Conversion of glutamate into proline by the leucine analog BCH enhances biphasic insulin secretion in pancreatic β-cells

Biphasic insulin secretion, which fails in type 2 diabetes, can be provoked by various nutrient stimuli, glucose being the superior physiological one. To identify pathways that may play a role in β-cell stimulus-secretion coupling, we compared β-cell and islet functional, secretory, and metabolic responses to glucose and 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH), a leucine analog, that acts as an allosteric activator of glutamate dehydrogenase. We employed a range of techniques, including insulin secretion assays, mitochondrial activity measurements, ATP/ADP ratio assessments, and cytosolic Ca²⁺ level quantifications. Metabolomics was used to analyze cellular metabolite profiles in response to glucose and BCH. Additionally, we investigated the role of proline synthesis by silencing ALDH18A1, encoding proline 5-carboxylate synthase, in both clonal β-cells and human islets. BCH and glucose similarly induced a biphasic insulin response in INS-1 832/13 cells, paralleled by increased mitochondrial activity and raised ATP/ADP ratios, plasma membrane depolarization, and elevated cytosolic Ca²⁺ levels. Metabolomics revealed that proline levels increased significantly only in BCH-stimulated β-cells. Silencing ALDH18A1 disrupted insulin secretion in response to both glucose and BCH, accompanied by reduced cytosolic Ca²⁺ levels, ATP/ADP ratios, and mitochondrial activity. Our findings demonstrated that BCH-induced activation of glutamate dehydrogenase leads to the conversion of glutamate into proline, which apparently enhances β-cell stimulus-secretion coupling. This work identifies a previously unrecognized role of proline metabolism in β-cell function and provides novel insights into the complex regulation of insulin secretion.