BDNF modulates glycine uptake in hippocampal synaptosomes by decreasing membrane insertion of glycine transporter 2.
Glycine transporter 2 (GlyT2) is localized in the nerve terminals of glycinergic neurons, promoting glycine uptake and ensuring the refilling of glycinergic vesicles. Brain-derived neurotrophic factor (BDNF) activates its high affinity TrkB receptors, which occur in two isoforms, full length (TrkB-FL) and truncated (TrkB-T1/T2). After BDNF binding to TrkB receptor, several intracellular cascades are triggered, specifically PLC, Akt and MAPK signalling pathways. We herein show that BDNF decreases [(3)H]glycine uptake mediated by GlyT2 in isolated nerve endings (synaptosomes) obtained from rat hippocampus, by reducing the maximum velocity (Vmax) of transport while not influencing the transporter affinity constant (Km) for glycine. Western Blot analysis detected both TrkB receptor isoforms in the synaptosomes but the BDNF effect seems to be mediated by TrkB-FL since: 1) the tyrosine kinase inhibitor, k252a, prevented the effect of BDNF, and 2) the effect of BDNF was lost in the presence of specific inhibitors of TrkB signalling pathways, namely U73122, LY294002 and U0126 (inhibitors of PLC, Akt and MAPK pathways, respectively). Monensin, a transporter recycling inhibitor, prevented the BDNF action upon glycine uptake, suggesting that BDNF reduces GlyT2 insertion in the plasma membrane. It is concluded that BDNF effect upon glycine uptake into glycinergic nerve terminals requires the activation of the TrkB-FL receptor and its canonical signalling pathways and occurs by inhibiting GlyT2 membrane incorporation.
Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal; Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal.
Rita I
RI
Aroeira
Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal; Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal.
Sandra H
SH
Vaz
Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal; Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal.
Ana M
AM
Sebastião
Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal; Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Av. Prof. Egas Moniz, 1649-028, Lisboa, Portugal. Electronic address: cvalentecastro@medicina.ulisboa.pt.
Cláudia A
CA
Valente
D000818:Animals; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005998:Glycine / Q000037:antagonists & inhibitors / Q000378:metabolism*; D050487:Glycine Plasma Membrane Transport Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*
D019208:Brain-Derived Neurotrophic Factor; D050487:Glycine Plasma Membrane Transport Proteins; C493269:Slc6a5 protein, rat; D005998:Glycine
Glycine transporter 2; Hippocampus; Neurotrophic factors; Synaptosomes