Mechanisms of insulin exocytosis and release

Abstract: Endocrine cells as well as neurons release their hormones and transmitters by regulated exocytosis. In the pancreatic B-cell, stimuli like glucose initiate biochemical and electrical processes that culminate in influx of Ca2+, which then triggers exocytosis of insulin-containing granules. Fusion of the secretory vesicles occurs rapidly upon Ca2+-influx but requires a granule to be ?primed? by an ATP-, Ca2+- and temperature-dependent reaction. Only a small fraction of the B-cell's granules (~ 0.5 %) are in the primed state and can undergo exocytosis immediately upon Ca2+-influx. These granules are referred to as the readily releasable pool (RRP), whereas the remaining granules form a ?reserve? pool. The functional organization of the granules in a reserve pool and a readily releasable pool could account for the fact that glucose-stimulated insulin secretion follows a biphasic time course, with the early rapid component (1st phase secretion) corresponding to RRP release and the second slower component reflecting time- and ATP-dependent mobilization of granules from the reserve pool. The commonest form of human diabetes (type-2 diabetes) is associated with disturbances in this release pattern. In this thesis, electrophysiology, fluorescence microscopy and biochemistry were combined to explore mechanisms of granule trafficking, priming, exocytosis, and release in insulin-secreting B-cells. Three aspects are discussed in detail: 1) The importance of a tight interaction between L-type Ca2+-channels and the exocytotic machinery for efficient secretion; 2) A novel ATP-dependent priming reaction that is regulated by a granular 65-kDa sulfonylurea-binding protein, and involves granule acidification and ClC-3 chloride channels; and 3) A previously overlooked delay between fusion of the granule with the plasma membrane and insulin release. Since regulated secretion is very similar in all (neuro)-endocrine cells, the data obtained are likely to be relevant for peptide secretion in general.