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Hexose metabolism in pancreatic islets. Glucose-induced and Ca(2+)-dependent activation of FAD-glycerophosphate dehydrogenase.

ABSTRACT: A rise in extracellular D-glucose concentration causes in pancreatic islets a preferential stimulation of aerobic, relative to total, glycolysis. The possible participation in such a phenomenon of a glucose-induced and Ca(2+)-dependent activation of FAD-glycerophosphate dehydrogenase was investigated. In islet homogenates, the activity of the mitochondrial and Ca(2+)-responsive FAD-glycerophosphate dehydrogenase was about two orders of magnitude lower than that of the cytosolic and Ca(2+)-insensitive NAD-glycerophosphate dehydrogenase. In islet mitochondria, Ca2+ increased the affinity of the FAD-glycerophosphate dehydrogenase for L-glycerol 3-phosphate, but did not affect the maximal reaction velocity. In the presence of 0.1 mM-L-glycerol 3-phosphate, the Ka for Ca2+ was close to 0.1 microM. When intact islets were preincubated in the presence of both D-glucose and Ca2+, the activity of FAD-glycerophosphate dehydrogenase measured in intact mitochondria incubated in the presence of 1.2 microM-Ca2+ was higher than that recorded under the same conditions in islets preincubated in the absence of D-glucose and/or Ca2+. These findings support the view that, in islets exposed to a high concentration of D-glucose, a Ca(2+)-induced activation of mitochondrial FAD-glycerophosphate dehydrogenase favours the transfer of reducing equivalents by the glycerol phosphate shuttle, and hence accounts, in part at least, for the preferential stimulation of aerobic glycolysis.

SUBMITTER: Rasschaert J

PROVIDER: S-EPMC1151345 | biostudies-other | 1991 Sep

REPOSITORIES: biostudies-other

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Project description:1. Rates of glucose oxidation, lactate output and the intracellular concentration of glucose 6-phosphate were measured in mouse pancreatic islets incubated in vitro. 2. Glucose oxidation rate, measured as the formation of (14)CO(2) from [U-(14)C]glucose, was markedly dependent on extracellular glucose concentration. It was especially sensitive to glucose concentrations between 1 and 2mg/ml. Glucose oxidation was inhibited by mannoheptulose and glucosamine but not by phlorrhizin, 2-deoxyglucose or N-acetylglucosamine. Glucose oxidation was slightly stimulated by tolbutamide but was not significantly affected by adrenaline, diazoxide or absence of Ca(2+) (all of which may inhibit glucose-stimulated insulin release), by arginine or glucagon (which may stimulate insulin release) or by cycloheximide (which may inhibit insulin synthesis). 3. Rates of lactate formation were dependent on the extracellular glucose concentration and were decreased by glucosamine though not by mannoheptulose; tolbutamide increased the rate of lactate output. 4. Islet glucose 6-phosphate concentration was also markedly dependent on extracellular glucose concentration and was diminished by mannoheptulose or glucosamine; tolbutamide and glucagon were without significant effect. Mannose increased islet fructose 6-phosphate concentration but had little effect on islet glucose 6-phosphate concentration. Fructose increased islet glucose 6-phosphate concentration but to a much smaller extent than did glucose. 5. [1-(14)C]Mannose and [U-(14)C]fructose were also oxidized by islets but less rapidly than glucose. Conversion of [1-(14)C]mannose into [1-(14)C]glucose 6-phosphate or [1-(14)C]glucose could not be detected. It is concluded that metabolism of mannose is associated with poor equilibration between fructose 6-phosphate and glucose 6-phosphate. 6. These results are consistent with the idea that glucose utilization in mouse islets may be limited by the rate of glucose phosphorylation, that mannoheptulose and glucosamine may inhibit glucose phosphorylation and that effects of glucose on insulin release may be mediated through metabolism of the sugar.

Project description:1. Glucose-phosphorylating and glucose 6-phosphatase activities, glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, NADP(+)-linked isocitrate dehydrogenase, ;malic' enzyme and pyruvate carboxylase were assayed in homogenates of normal mouse islets. 2. Two glucose-phosphorylating activities were detected; the major activity had K(m) 0.075mm for glucose and was inhibited by glucose 6-phosphate (non-competitive with glucose) and mannoheptulose (competitive with glucose). The other (minor) activity had a high K(m) for glucose (mean value 16mm) and was apparently not inhibited by glucose 6-phosphate. 3. Glucose 6-phosphatase activity was present in amounts comparable with the total glucose-phosphorylating activity, with K(m) 1mm for glucose 6-phosphate. Glucose was an inhibitor and the inhibition showed mixed kinetics. No inhibition of glucose 6-phosphate hydrolysis was observed with mannose, citrate or tolbutamide. The inhibition by glucose was not reversed by mannoheptulose. 4. 6-Phosphogluconate dehydrogenase had K(m) values of 2.5 and 21mum for NADP(+) and 6-phosphogluconate respectively. 5. Glucose 6-phosphate dehydrogenase had K(m) values of 4 and 22mum for NADP(+) and glucose 6-phosphate. The K(m) for glucose 6-phosphate was considerably below the intra-islet concentration of glucose 6-phosphate at physiological extracellular glucose concentrations. The enzyme had no apparent requirement for cations. Of a number of possible modifiers of glucose 6-phosphate dehydrogenase, only NADPH was inhibitory. The inhibition by NADPH was competitive with NADP(+) and apparently mixed with respect to glucose 6-phosphate. 6. NADP(+)-isocitrate dehydrogenase was present but the islet homogenate contained little, if any, ;malic' enzyme. The presence of pyruvate carboxylase was also demonstrated. 7. The results obtained are discussed with reference to glucose phosphorylation and glucose 6-phosphate oxidation in the intact mouse islet, and the possible nature of the beta-cell glucoreceptor mechanism.

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Hexose metabolism in pancreatic islets. Glucose-induced and Ca(2+)-dependent activation of FAD-glycerophosphate dehydrogenase.

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