s) increase in islet respiration which attained a new steady-state within 10-40 min. The respiratory changes were immediately reversible upon removal of the substrate and were paralleled by changes in insulin release and substrate oxidation. The concentration dependence of glucose-induced respiratory changes was sigmoidal with a threshold at 3 mmol/l. The concentration dependence with 4-methyl-2-oxopentanoate was characterised by a hyperbolic relationship. The weak insuln secretagogues 3-methyl-2-oxobutyrate and d,l-3-methyl-2-oxopentanoate, although stimulating islet respiration were not more effective than 4- methyl-2-oxopentanoate at non-insulinotropic concentrations. Rotenone, antimycin and oligomycin inhibited both basal O2 consumption and the ability of glucose and 4-methyl-2-oxopentanoate to increase islet respiration. 2,4-Dinitrophenol increased islet O2 consumption. The omission of Ca2+ and Mg2+ from the perifusing media, or the addition of the ionophore A23187, had little effect on respiration. The omission of K+ inhibited glucose-induced changes but had a lesser effect in the absence of substrate or in the presence of 4-methyl-2-oxopentanoate. The omission of HCO3-reduced both basal and secretagogue-induced changes in islet respiration. It is concluded that mitochondrial O2 consumption linked to oxidative phosphorylation is a major component in the respiratory response, and that some energy consuming process in the islets depends on the availability of HCO3-. Mitochondrial reactions may generate a signal initiating the secretory process.

Acetoacetates↗
PubMed1980

The stimulus-secretion coupling of glucose-induced insulin release. XLII. Effects of extracellular pH on insulin release: their dependency on nutrient concentration.

Changes in extracellular pH affected insulin output from pancreatic islets stimulated with either glucose or alpha-ketoisocaproate. The extracellular pH at which the highest secretory response occurred was not identical in all cases, being shifted to alkaline values as the concentration of the nutrient secretagogue was increased. The output of insulin correlated with the nutrient-induced increment in 45Ca net uptake, but not with the rate of nutrient oxidation. By reference to changes in intracellular pH observed in islets exposed to either acidic media or increasing concentrations of alpha-ketoisocaproate, and taking into account a titration curve for the buffering capacity of islet homogenates, the optimal intracellular pH for insulin release was found to approximate a value of 0.09 pH unit below basal pH. It is postulated that, in the process of nutrient-induced insulin release, modest changes in intracellular pH participate in the multifactorial coupling between metabolic, ionic and secretory events.

Animals↗
PubMed1980

Similarities in the stimulus-secretion coupling mechanisms of glucose- and 2-keto acid-induced insulin release.

The stimulus-secretion coupling of 2-keto acid-induced insulin release was investigated using 2-ketoisocaproate (4-methyl-2-oxopentanoate) as the principal model secretagogue. 2-Ketoisocaproate and 2-ketocaproate (2-oxo-, hexanoate) provoked changes in B cell electrical behavior characterized by an initial depolarization of the membrane potential, followed by rapid spike activity, which appeared either in a bursting pattern or as continuous activity. The onset of spike activity induced by 2-ketoisocaproate (5 mM) was biphasic in nature. The dynamic pattern of 2-ketoisocaproate-induced insulin release was also biphasic. 2-[U-14C]Ketoisocaproate (10 mM) was oxidized in islet tissue at a rate equivalent to that of [U-14C]glucose (17 mM) and a t a higher rate than 2-ketoisovalerate (3-methyl-2-oxobutyrate) and 2-keto-3-methyl-valerate, which were poor secretagogues. Like glucose, 2-ketoisocaproate provoked characteristic changes in 86Rb and 45Ca efflux from prelabeled islets and stimulated 45Ca net uptake. Proinsulin synthesis was stimulated by 2-ketoisocaproate through both a general effect on protein synthesis and a specific effect on hormonal biosynthesis. 2-Ketoisocaproate and 2-ketocaproate reproduced the effect of glucose on the islet content of ATP, ADP, AMP, NAD+, NADH, NADP+, and NADPH. These findings together with a series of observations on the effects upon the above parameters of site-specific inhibitors, e.g. respiratory inhibitors, suloctidil, theophylline, and epinephrine, suggested that the stimulus-secretion-coupling mechanisms for 2-ketoisocaproate- and glucose-induced release are similar. It is postulated that glucose- and 2-keto acid-induced insulin release may be initiated by a common signal.

