title: Dual regulation of Ca2+-dependent glutamate release from astrocytes: vesicular glutamate transporters and cytosolic glutamate levels.
abstract: Vesicular glutamate transporters (VGLUTs) are responsible for vesicular glutamate storage and exocytotic glutamate release in neurons and astrocytes. Here, we selectively and efficiently overexpressed individual VGLUT proteins (VGLUT1, 2, or 3) in solitary astrocytes and studied their effects on mechanical stimulation-induced Ca2+-dependent glutamate release. Neither VGLUT1 nor VGLUT2 overexpression changed the amount of glutamate release, whereas overexpression of VGLUT3 significantly enhanced Ca2+-dependent glutamate release from astrocytes. None of the VGLUT overexpression affected mechanically induced intracellular Ca2+ increase. Inhibition of glutamine synthetase activity by L-methionine sulfoximine in astrocytes, which leads to increased cytosolic glutamate concentration, greatly increased their mechanically induced Ca2+-dependent glutamate release, without affecting intracellular Ca2+ dynamics. Taken together, these data indicate that both VGLUT3 and the cytosolic concentration of glutamate are key limiting factors in regulating the Ca2+-dependent release of glutamate from astrocytes.
authors.affiliation: National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Yingchun
authors.initials: Y
authors.lastname: Ni
authors.forename: Vladimir
authors.initials: V
authors.lastname: Parpura
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D005974:Glutamate-Ammonia Ligase / Q000037:antagonists & inhibitors; D018698:Glutamic Acid / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D008717:Methionine Sulfoximine / Q000494:pharmacology; D010812:Physical Stimulation; D034622:RNA Interference; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection; D050598:Vesicular Glutamate Transport Protein 1 / Q000235:genetics / Q000378:metabolism; D050599:Vesicular Glutamate Transport Protein 2 / Q000235:genetics / Q000378:metabolism; D050496:Vesicular Glutamate Transport Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: C493475:Slc17a6 protein, rat; C493466:Slc17a7 protein, rat; C482103:Slc17a8 protein, rat; D050598:Vesicular Glutamate Transport Protein 1; D050599:Vesicular Glutamate Transport Protein 2; D050496:Vesicular Glutamate Transport Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D008717:Methionine Sulfoximine; D018698:Glutamic Acid; D005974:Glutamate-Ammonia Ligase; D002118:Calcium