1737-P: Bulk GABA Secretion from Pancreatic Islets Is Not Co-released with Insulin

Introduction & Objective: Gamma Aminobutyric Acid (GABA) is a neurotransmitter found in inhibitory neurons but is also synthesized and secreted at high levels in pancreatic islets by beta cells. In neurons, GABA is packaged into synaptic vesicles by the vesicular GABA transporter (VGAT) for regulated secretion. In beta cells, evidence exists for regulated GABA secretion from insulin granules and for tonic GABA release from the cytosol via volume regulated anion channels (VRAC) with an osmotic dependence. Since GABA is a potent paracrine and autocrine signal in the islet, determining the dominant mechanism of release is vital to understand its function in the islet. Methods: To interrogate GABA and insulin co-secretion, isolated BL/6 mouse islets were stimulated with glucose, treated with and without diazoxide and forskolin, and analyzed for supernatant insulin and GABA by ELISA and HPLC. The relationship between GABA secretion and VGAT in beta cells was investigated via immunostaining, single-cell RNA-seq dataset (HPAP), Vgat-ires-Cre x Ai14 tdTomato reporter mice, and VGAT-GFP adenoviral beta cell transduction. Results: Diazoxide blocked while forskolin enhanced glucose-stimulated insulin release, but neither drug affected GABA secretion, indicating bulk GABA secretion is not correlated with insulin at the whole islet level. VGAT reporter mice showed many tdTomato+ neurons in the CNS but no tdTomato expression in islets. Immunostaining of islets for VGAT and analysis of the HPAP dataset showed VGAT was undetectable in beta cells. VGAT-GFP overexpression in human beta cells failed to localize VGAT to vesicles, although mouse hippocampal neurons with the same construct showed co-localization with synaptic boutons. Conclusion: Low expression and/or poor vesicular localization of VGAT indicates regulated secretion of GABA from insulin granules appears unlikely to be the dominant release mechanism from whole islets, suggesting tonic release of GABA plays a more significant role in islet physiology. Disclosure A.E. Stis: None. S. Ferreira: None. C. Lazimi: None. E. Phelps: Research Support; Immunocore, Ltd, MESO SCALE DIAGNOSTICS, LLC. Funding National Institutes of Health (R01DK124267)