Adenine Nucleotides↗
PubMed1980

The stimulus-secretion coupling of amino acid-induced insulin release: metabolism and cationic effects of leucine.

When isolated rat pancreatic islets are exposed to L-leucine (20 mM), the rate of NH4 production is close to the summed rates of L-[1-14C] leucine decarboxylation and alpha-ketoisocarproate production, whereas the rates of acetoacetate production and L-[U-14C]-leucine oxidation are compatible with conversion of each mole of the amino acid to one mole of acetoacetate and three moles of CO2. ATP content, ATP/ADP ratio, and adenylate charge are maintained at normal values by L-leucine, whereas the NADH/NAD+ ratio (but not the NADPH/NADP+ ratio) is significantly increased. The release of insulin evoked by L-leucine is potentiated by 2-ketoisovalerate, unaffected by L-valine, and inhibited by menadione. L-leucine mimicks the effect of D-glucose on 86Rb+ and 45Ca2+ handling by the islets. However, relative to its rate of oxidation, the insulinotropic effect of L-leucine is less marked than that of D-glucose. This may be due, in part at least, to a decrease in the oxidation of endogenous nutrients. It is concluded that the metabolic, cationic, and secretory effects of L-leucine in isolated islets are not incompatible with the fuel hypothesis for insulin release.

Acetoacetates↗
PubMed1980

Fuel and signal function of 2-keto acids in insulin secretion.

The stimulation of insulin release from pancreatic islet tissue by 2-ketoisocaproate was accompanied by a characteristic electrophysiological response and by increases in islet net uptake of 45Ca and respiratory activity. These parameters were closely correlated to changes in islet keto acid content, islet NADPH/NADP+ ratio and to the rates of 2-ketoisocaproate catabolism and amination. The production of 14CO2 from [U-14C]2-ketoisocaproate was reduced by the addition of valine, isovalerate, glutamine or pyruvate to the incubation media. Such an effect, however, did not correspond to the response of insulin secretion of 45Ca uptake observed in the presence of these substrates. These findings suggest that the metabolic sequence between the initial conversion of 2-ketoisocaproate to isovaleryl CoA and its subsequent transformation to acetoacetate and acetyl CoA was not the site of generation of a signal which initiated insulin release. The total rate of oxidation deduced from the sum of the rates of 14CO2 production from [U-14C]2-ketoisocaproate and the [U-14C]-labelled second substrate, however, did follow the secretory response of the tissue. It is proposed that reactions associated with the tricarboxylic acid cycle are important to the stimulus-secretion coupling mechanism of 2-ketoisocaproate-induced insulin release. The question of whether such reactions participate in the generation of a signal which initiates insulin secretion or merely reflect the energetic demands of the secretory process is discussed.

Animals↗
PubMed1980

The interplay between metabolic and cationic events in islet cells: coupling factors and feedback mechanisms.

In the mechanism of glucose-stimulated insulin release, the coupling between glucose metabolism and the remodelling of cationic fluxes in the B-cell apparently represents a multifactorial process involving changes in the generation rate of H+, reducing equivalents and ATP. This process is susceptible to feedback regulatory mechanisms through which primary changes in cationic movements affect glucose metabolism. The interplay between metabolic and ionic events may participate in the rhythmogenesis of bioelectrical and secretory phenomena.

