title: Multivesicular exocytosis in rat pancreatic beta cells.
abstract: OBJECTIVE
  To establish the occurrence, modulation and functional significance of compound exocytosis in insulin-secreting beta cells.
  
  
  METHODS
  Exocytosis was monitored in rat beta cells by electrophysiological, biochemical and optical methods. The functional assays were complemented by three-dimensional reconstruction of confocal imaging, transmission and block face scanning electron microscopy to obtain ultrastructural evidence of compound exocytosis.
  
  
  RESULTS
  Compound exocytosis contributed marginally (<5% of events) to exocytosis elicited by glucose/membrane depolarisation alone. However, in beta cells stimulated by a combination of glucose and the muscarinic agonist carbachol, 15-20% of the release events were due to multivesicular exocytosis, but the frequency of exocytosis was not affected. The optical measurements suggest that carbachol should stimulate insulin secretion by ∼40%, similar to the observed enhancement of glucose-induced insulin secretion. The effects of carbachol were mimicked by elevating [Ca(2+)](i) from 0.2 to 2 μmol/l Ca(2+). Two-photon sulforhodamine imaging revealed exocytotic events about fivefold larger than single vesicles and that these structures, once formed, could persist for tens of seconds. Cells exposed to carbachol for 30 s contained long (1-2 μm) serpentine-like membrane structures adjacent to the plasma membrane. Three-dimensional electron microscopy confirmed the existence of fused multigranular aggregates within the beta cell, the frequency of which increased about fourfold in response to stimulation with carbachol.
  
  
  CONCLUSIONS
  Although contributing marginally to glucose-induced insulin secretion, compound exocytosis becomes quantitatively significant under conditions associated with global elevation of cytoplasmic calcium. These findings suggest that compound exocytosis is a major contributor to the augmentation of glucose-induced insulin secretion by muscarinic receptor activation.
authors.affiliation: Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Oxford, UK.
authors.forename: M B
authors.initials: MB
authors.lastname: Hoppa
authors.forename: E
authors.initials: E
authors.lastname: Jones
authors.forename: J
authors.initials: J
authors.lastname: Karanauskaite
authors.forename: R
authors.initials: R
authors.lastname: Ramracheya
authors.forename: M
authors.initials: M
authors.lastname: Braun
authors.forename: S C
authors.initials: SC
authors.lastname: Collins
authors.forename: Q
authors.initials: Q
authors.lastname: Zhang
authors.forename: A
authors.initials: A
authors.lastname: Clark
authors.forename: L
authors.initials: L
authors.lastname: Eliasson
authors.forename: C
authors.initials: C
authors.lastname: Genoud
authors.forename: P E
authors.initials: PE
authors.lastname: Macdonald
authors.forename: A G
authors.initials: AG
authors.lastname: Monteith
authors.forename: S
authors.initials: S
authors.lastname: Barg
authors.forename: J
authors.initials: J
authors.lastname: Galvanovskis
authors.forename: P
authors.initials: P
authors.lastname: Rorsman
mesh_terms: D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002478:Cells, Cultured; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism*; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D022142:Secretory Vesicles / Q000187:drug effects / Q000378:metabolism*
chemical_list: D007328:Insulin; D002118:Calcium