title: Caffeine potentiates the release of GABA mediated by NMDA receptor activation: Involvement of A1 adenosine receptors.
abstract: Caffeine, a methylated derivative of xanthine and widely consumed psychoactive substance, acts in several targets in the nervous system. We investigated its role in retinal explants of chick embryo analyzing the role of purinergic receptors in [(3)H]-GABA release induced by d-aspartate (d-asp). d-Asp increases GABA-release 4.5-fold when compared to basal levels from 13-day-old chick embryo retinal explants. Caffeine 500μM elevated d-asp-induced GABA release in 60%. The release was inhibited in the presence of NNC-711, a GABA transporter-1 (GAT-1) blocker or by MK-801, an N-methyl-d-aspartate receptor (NMDAR) antagonist. Caffeine did not modify [(3)H]-GABA uptake carried out for 5, 10, 30 and 60min and did not increase the release of d-asp or glutamate at basal or stimulated conditions. The caffeine effect was mimicked by the adenosine A1 receptor antagonist DPCPX and by the adenylyl cyclase (AC) activator forskolin. It was also blocked by the protein kinase A (PKA) inhibitor H-89, tyrosine kinase inhibitor genistein or by the src family kinase (SFK) inhibitor PP1. Forskolin-stimulated cyclic adenosine monophosphate (cAMP) levels were reduced in the presence of the A1 receptor agonist CHA. Western blot analysis revealed that 500μM caffeine increased phosphoGluN2B expression levels in approximately 60% when compared to total GluN2B levels in embryonic E13 retina. The GluN2B subunit-containing NMDAR antagonist ifenprodil inhibited the caffeine effect. Our results suggest that caffeine potentiates d-asp-induced GABA release, which is mediated by GAT-1, via inhibition of adenosine A1 receptor and activation of the PKA pathway. Regulation of NMDAR by phosphorylation of GluN2B subunit by a SFK may also be involved in the effect promoted by caffeine.
authors.affiliation: Laboratório de Neurofarmacologia, Departamento de Fisiologia e Farmacologia, Programa de Pós-graduação em Neurociências, Universidade Federal Fluminense, Niterói, Brazil. Electronic address: ferreira@vm.uff.br.
authors.forename: D D P
authors.initials: DD
authors.lastname: Ferreira
authors.affiliation: Laboratório de Neuroquímica, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal de Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: stutz@biof.ufrj.br.
authors.forename: B
authors.initials: B
authors.lastname: Stutz
authors.affiliation: Laboratório de Neuroquímica, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal de Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: fgmello@biof.ufrj.br.
authors.forename: F G
authors.initials: FG
authors.lastname: de Mello
authors.affiliation: Laboratório de Neuroquímica, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal de Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: ramreis@biof.ufrj.br.
authors.forename: R A M
authors.initials: RA
authors.lastname: Reis
authors.affiliation: Laboratório de Neurofarmacologia, Departamento de Fisiologia e Farmacologia, Programa de Pós-graduação em Neurociências, Universidade Federal Fluminense, Niterói, Brazil. Electronic address: kubrusly@vm.uff.br.
authors.forename: R C C
authors.initials: RC
authors.lastname: Kubrusly
mesh_terms: D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D000697:Central Nervous System Stimulants / Q000494:pharmacology*; D002642:Chick Embryo; D017868:Cyclic AMP-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D026603:D-Aspartic Acid / Q000494:pharmacology*; D050485:GABA Plasma Membrane Transport Proteins / Q000187:drug effects / Q000378:metabolism*; D043682:Receptor, Adenosine A1 / Q000187:drug effects / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000187:drug effects / Q000378:metabolism*; D012160:Retina / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D005680:gamma-Aminobutyric Acid / Q000187:drug effects / Q000378:metabolism*
chemical_list: D000697:Central Nervous System Stimulants; D050485:GABA Plasma Membrane Transport Proteins; C121001:NR2B NMDA receptor; D043682:Receptor, Adenosine A1; D016194:Receptors, N-Methyl-D-Aspartate; D002110:Caffeine; D026603:D-Aspartic Acid; D005680:gamma-Aminobutyric Acid; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: GABA; NMDA receptor; adenosine receptors; caffeine; retina