Adenosine Triphosphate↗
PubMed1979

The metabolism of 4-methyl-2-oxopentanoate in rat pancreatic islets.

1. Radioactively labelled 4-methyl-2-oxopentanoate was taken up by isolated pancreatic islets in a concentration- and pH-dependent manner and led to the intracellular accumulation of labelled amino acid and to a decrease in the intracellular pH. Uptake of 4-methyl-2-oxopentanoate did not appear to be either electrogenic or Na+-dependent. The islet content of 2-oxo acid radioactivity was not affected by either 2-cyano-3-hydroxy-cinnamate (10mM) or pyruvate (10mM), although both these substances inhibited the oxidation of [U-14C]4-methyl-2-oxopentanoate by islet tissue. 2. 4-Methyl-2-oxopentanoate markedly stimulated islet-cell respiration, ketone-body formation and biosynthetic activity. The metabolism of endogenous nutrients by islets appeared to be little affected by the compound. 3. Studies with the 3H- and 14C-labelled substrate revealed that 4-methyl-2-oxopentanoate was incorporated by islets into CO2, water, acetoacetate, L-leucine and to a lesser extent into islet protein and lipid. Carbon atoms C-2, C-3 and C-4 of the acetoacetate produced were derived from the carbon skeleton of the 4-methyl-2-oxopentanoate, but the acetoacetate carboxy group was derived from the incorporation of CO2. These results, and consideration of the relative rates of 14CO2 and acetoacetate formation from 1-14C-labelled as opposed to U-14C-labelled 4-methyl-2-oxopentanoate, led to the conclusion that the pathway of catabolism of this 2-oxo acid in pancreatic islets is identical with that described in other tissues. The amination of 4-methyl-2-oxopentanoate by islets was attributed to the presence of a branched-chain amino acid aminotransferase (EC 2.6.1.42) activity in the tissue. Although glutamate dehydrogenase activity was demonstrated in islet tissue, the reductive amination of 2-oxoacids did not seem to be of importance in the formation of leucine from 4-methyl-2-oxopentanoate. 4. The results of experiments with respiratory inhibitors and uncouplers, and the finding that 14CO2 production and islet respiration were linked in a 1:1 stoicheiometry suggested that 4-methyl-2-oxopentanoate catabolism was coupled to mitochondrial oxidative phosphorylation. The catabolism of 4-methyl-2-oxopentanoate in islet tissue appeared to be regulated at the level of the initial 2-oxo acid dehydrogenase (EC 1.2.1.25) reaction.

Acetoacetates↗
PubMed1979

The stimulus--secretion coupling 4-methyl-2-oxopentanoate-induced insulin release.

1. Pancreatic islet insulin secretion and 45Ca uptake showed similar responses to variation in the extracellular concentration of 4-methyl-2-oxopentanoate with a threshold at 4 mM and a maximal response at a 25 mM concentration. 2. Islet respiration, acetoacetate production and rates of substrate utilization, oxidation and amination all changed as a simple hyperbolic function of 4-methyl-2-oxopentanoate concentration and exhibited a maximal response at 25 mM. 3. The responses of ATP content, [ATP]/[ADP] ratio, adenylate energy charge and [NADH]/[NAD+] ratio were also hyperbolic in nature but were maximally elevated at lower concentrations of the secretagogue. The islet [NADPH]/[NADP+] ratio, however, was tightly correlated with parameters of metabolic flux, 45Ca uptake and insulin release. 4. NH4+ and menadione, agents that promote a more oxidized state in islet NADP, did not affect islet ATP content or the rates of [U-14C]4-methyl-2-oxopentanoate oxidation or amination, but markedly inhibited islet 45Ca uptake and insulin release. 5. It is proposed that changes in the redox state of NADP and Ca transport may serve as mediators in the stimulus-secretion coupling mechanism of insulin release induced by 4-methyl-2-oxopentanoate.

