With ATP sites on K(ir)6.2 that inhibit activity and ADP sites on SUR1 that antagonize the inhibition, ATP-sensitive 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 anti-insulin. 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.