Transfection analysis of functional roles of complexin I and II in the exocytosis of two different types of secretory vesicles

Makoto Itakura, Hidemi Misawa, Mariko Sekiguchi, Seiichi Takahashi, Masami Takahashi

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

Classical neurotransmitters such as γ-aminobutyric acid and glutamate are released from synaptic nerve terminals by exocytosis of synaptic vesicles. PC12 cells also have SSVs capable of storing acetylcholine (ACh). A novel method to examine the effect of transient transfection of any gene of interest on the exocytosis of SSVs was developed. The transfection of choline acetyltransferase (ChAT) into PC12 cells which have lost ACh synthesizing activity resulted in the accumulation of a substantial amount of ACh. Synthesized ACh was released in Ca2+-dependent manner. Release was thought to occur by an exocytosis of SSVs because: (1) release was abolished by treating the cells with vesamicol, a specific inhibitor of the vesicular ACh transporter (VAChT) localizing specifically in SSVs; and (2) the release was further increased by cotransfecting rat VAChT with the ChAT. By means of this method, we showed that overexpression of complexin I or II with ChAT markedly suppressed high-K+-dependent ACh release of SSVs.

Original languageEnglish
Pages (from-to)691-696
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume265
Issue number3
DOIs
Publication statusPublished - 1999 Nov 30
Externally publishedYes

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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