Adenine Nucleotides↗
PubMed1979

The stimulus-secretion coupling of glucose-induced insulin release. XXXV. The links between metabolic and cationic events.

When isolated rat islets were exposed to glucose, the concentrations of NADH and NADPH, and the NADH/NAD+ and NADPH/NADP+ ratios were increased. The dose-response curve resembled that characterising the glucose-induced secondary rise in 45Ca efflux, displaying a sigmoidal pattern with a half-maximal value at glucose 7.5 mmol/l. The glucose-induced increase in NAD(P)H was detectable within 1 min of exposure to the sugar. Except for the fall in ATP concentration and ATP/ADP ratio found at very low glucose concentrations (zero to 1.7 mmol/l) no effect of glucose (2.8-27.8 mmol/l) upon the steady-state concentration of adenine nucleotides was observed. However, a stepwise increase in glucose concentration provoked a dramatic and transient fall in the ATP concentration, followed by a sustained increase in both O2 consumption and oxidation of exogenous + endogenous nutrients. This may be essential to meet the energy requirements in the stimulated B-cell. Although no significant effect of glucose upon intracellular pH was detected by the 5,5-dimethyloxazolidine-2,4-dione method, the net release of H+ was markedly increased by glucose, with a hyperbolic dose-response curve (half-maximal response at glucose 2.9 mmol/l) similar to that characterising the glucose-induced initial fall in 45Ca efflux. It is proposed that the generation of both NAD(P)H and H+ participates in the coupling of glucose metabolism to distal events in the secretory sequence, especially the ionophoretic process of Ca2+ inward and outward transport, and that changes in these parameters occur in concert with an increased turn-over rate of high-energy phosphate intermediates.

Adenine Nucleotides↗
PubMed1979

Insulin release: the fuel hypothesis.

The immediate and direct regulation of insulin release by circulating nutrients, especially glucose, is thought to be mediated in the pancreatic B-cell by a sequence of metabolic, ionic, and motile events. On the basis of previous work, it is assumed that the process by which glucose is recognized as an insulinotropic agent entirely depends on the metabolic changes evoked by the sugar in the islet cells. Several factors are considered as possible candidates for the coupling between these metabolic changes and subsequent ionic events such as altered phosphate, chloride, sodium, potassium, and calcium handling. It is acknowledged that changes in the concentrations of glycolytic intermediates and cyclic nucleotides (adenosine- or guanosine-3', 5'-cyclic monophosphate), or both, could play a modulatory role upon stimulated insulin release. However, the initiation of insulin release seems to depend on the generation of two essential coupling factors: H+ and reduced pyridine nucleotides. The changes in H+ fluxes may account for the glucose-induced decrease in K+ and Ca2+ fractional outflow rate, all three parameters displaying hyperbolic-like dose-response curves with half-maximal values at noninsulinotropic glucose concentrations. The changes in NAD(P)H concentration may account for a glucose-induced Ca2+--Ca2+ exchange process due to a change in affinity of a native ionophoretic system. The dose-response curves for these parameters yield a sigmoidal pattern analogous to that which depicts the rate of insulin release at increasing glucose concentrations. It is proposed that such a coupling between metabolic and cationic events is operative in response to other insulinotropic nutrients and that its time course may be relevant to the phasic aspect of insulin release. Thus, the nutrient-induced release of insulin (and possibly other pancreatic hormones), which is essential for the regulation of fuel homeostasis, would depend on the capacity of circulating nutrients to act as a fuel in the islet cells. This concept raises a question as to the existence and nature of feedback mechanisms regulating the metabolic fluxes in the islet cells as a function of their energy expenditure.

Adenosine Triphosphate↗
PubMed1979

Role of microtubules in the synthesis, conversion, and release of (pro)insulin. A biochemical and radioautographic study in rat islets.

