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Overexpression of rat neuronal calcium sensor‐1 in rodent NG108‐15 cells enhances synapse formation and transmission
Author(s) -
Chen XiaoLiang,
Zhong ZhenGuo,
Yokoyama Shigeru,
Bark Christina,
Meister Björn,
Berggren PerOlof,
Roder John,
Higashida Haruhiro,
Jeromin Andreas
Publication year - 2001
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 - 10.1111/j.1469-7793.2001.0649e.x
Subject(s) - neurite , transfection , green fluorescent protein , microbiology and biotechnology , golgi apparatus , synapse , myogenesis , cell culture , chemistry , synaptic vesicle , biology , biophysics , myocyte , vesicle , biochemistry , in vitro , endoplasmic reticulum , neuroscience , membrane , gene , genetics
1 The role of rat neuronal calcium sensor‐1 (NCS‐1), a Ca 2+ ‐binding protein, in synapse formation and transmitter release was examined in mouse neuroblastoma × rat glioma hybrid NG108‐15 cells in culture. 2 Wild‐type NG108‐15 cells expressed rodent NCS‐1. Endogenous NCS‐1 was partially co‐localized with the synaptic protein SNAP‐25 at the plasma membrane in both cell bodies and processes, but not with the Golgi marker β‐COP, an individual coat subunit of the coatomer complex present on Golgi‐derived vesicles. 3 In NG108‐15 cells co‐cultured with rat myotubes, partial co‐localization of SNAP‐25 and NCS‐1 was observed at the plasma membrane of neurites and growth cones, some of which had synaptic contacts to muscle cells. 4 Transient co‐transfection of the rat NCS‐1 cDNA and green fluorescent protein (GFP) resulted in NCS‐1 overexpression in about 30 % of the cells as determined by fluorescence microscopy. 5 The rate of functional synapse formation with co‐cultured rat myotubes increased 2‐fold as determined by the presence of miniature endplate potentials (MEPPs) in NCS‐1‐overexpressing NG108‐15 cells compared to non‐ and mock‐transfected cells. 6 The number of neurites per cell, branches per neurite and length of neurites was slightly less in cells that were either transiently transfected (GFP‐NCS‐1‐fluorescence positive) or stably transformed with NCS‐1 compared to GFP‐NCS‐1‐negative, non‐transfected or mock‐transfected NG108‐15 cells. 7 The number of action potentials that elicited endplate potentials increased in NG108‐15 cells stably transformed with rat NCS‐1. The mean number of quanta per impulse ( m ) increased 5‐fold. 8 These results show that NCS‐1 functions to facilitate synapse formation, probably because of the increased quantal content of evoked acetylcholine release.