title: Endocytosis of secretory granules in mouse pancreatic beta-cells evoked by transient elevation of cytosolic calcium.
abstract: 1. To investigate the mechanisms regulating the reuptake of secretory granule membranes following regulated exocytosis, we have monitored changes in cell capacitance in single pancreatic beta-cells. 2. Membrane retrieval (endocytosis) occurred both in a continuous manner and in abrupt steps, corresponding to the simultaneous retrieval of 50-100 granules. The large endocytotic steps were associated with a conductance change of about 1 nS which we attribute to the formation of a fission pore with a pore radius of approximately 1 nm. 3. In some cells, we observed large amplitude capacitance fluctuations, suggesting that aggregates of granules are connected to the plasma membrane by a single pore and are subsequently retrieved as a single unit. 4. Endocytosis was evoked by elevation of cytosolic [Ca2+]i, but once initiated, a sustained increase in [Ca2+]i was not required for endocytosis to continue. 5. The [Ca2+]i dependence of exo- and endocytosis was studied by photorelease of Ca2+ from the 'caged' precursor Ca(2+)-nitrophenyl-EGTA (Ca(2+)-NP-EGTA). Both exo- and endocytosis were initiated at between 0.5 and 2 microM Cai(2+). The rate of endocytosis saturated above 2 microM Cai(2+), whereas exocytosis continued to increase up to 4 microM Cai(2+). The maximum rate of endocytosis was < 25% of that of exocytosis. 6. Unlike exocytosis, endocytosis proceeded equally well in the presence or absence of Mg-ATP. 7. Our data indicate that in the pancreatic beta-cell, exocytosis and endocytosis are regulated by different mechanisms.
authors.affiliation: Department of Physiology and Pharmacology, University Laboratory of Physiology, Oxford, UK.
authors.forename: L
authors.initials: L
authors.lastname: Eliasson
authors.forename: P
authors.initials: P
authors.lastname: Proks
authors.forename: C
authors.initials: C
authors.lastname: Ammälä
authors.forename: F M
authors.initials: FM
authors.lastname: Ashcroft
authors.forename: K
authors.initials: K
authors.lastname: Bokvist
authors.forename: E
authors.initials: E
authors.lastname: Renström
authors.forename: P
authors.initials: P
authors.lastname: Rorsman
authors.forename: P A
authors.initials: PA
authors.lastname: Smith
mesh_terms: D000255:Adenosine Triphosphate / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D003594:Cytoplasmic Granules / Q000378:metabolism / Q000502:physiology*; D003600:Cytosol / Q000378:metabolism / Q000502:physiology*; D004594:Electrophysiology; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D066298:In Vitro Techniques; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D007700:Kinetics; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D017551:Microdialysis; D018408:Patch-Clamp Techniques; D013050:Spectrometry, Fluorescence
chemical_list: D000255:Adenosine Triphosphate; D002118:Calcium