In the pancreatic B cell, microtubules are thought to be involved in the process of insulin release. Their possible participation in the sequence of events leading from the biosynthesis and conversion of proinsulin to the release of newly synthesized insulin was investigated in rat isolated islets exposed to colchicine (0.1 mM). When the islets were preincubated for 30 min with colchicine and [3H]-leucine and, thereafter, incubated for two successive periods of 90 min each, still in the presence of colchicine, the release of preformed insulin was progressively inhibited and that of newly synthesized hormone delayed. When the islets were preincubated for 120 min with colchicine, subsequently pulse-labeled with [3H]leucine, and eventually examined by ultrastructural autoradiography, the export of newly synthesized proinsulin out of the rough endoplasmic reticulum, its transit through the Golgi complex, and its eventual packaging in secretory granules were all retarded. This situation was associated with a delayed conversion of proinsulin to insulin. Under the same experimental conditions, colchicine failed to affect the oxidation of glucose and adenylate charge in the islets. The effect of colchicine upon the release of preformed and newly synthesized insulin was not reproduced by lumicolchicine. It is concluded that colchicine interferes with the system controlling the intracellular transfer of secretory material from site of synthesis to site of release. This interference is likely to be linked to the effect of colchicine on microtubules.

Adenine Nucleotides↗
PubMed1979

Relationship of endogenous to dietary sorbitol. A study in normal and diabetic rats.

The relationship of endogenous to dietary sorbitol was investigated by measuring the sorbitol content of plasma, lens and kidney in normal an streptozotocin-injected rats fed for two-three weeks a control diet or a diet rich in either sorbitol or sucrose. The insulin content and secretory capacity of the pancreatic islets removed from these rats was also measured. In normal rats, the carbohydrate-rich diet increased the concentration of sorbitol in plasma but not lens or kidney. The carbohydrate-rich diet also increased the secretory responsiveness of the B-cell to glucose. Accumulation of sorbitol in lens and kidney occurred in diabetic rats. When the diabetic rats were treated with insulin, the sorbitol content of plasma and lens was less elevated than in untreated diabetic animals, despite poor control of the glycemia. The insulin treatment failed to prevent sorbitol accumulation in the kidney. There was no obvious difference between dietary sorbitol and sucrose respectively, in terms of sorbitol accumulation in tissues or stimulation of B-cell secretory potential.

Animals↗
PubMed1978

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.

Animals↗
PubMed1978

The stimulus-secretion coupling of glucose-induced insulin release. Metabolic effects of menadione in isolated islets.

Pancreatic islets contain an enzyme system which catalyzes the donation of hydrogen from NAD(P)H to menadione (2-methyl-1,4-naphthoquinone). In high concentrations (20 to 50 micrometer), menadione, in addition to lowering the concentration of reduced pyridine nucleotides in the islets, also impairs glycolysis and glucose oxidation, decreases ATP concentration, and inhibits proinsulin biosynthesis. However, at a 10 micrometer concentration, menadione fails to affect the concentration of adenine nucleotides, the utilization of glucose, the production of lactate and pyruvate, the oxidation of [6-14C]glucose and the synthesis of proinsulin; whereas the metabolism of glucose through the pentose shunt is markedly increased. The sole inhibitory effect of menadione 10 micrometer upon metabolic parameters is to reduce the concentration of both NADH and NADPH, such an effect being noticed in islets exposed to glucose 11.1 mM but not in those incubated at a higher glucose level (27.8 mM). Since, in the presence of glucose 11.1 mM, menadione 10 micrometer also severely decreases glucose-stimulated45 calcium net uptake and subsequent insulin release, it is concluded that the availability of reduced pyridine nucleotides may play an essential role in the secretory sequence by coupling metabolic to cationic events. Thus, when insulinotropic nutrients are oxidized in the B-cell, the increased availability of reduced pyridine nucleotides could modify the affinity for cations of native ionophoretic systems, eventually leading to the accumulation of calcium up to a level sufficient to trigger insulin release.

Adenine Nucleotides↗