Functional plasticity of GAT-3 in avian Müller cells is regulated by neurons via a glutamatergic input

• Published in: Neurochemistry international Vol. 82; pp. 42 - 51
• Main Authors: Schitine, Clarissa S., Mendez-Flores, Orquidia G., Santos, Luis E., Ornelas, Isis, Calaza, Karin C., Pérez-Toledo, Karla, López-Bayghen, Esther, Ortega, Arturo, Gardino, Patrícia F., de Mello, Fernando G., Reis, Ricardo A.M.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.03.2015
• Subjects: Animals; Biological Transport, Active; Biotinylation; Calcium - analysis; Cell Membrane - metabolism; Cells, Cultured; Chick Embryo; Chickens; Culture Media, Conditioned; Ependymoglial Cells - drug effects; Ependymoglial Cells - physiology; GABA Plasma Membrane Transport Proteins - genetics; GABA Plasma Membrane Transport Proteins - physiology; GABA transporters; gamma-Aminobutyric Acid - metabolism; Gene Expression Profiling; Glutamate receptors; Glutamic Acid - pharmacology; Glutamic Acid - physiology; Kainic Acid - pharmacology; Müller glia; N-Methylaspartate - administration & dosage; N-Methylaspartate - pharmacology; Protein Kinase C - antagonists & inhibitors; Protein Kinase C - physiology; Protein Kinase Inhibitors - pharmacology; Retina - growth & development; RNA, Messenger - biosynthesis; RNA, Messenger - genetics; Tetradecanoylphorbol Acetate - pharmacology; Glutamate receptors; Müller glia; GABA transporters
• DOI: 10.1016/j.neuint.2015.02.004
• ISSN: 0197-0186

•Glutamate decreases [3H]-GABA uptake in Müller glia via ionotropic receptors.•Glutamate increases intracellular Ca2+without causing toxicity to Müller cells.•GAT-1 and GAT-3 mRNA levels are also decreased by glutamate.•Inhibition on GAT-3 activity is not reverted by PKC inhibitors.•Intra-vitreous injection of NMDA results in GAT-3 expression in Müller cells. GABA (γ-amino butyric acid) is the major inhibitory transmitter in the central nervous system and its action is terminated by specific transporters (GAT), found in neurons and glial cells. We have previously described that GAT-3 is responsible for GABA uptake activity in cultured avian Müller cells and that it operates in a Na+ and Cl− dependent manner. Here we show that glutamate decreases [3H] GABA uptake in purified cultured glial cells up to 50%, without causing cell death. This effect is mediated by ionotropic glutamatergic receptors. Glutamate inhibition on GABA uptake is not reverted by inhibitors of protein kinase C or modified by agents that modulate cyclic AMP/PKA. Biotinylation experiments demonstrate that this reduction in GABA uptake correlates with a decrease in GAT-3 plasma membrane levels. Interestingly, both GAT-1 and GAT-3 mRNA levels are also decreased by glutamate. Conditioned media (CM) prepared from retinal neurons could also decrease GABA influx, and glutamate receptor antagonists (MK-801 + CNQX) were able to prevent this effect. However, glutamate levels in CM were not different from those found in fresh media, indicating that a glutamatergic co-agonist or modulator could be regulating GABA uptake by Müller cells in this scenario. In the whole avian retina, GAT-3 is present from embryonic day 5 (E5) increasing up to the end of embryonic development and post-hatch period exclusively in neuronal layers. However, this pattern may change in pathological conditions, which drive GAT-3 expression in Müller cells. Our data suggest that in purified cultures and upon extensive neuronal lesion in vivo, shown as a Brn3a reduced neuronal cells and an GFAP increased gliosis, Müller glia may change its capacity to take up GABA due to GAT-3 up regulation and suggests a regulatory interplay mediated by glutamate between neurons and glial cells in this process.