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Two cAMP‐dependent pathways differentially regulate exocytosis of large dense‐core and small vesicles in mouse β‐cells
Author(s) -
Hatakeyama Hiroyasu,
Takahashi Noriko,
Kishimoto Takuya,
Nemoto Tomomi,
Kasai Haruo
Publication year - 2007
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2007.135228
Subject(s) - exocytosis , forskolin , vesicle , munc 18 , microbiology and biotechnology , stimulation , chemistry , protein kinase a , biophysics , secretion , biology , synaptic vesicle , endocrinology , biochemistry , kinase , membrane
It has been reported that cAMP regulates Ca 2 + ‐dependent exocytosis via protein kinase A (PKA) and exchange proteins directly activated by cAMP (Epac) in neurons and secretory cells. It has, however, never been clarified how regulation of Ca 2 + ‐dependent exocytosis by cAMP differs depending on the involvement of PKA and Epac, and depending on two types of secretory vesicles, large dense‐core vesicles (LVs) and small vesicles (SVs). In this study, we have directly visualized Ca 2 + ‐dependent exocytosis of both LVs and SVs with two‐photon imaging in mouse pancreatic β‐cells. We found that marked exocytosis of SVs occurred with a time constant of 0.3 s, more than three times as fast as LV exocytosis, on stimulation by photolysis of a caged‐Ca 2 + compound. The diameter of SVs was identified as ∼80 nm with two‐photon imaging, which was confirmed by electron‐microscopic investigation with photoconversion of diaminobenzidine. Calcium‐dependent exocytosis of SVs was potentiated by the cAMP‐elevating agent forskolin, and the potentiating effect was unaffected by antagonists of PKA and was mimicked by the Epac‐selective agonist 8‐(4‐chlorophenylthio)‐2′‐ O ‐methyl cAMP, unlike that on LVs. Moreover, high‐glucose stimulation induced massive exocytosis of SVs in addition to LVs, and photolysis of caged cAMP during glucose stimulation caused potentiation of exocytosis with little delay for SVs but with a latency of 5 s for LVs. Thus, Epac and PKA selectively regulate exocytosis of SVs and LVs, respectively, in β‐cells, and Epac can regulate exocytosis more rapidly than PKA.