Glutamate enhances the surface distribution and release of Munc18 in cerebral cortical neurons.

Neuroscience Bulletin
Ping WanCui-Qing Zhu

Abstract

Munc18 is considered as an intracellular protein that plays an important role in exocytosis of neurotransmitters. Previous studies have demonstrated the presence of autoantibodies against Munc18 in a subgroup of Rasmussen's encephalitis patients. However, the machinery of Munc18 autoimmunity is still elusive. The present study was aimed to investigate Munc18 release from primary cultured neurons, Munc18 distribution on the outer plasma membrane of neurons, and the neurotoxicity of Munc18 antibody. The cerebral cortical neurons from embryonic day 17 Sprague-Dawley rats were prepared and cultured in neurobasal medium. The proteins in culture medium were precipitated with 10 % trichloroacetic acid, and analyzed by immunoblotting. The proteins on neuronal surface were biotinylated with EZ-Link-sulfo-NHS-LC-Biotin, and collected with avidin-conjugated agarose beads followed by immunoblotting analysis. For cell surface immunofluorescent staining, the living neurons were labeled with anti-Munc18 antibody at 4 degrees C. Neuronal injury was assessed by lactate dehydrogenase(LDH) release. Munc18 was detected in culture medium by immunoblotting analysis. After treatment with 50 micromol/L glutamate for 1 h, Munc18 content in medium was i...Continue Reading

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Citations

Dec 17, 2010·Journal of Bioenergetics and Biomembranes·Florence Molinari

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