The Insulin Secretory Granule Is the Major Site of KATP Channels of the Endocrine Pancreas

Abstract

With ATP sites on K ir 6.2 that inhibit activity and ADP sites on SUR1 that antagonize the inhibition, ATPsensitive potassium channels (K ATP channels) are designed as exquisite sensors of adenine nucleotide levels that signal changes in glucose metabolism. If pancreatic K ATP channels localize to the insulin secretory granule, they would be well positioned to transduce changes in glucose metabolism into changes in granule transport and exocytosis. Tests for pancreatic K ATP channels localized to insulin secretory granules led to the following observations: fluorescent sulfonylureas that bind the pancreatic K ATP channel specifically label intracellular punctate structures in cells of the endocrine pancreas. The fluorescent glibenclamides colocalize with Ins-C-GFP, a live-cell fluorescent reporter of insulin granules. Expression of either SUR1-GFP or K ir 6.2-GFP fusion proteins, but not expression of GFP alone, directs GFP fluorescence to insulin secretory granules. An SUR1 antibody specifically labels insulin granules identified by anti-insulin. Two different K ir 6.2 antibodies specifically label insulin secretory granules identified by antiinsulin. Immunoelectron microscopy showed K ir 6.2 antibodies specifically label perimeter membrane regions of the secretory granule. Relatively little or no labeling of other structures, including the plasma membrane, was found. Our results demonstrate that the insulin secretory granule is the major site of K ATP channels of the endocrine pancreas. Diabetes 52: 767-776, 2003 A major question in insulin secretion is the cellular site of action of sulfonylureas, which are taken daily by millions of diabetic subjects to correct hyperglycemia. One site of sulfonylurea action is at the cytoplasmic face of the plasma membrane sulfonylurea receptor (1-5) of the ATP-sensitive potassium channel (K ATP channel) (6). Hundreds of K ATP channels are localized to the plasma membrane of insulin-secreting ␤-cells (7). The pancreatic K ATP channel comprises four regulatory sulfonylurea receptor (SUR1) subunits and four potassium pore-forming (K ir 6.2) subunits (8 -10). The plasma membrane K ATP channel acts like an on/off switch. When on, potassium ions flow out through the channel electrically hyperpolarizing the ␤-cell, putting a brake on the signal flow controlling insulin secretion (11). When off, the K ATP channel, which is inhibited by sulfonylureas or by the increased ATP/ADP ratio from glucose metabolism (12,13), removes this brake, allowing initiation of insulin release by elevating intracellular calcium (14 -17), which triggers the exocytotic fusion of insulin granule and ␤-cell membrane.The calcium signal, however, is insufficient for regulating insulin release in response to glucose under typical conditions (11,18). While pharmacologically either opening or inhibiting the plasma membrane K ATP channel, maneuvers that elevate not only intracellular calcium but also glucose metabolism are necessary to confer glucose dose dependency to stimulated insulin release. By an unknown m...

β IV -Spectrin and CaMKII facilitate Kir6.2 regulation in pancreatic beta cells

“…Whereas we and others have observed Kir6.2 and SUR1 across the beta cell (immunostaining throughout cytosol and membrane, Figs. 1 and 3), K ATP channel subunits have also been observed at the insulin granule (27). Compared with the HEK cell or cardiomyocyte, the pancreatic beta cell is small with few well-defined membrane features.…”

Visualising insulin secretion. The Minkowski Lecture 2004

“…"Mixed" events allow the release of selected vesicle membrane proteins (e.g. VAMP2) [81], with K ATP channel subunits [111] possibly providing a mechanism for delivering proteins ultimately destined to function at the plasma membrane. Finally, (quasi) "full" events permit the release of soluble vesicle cargoes including NPY.Venus (which has approximately the same molecular dimensions as insulin), but do not permit the release of larger proteins such as tissue plasminogen activator (tPA, 70 M r ) or phogrin (Fig.…”

SUR1: a unique ATP-binding cassette protein that functions as an ion channel regulator

“…Thus, it is not inconceivable that it serves as a transporter of an as yet unidentified substrate.There is accumulating evidence, however, that SUR1 also plays a role in insulin granule exocytosis. First, K ATP channels are found at much higher density on insulin granules than in the plasma membrane (Geng et al 2003). Second, insulin granule fusion is reduced in SUR1 knockout mice (Kikuta et al 2005).…”