Imaging analysis reveals mechanistic differences between first- and second-phase insulin exocytosis.
The mechanism of glucose-induced biphasic insulin release is unknown. We used total internal reflection fluorescence (TIRF) imaging analysis to reveal the process of first- and second-phase insulin exocytosis in pancreatic beta cells. This analysis showed that previously docked insulin granules fused at the site of syntaxin (Synt)1A clusters during the first phase; however, the newcomers fused during the second phase external to the Synt1A clusters. To reveal the function of Synt1A in phasic insulin exocytosis, we generated Synt1A-knockout (Synt1A(-/-)) mice. Synt1A(-/-) beta cells showed fewer previously docked granules with no fusion during the first phase; second-phase fusion from newcomers was preserved. Rescue experiments restoring Synt1A expression demonstrated restoration of granule docking status and fusion events. Inhibition of other syntaxins, Synt3 and Synt4, did not affect second-phase insulin exocytosis. We conclude that the first phase is Synt1A dependent but the second phase is not. This indicates that the two phases of insulin exocytosis differ spatially and mechanistically.
Ohara-Imaizumi
Mica
M
Department of Biochemistry, Kyorin University School of Medicine, Mitaka, Tokyo, Japan.
Fujiwara
Tomonori
T
Nakamichi
Yoko
Y
Okamura
Tadashi
T
Akimoto
Yoshihiro
Y
Kawai
Junko
J
Matsushima
Satsuki
S
Kawakami
Hayato
H
Watanabe
Takashi
T
Akagawa
Kimio
K
Nagamatsu
Shinya
S
D000818:Animals; D002478:Cells, Cultured; D005089:Exocytosis / Q000502:physiology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence
D007328:Insulin