Elimination of KATP Channels in Mouse Islets Results in Elevated [U-13C]Glucose Metabolism, Glutaminolysis, and Pyruvate Cycling but a Decreased γ-Aminobutyric Acid Shunt

Changhong Li,Ilana Nissim,Habiba Najafi,Yevgeny Daikhin,Itzhak Nissim,Heather W Collins,Carol Buettger,Pan Chen,Marc Yudkoff,Franz M Matschinsky,Charles A Stanley

doi:10.1074/jbc.m709235200

Abstract

Pancreatic beta cells are hyper-responsive to amino acids but have decreased glucose sensitivity after deletion of the sulfonylurea receptor 1 (SUR1) both in man and mouse. It was hypothesized that these defects are the consequence of impaired integration of amino acid, glucose, and energy metabolism in beta cells. We used gas chromatography-mass spectrometry methodology to study intermediary metabolism of SUR1 knock-out (SUR1(-/-)) and control mouse islets with d-[U-(13)C]glucose as substrate and related the results to insulin secretion. The levels and isotope labeling of alanine, aspartate, glutamate, glutamine, and gamma-aminobutyric acid (GABA) served as indicators of intermediary metabolism. We found that the GABA shunt of SUR1(-/-) islets is blocked by about 75% and showed that this defect is due to decreased glutamate decarboxylase synthesis, probably caused by elevated free intracellular calcium. Glutaminolysis stimulated by the leucine analogue d,l-beta-2-amino-2-norbornane-carboxylic acid was, however, enhanced in SUR1(-/-) and glyburide-treated SUR1(+/+) islets. Glucose oxidation and pyruvate cycling was increased in SUR1(-/-) islets at low glucose but was the same as in controls at high glucose. Malic enzyme isoforms 1, 2, and 3, involved in pyruvate cycling, were all expressed in islets. High glucose lowered aspartate and stimulated glutamine synthesis similarly in controls and SUR1(-/-) islets. The data suggest that the interruption of the GABA shunt and the lack of glucose regulation of pyruvate cycling may cause the glucose insensitivity of the SUR1(-/-) islets but that enhanced basal pyruvate cycling, lowered GABA shunt flux, and enhanced glutaminolytic capacity may sensitize the beta cells to amino acid stimulation.

Highlights

The pancreatic beta cells function as the predominant sensors and regulators of glucose, amino acid, and fatty acid levels
Profiles of Major Amino Acids in SUR1Ϫ/Ϫ and Control Islets as Influenced by Glucose, methionine sulfoximine (MSO), and VGB—Earlier studies had indicated that incubation of cultured mouse pancreatic islets for 2 h in Krebs-Ringer bicarbonate buffer solution that lacked amino acids resulted in a substantial reduction of the intracellular amino acid pool by 1/2 to 2/3 [6, 13]
Respiration in the absence of external fuel is comparable in SUR1Ϫ/Ϫ islets and controls, but stimulation of respiration by high glucose is substantially less pronounced in SUR1Ϫ/Ϫ islets as compared with controls [7]

Summary

Introduction

The pancreatic beta cells function as the predominant sensors and regulators of glucose, amino acid, and fatty acid levels. We used the conversion or incorporation of 13C carbon into CO2 and various amino acids, respectively, as readout for the integration of metabolic pathways This approach allowed us to examine for the first time in isolated mouse islets and in one comprehensive study the concentration dependences of glucose oxidation and glutaminolysis, the operation of the so called “pyruvate cycling,” the nature of the glucose induced aspartate decrease ( referred to as “aspartate switch”), the regulatory role of glutamate decarboxylase (GAD) in the operation of the beta cell GABA shunt, and glucose stimulation of glutamine synthesis. Metabolic Integration and Insulin Secretion creatic islet cells as contrasted with studies using tumor-derived beta cell lines [8, 9]

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