Interaction of SNARE Complexes with P/Q-type Calcium Channels in Rat Cerebellar Synaptosomes (*)
- Nicole Martin-Moutot(§),
- Nathalie Charvin,
- Christian Leveque,
- Kazuki Sato(1),
- Tei-ich Nishiki(1),
- Shunji Kozaki(2),
- Masami Takahashi(1) and
- Michael Seagar
- From the (1)From INSERM, Unité 374, Institut Jean Roche, Faculté de Médecine Secteur Nord, Boulevard Pierre Dramard, 13916 Marseille Cedex 20, France, Mitsubishi Kasei Institute of Life Science, Machida, Tokyo 194, Japan, and the
- (2)Department of Veterinary Science, University of Osaka Prefecture, Osaka 593, Japan
- § To whom correspondence should be addressed. Tel.: 33-91698834; Fax: 33-91090506.
Abstract
P- and Q-type calcium channels, which trigger rapid neurotransmitter release at many mammalian synapses, are blocked by 
-conotoxin MVIIC. 
I-
-Conotoxin MVIIC binding to rat cerebellar synaptosomes was not displaced by 
-conotoxins GVIA or MVIIA (K
> 1 μM), which are selective for N-type calcium channels. Solubilized 
I-
-conotoxin MVIIC receptors were specifically recognized by antibodies directed against α
A calcium channel subunits, proteins known to constitute a pore with P/Q-like channel properties. Antibodies against syntaxin
1, SNAP 25, and VAMP 2 (synaptobrevin) each immunoprecipitated a similar fraction (20-40%) of 
-conotoxin MVIIC receptors. Immunoprecipitation was not additive, suggesting that heterotrimeric (SNARE) complexes containing
these three proteins interact with P/Q-type calcium channels. Immobilized monoclonal anti-syntaxin antibodies retained α
A calcium channel subunits of 220, 180 and 160 kDa monitored by immunoblotting with site directed antibodies. Synaptotagmin
was detected in channel-associated complexes, but not synaptophysin, Rab 3A nor rat cysteine string protein. Trimeric SNARE
complexes are implicated in calcium-dependent exocytosis, a process thought to be regulated by synaptotagmin. Our results
indicate that these proteins interact with P/Q-type calcium channels, which may optimize their location within domains of
calcium influx.
Footnotes
-
↵* This work was supported by CNRS, the Association Française contre les Myopathies, a grant-in-aid for scientific research on priority areas (“Fundamental Development of Neural Circuits”) from the Japanese Ministry of Education, Science, Sports and Culture, and a joint project grant from INSERM and the Japanese Society for the Promotion of Science. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵1 The abbreviations used are:
GVIA-
-conotoxin GVIA
MVIIA-
-conotoxin MVIIA
AgaIVA-
-agatoxin IVA
MVIIC-
-conotoxin MVIIC
- mAb
-
monoclonal antibody
- CHAPS
-
3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid.
-
- Received December 29, 1995.
- Revision received January 25, 1996.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
