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P4‐257: Activation glycogen synthase kinase 3 retards exocytosis
Author(s) -
Zhu Ling-Qiang,
Liu Dan,
Wang Jian-Zhi
Publication year - 2008
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1016/j.jalz.2008.05.2326
Subject(s) - exocytosis , microbiology and biotechnology , biology , synaptosome , gsk 3 , syntaxin , förster resonance energy transfer , chemistry , biochemistry , kinase , secretion , fluorescence , physics , quantum mechanics , membrane
Background: Glycogen synthase kinase 3 (GSK-3) plays an important role in the pathogenesis of Alzheimer’s disease. Activation GSK-3 induces tau hyperphosphorylation, memory retention deficits and Long-term Potentiation (LTP) inhibition in vivo. Presynaptic vesicle exocytosis is important to neurotransmitter release, which in turn affects LTP. Methods: Wild type and dominant negative GSK-3 plasmids were transfected to regulate the activity of GSK-3. FM4-64 releasing by time lapse with confocal microscopy in primary hippocampus neurons and pHluorin-VAMP2 fluorescence detecting in Neuro2A cells by total internal reflecting fluorescence microscopy (TIRFM) were used to evaluate the exocytosis stimulated by 90mM K , Fluorescence resonance energy transfer (FRET) and immunoprecipitation in synaptosome were used to assay the formation of SNARE complex, which play an important role in exocytosis. GSK-3 activity was determined by P-labelling assay. Results: Overexpressing wild type GSK-3 not only retards the velocity of FM4-64 releasing in the axon terminal of primary hippocampus neurons, but also delays the fluorescence intensity increasing of pHluorin-VAMP2 in N2a cells. FRET analysis with ECFP-VAMP2 and EYFP-Syp I shows that overexpressing wtGSK-3 inhibits the dissociation of VAMP2 and Syp I in both N2a cells and hippocampus neurons, especially in the terminal of axon. Immunoprecipitation with anti-VAMP2 in synaptosome reveals the less amount of SNAP25 and syntaxin I (t-SNARE) when GSK-3 is activated. Conclusions: GSK-3 activation inhibits exocytosis through retards SNARE complex formation, which plays an important role in synaptic dysfunction in Alzheimer’s disease.