title: BDNF, via truncated TrkB receptor, modulates GlyT1 and GlyT2 in astrocytes.
abstract: Glycine transporters (GlyT), GlyT1 and GlyT2, are responsible for the termination of glycine-mediated synaptic activity through removal of neurotransmitter from synaptic cleft. Brain-derived neurotrophic factor (BDNF) activates its high affinity tropomyosin-related kinase (Trk) receptors, namely TrkB, which includes full length (TrkB-FL) and truncated (TrkB-T) isoforms. In this article we evaluated the influence of BDNF upon the activity of glycine transporters in astrocytes. We report that BDNF decreases GlyT1- and GlyT2- mediated [(3) H]glycine transport in primary cultures of astrocytes from rat cerebral cortex. BDNF decreased Vmax but not Km values of transport, which suggests that BDNF induces transporter internalization. Accordingly, dynasore, an inhibitor of dynamin/clathrin-dependent endocytosis, prevented the influence of BDNF upon GlyT-mediated transport. While quantifying mRNA and protein levels, we detected a predominance of truncated isoforms over the TrkB-FL receptor. The effect of BDNF was not abolished by specific inhibitors of PLCγ, PI3K and MAPK, indicating that it did not occur through TrkB-FL canonical pathways. However, BDNF action was lost in the presence of a Rho family-specific blocker (toxin B), a signaling pathway that has been associated to TrkB-T1. Furthermore, the effect of BDNF was abolished upon TrkB-T knockdown in astrocytes by RNA interference. Immunofluorescence assays confirmed an increased GlyT expression in endosomes upon BDNF incubation, which was prevented in the presence of either dynasore or toxin B. We conclude that BDNF, acting on TrkB-T1 receptors, inhibits glycine uptake in astrocytes by promoting GlyT internalization through a Rho-GTPase activity dependent mechanism.
authors.affiliation: Faculty of Medicine, Institute of Pharmacology and Neurosciences, and Unit of Neurosciences, Institute of Molecular Medicine, University of Lisbon, Av. Prof. Egas Moniz, Lisbon, Portugal.
authors.forename: Rita I
authors.initials: RI
authors.lastname: Aroeira
authors.affiliation: Faculty of Medicine, Institute of Pharmacology and Neurosciences, and Unit of Neurosciences, Institute of Molecular Medicine, University of Lisbon, Av. Prof. Egas Moniz, Lisbon, Portugal.
authors.forename: Ana M
authors.initials: AM
authors.lastname: Sebastião
authors.affiliation: Faculty of Medicine, Institute of Pharmacology and Neurosciences, and Unit of Neurosciences, Institute of Molecular Medicine, University of Lisbon, Av. Prof. Egas Moniz, Lisbon, Portugal.
authors.forename: Cláudia A
authors.initials: CA
authors.lastname: Valente
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D019208:Brain-Derived Neurotrophic Factor / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D005998:Glycine / Q000378:metabolism; D050487:Glycine Plasma Membrane Transport Proteins / Q000378:metabolism*; D006835:Hydrazones / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D020813:Receptor, trkB / Q000378:metabolism*; D014316:Tritium; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D019208:Brain-Derived Neurotrophic Factor; D050487:Glycine Plasma Membrane Transport Proteins; D006835:Hydrazones; C511794:N'-(3,4-dihydroxybenzylidene)-3-hydroxy-2-naphthahydrazide; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; C493269:Slc6a5 protein, rat; C493266:Slc6a9 protein, rat; D014316:Tritium; D020813:Receptor, trkB; D054460:rho-Associated Kinases; D002118:Calcium; D005998:Glycine
keywords: RNA interference; brain; glial cells; glycine transporter 1; glycine transporter 2; neurotrophins