Visualization of sequential exocytosis in rat pancreatic islet beta cells

Biochem Biophys Res Commun. 2002 Apr 12;292(4):980-6. doi: 10.1006/bbrc.2002.6712.


The examination of insulin exocytosis at the single cell level by conventional electrophysiologic and amperometric methods possesses inherent limitations, and may not accurately reflect the morphologic events of exocytosis of the insulin granule. To overcome some of these limitations, we show by epifluorescent microscopy of a fluorescent dye, FM1-43, its incorporation into the plasma membrane and oncoming insulin granules undergoing exocytosis, and their core proteins. Using this method, we tracked exocytosis in real-time in insulinoma INS-1 and single rat islet beta cells in response to KCl and glucose. We observed both single transient and multi-stepwise increases in membrane FM1-43 fluorescence, suggesting single granule exocytosis as well as sequential and compound exocytosis, respectively. Confocal microscopy of nonpermeabilized cells shows that some of the exocytosed insulin granules labeled by the FM1-43 dye could also be labeled with insulin antibodies, suggesting prolonged openings of the fusion pores and slow dissolution of the granule core proteins on the membrane surface.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Membrane / drug effects
  • Cells, Cultured
  • Exocytosis / drug effects
  • Exocytosis / physiology*
  • Fluorescent Dyes
  • Glucose / pharmacology
  • Insulin / metabolism
  • Insulin Secretion
  • Insulinoma / drug therapy
  • Insulinoma / metabolism
  • Islets of Langerhans / cytology
  • Islets of Langerhans / drug effects
  • Islets of Langerhans / metabolism*
  • Male
  • Microscopy, Confocal
  • Microscopy, Fluorescence
  • Potassium Chloride / pharmacology
  • Rats
  • Rats, Sprague-Dawley
  • Secretory Vesicles / metabolism


  • Fluorescent Dyes
  • Insulin
  • Potassium Chloride
  • Glucose