ORF14-07 - Glutamine Metabolism Supports Glucagon Secretion and Alpha-Cell Mass

Pancreatic α cells are central regulators of metabolic homeostasis through glucagon secretion and amino acid sensing, yet the metabolic mechanisms coupling nutrient availability to α-cell growth and function remain poorly defined. Here, we identify glutaminase, the enzyme that catalyzes glutamine catabolism, as an essential regulator of α-cell proliferation and hormone secretion. Single-cell RNA sequencing of human and mouse islets revealed marked enrichment of glutaminase in α cells (log2fold expression: 1.659±0.001993 ) relative to β (log2fold expression: 0.1968±0.0008958) and δ (log2fold expression: 0.2645±0.003942) cells, with expression of both cytosolic (KGA) and mitochondrial (GAC) isoforms. Consistent with this enrichment, α cells accounted for approximately 70% of total glutaminase activity in intact mouse islets, despite representing a minority of islet cells. Pharmacological inhibition of glutaminase via CB-839 in isolated islets reduced α-cell proliferation (DMSO: 6.773±1.361% Ki67+ α cells, CB-839: 0.1141±0.05521% Ki67+ α cells), mimicking results from glutamine deprivation (high glutamine: 7.457±1.052% Ki67+ α cells, low glutamine: 0.3231±0.1752% Ki67+ α cells). In vivo, tamoxifen-inducible, α-cell-specific glutaminase knockout mice (αGlsKO) had reduced glutaminase activity by ~3.25-fold in isolated islets compared to control mice (αGlsWT). αGlsKO mice also had reduced α-cell expansion as indicated by reduced α-cell mass (αGlsWT mAb: 4.144±0.4501mg; αGlsKO mAb: 2.182±0.1466) and mTORC1 activation (αGlsWT mAb: 88.36±1.730% pS6+ α cells; αGlsKO mAb: 38.08±4.943% pS6+ α cells) in response to elevated amino acids following glucagon receptor blockade. Mechanistically, glutaminase was required for nutrient-dependent mTORC1 activation, suppression of the AMPK–TSC2 axis, activation of mTORC2–AKT signaling, and sustained expression of the glutamine transporter SLC38A5, establishing a feed-forward loop linking glutamine metabolism to amino acid sensing and α-cell growth. Notably, loss of glutaminase activity impaired glucagon secretion in response to low glucose, arginine, and glutamine in both mouse and human isolated islets, indicating a fundamental role for glutamine metabolism in α-cell secretory competence. Together, these findings identify glutaminase as a central metabolic regulator of α-cell growth and glucagon secretion and highlight glutamine metabolism as a key driver of α-cell adaptation under metabolic stress.