Thrombin-stimulated glutamate uptake in human platelets is predominantly mediated by the glial glutamate transporter EAAT2
| Autor: | Henk A. Spierenburg, P.N.E. de Graan, Frank Kupper, G. Van Willigen, Ineke W. M. Bos, O. van Nieuwenhuizen, G. Hoogland |
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| Přispěvatelé: | Science Communication |
| Rok vydání: | 2005 |
| Předmět: |
Blood Platelets
Platelet Aggregation Blotting Western Glutamate decarboxylase Glutamic Acid Cell Separation In Vitro Techniques Biology Cytoplasmic Granules Phosphatidylinositol 3-Kinases Cellular and Molecular Neuroscience Glutamate aspartate transporter Humans Cell Shape Protein Kinase C Metabotropic glutamate receptor 5 Sodium Thrombin Glutamate receptor Metabotropic glutamate receptor 6 Cell Biology Adenosine Diphosphate Kinetics Excitatory Amino Acid Transporter 2 Biochemistry Glutamate dehydrogenase 1 Metabotropic glutamate receptor Chromatography Gel biology.protein NMDA receptor Neuroglia |
| Zdroj: | Kupper, F 2005, ' Thrombin-stimulated glutamate uptake in human platelets is predominantly mediated by the glial glutamate transporter EAAT2 ', Neurochemistry International, vol. 47, no. 7 . https://doi.org/10.1016/j.neuint.2005.06.006 Neurochemistry International, 47(7). Elsevier Limited |
| ISSN: | 0197-0186 |
| Popis: | Glutamate toxicity has been implicated in the pathogenesis of various neurological diseases. Glial glutamate transporters play a key role in the regulation of extracellular glutamate levels in the brain by removing glutamate from the extracellular fluid. Since human blood platelets possess an active glutamate uptake system, they have been used as a peripheral model of glutamate transport in the central nervous system (CNS). The present study is aimed at identifying the glutamate transporter on blood platelets, and to asses the influence of platelet activation on glutamate uptake. Platelets from healthy donors showed Na+-dependent glutamate uptake (Km, 3.5+/-0.9 microM; Vmax, 2.8+/-0.2 pmol glutamate/75 x 10(6)platelets/30 min), which could be blocked dose-dependently by the EAAT specific inhibitors DL-threo-E-benzyloxyaspartate (TBOA), L-trans-pyrrolidine-2,4-dicarboxylic acid (tPDC) and high concentrations of the EAAT2 inhibitor dihydrokainate (DHK). Analysis of platelet homogenates on Western blots showed EAAT2 as the predominant glutamate transporter. Platelet activation by thrombin caused an increase in glutamate uptake, which could be inhibited by TBOA and the EAAT2 inhibitor DHK. Kinetic analysis showed recruitment of new transporters to the membrane. Indeed, Western blot analysis of subcellular fractions revealed that alpha-granules, which fuse with the membrane upon thrombin stimulation, contained significant EAAT2 immunoreactivity. Inhibition of the second messengers involved in alpha-granule secretion (protein kinase C, phosphatidylinositol-3-kinase) inhibited thrombin-stimulated uptake, but not basal uptake. These data show that the glial EAAT2 is the predominant glutamate transporter on blood platelets and suggest, that thrombin increases glutamate uptake capacity by recruiting new transporters (EAAT2) from alpha-granules. |
| Databáze: | OpenAIRE |
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