Readily releasable ß cells with tight Ca2+-exocytosis coupling dictate biphasic glucose-stimulated insulin secretion.

Biphasic glucose-stimulated insulin secretion (GSIS) is essential for blood glucose regulation, but a mechanistic model incorporating the recently identified islet ß cell heterogeneity remains elusive. Here, we show that insulin secretion is spatially and dynamically heterogeneous across the islet. Using a zinc-based fluorophore with spinning-disc confocal microscopy, we reveal that approximately 40% of islet cells, which we call readily releasable ß cells (RRßs), are responsible for 80% of insulin exocytosis events. Although glucose up to 18.2 mM fully mobilized RRßs to release insulin synchronously (first phase), even higher glucose concentrations enhanced the sustained secretion from these cells (second phase). Release-incompetent ß cells show similarities to RRßs in glucose-evoked Ca2+ transients but exhibit Ca2+-exocytosis coupling deficiency. A decreased number of RRßs and their altered secretory ability are associated with impaired GSIS progression in ob/ob mice. Our data reveal functional heterogeneity at the level of exocytosis among ß cells and identify RRßs as a subpopulation of ß cells that make a disproportionally large contribution to biphasic GSIS from mouse islets.

Doc2b serves as a scaffolding platform for concurrent binding of multiple Munc18 isoforms in pancreatic islet ß-cells.

Biphasic glucose-stimulated insulin secretion (GSIS) from pancreatic ß-cells involves soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor (SNARE) protein-regulated exocytosis. SNARE complex assembly further requires the regulatory proteins Munc18c, Munc18-1 and Doc2b. Munc18-1 and Munc18c are required for first- and second-phase GSIS respectively. These distinct Munc18-1 and Munc18c roles are related to their transient high-affinity binding with their cognate target (t-)SNAREs, Syntaxin 1A and Syntaxin 4 respectively. Doc2b is essential for both phases of GSIS, yet the molecular basis for this remains unresolved. Because Doc2b binds to Munc18-1 and Munc18c via its distinct C2A and C2B domains respectively, we hypothesized that Doc2b may provide a plasma membrane-localized scaffold/platform for transient docking of these Munc18 isoforms during GSIS. Towards this, macromolecular complexes composed of Munc18c, Doc2b and Munc18-1 were detected in ß-cells. In vitro interaction assays indicated that Doc2b is required to bridge the interaction between Munc18c and Munc18-1 in the macromolecular complex; Munc18c and Munc18-1 failed to associate in the absence of Doc2b. Competition-based GST-Doc2b interaction assays revealed that Doc2b could simultaneously bind both Munc18-1 and Munc18c. Hence these data support a working model wherein Doc2b functions as a docking platform/scaffold for transient interactions with the multiple Munc18 isoforms operative in insulin release, promoting SNARE assembly.