Glutamate dehydrogenase (GDH) is regulated by both positive (leucine and ADP) and negative (GTP and ATP) allosteric factors. We hypothesized that the phosphate potential of ␤-cells regulates the sensitivity of leucine stimulation. These predictions were tested by measuring leucine-stimulated insulin secretion in perifused rat islets following glucose depletion and by tracing the nitrogen flux of [2-15 N]glutamine using stable isotope techniques. The sensitivity of leucine stimulation was

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... d by long time (120-min) energy depletion and inhibited by glucose pretreatment. After limited 50-min glucose depletion, leucine, not ␣-ketoisocaproate, failed to stimulate insulin release. ␤-Cells sensitivity to leucine is therefore proposed to be a function of GDH activation. Leucine increased the flux through GDH 3-fold compared with controls while causing insulin release. High glucose inhibited flux through both glutaminase and GDH, and leucine was unable to override this inhibition. These results clearly show that leucine induced the secretion of insulin by augmenting glutaminolysis through activating glutaminase and GDH. Glucose regulates ␤-cell sensitivity to leucine by elevating the ratio of ATP and GTP to ADP and P i and thereby decreasing the flux through GDH and glutaminase. These mechanisms provide an explanation for hypoglycemia caused by mutations of GDH in children.