title: Activation of glutamate receptors promotes a calcium-dependent and transporter-mediated release of purines in cultured avian retinal cells: possible involvement of calcium/calmodulin-dependent protein kinase II.
abstract: Calcium-dependent release of purines was previously demonstrated in cultures of chick retinal cells stimulated with high potassium concentrations but there is no evidence for an exocytotic mechanism of adenosine release from presynaptic terminals. Here we show that activation of NMDA or alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA)/kainate glutamate ionotropic receptors promotes a two- to three-fold increase in the release of purines from these cultures. Approximately 96% of intracellular radioactivity is found as nucleotides after incubation with [(3)H]adenosine, but more than 85% of glutamate-stimulated released material is found as inosine (60%), hypoxanthine (19.9%) and adenosine (7.8%). The release is prevented by removal of extracellular calcium, by the transporter blocker nitrobenzylthioinosine, or inhibitors of calcium/calmodulin-dependent protein kinase II (CAMK II). The uptake of [(3)H]adenosine, but not of [(3)H]GABA or [(3)H]choline, is also blocked by 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-l-tyrosyl]-4-phenylpiperazine (KN62), N-[2-(N-(4-chlorocinnamyl)-N-methylaminomethyl)phenyl-N-[2-hydroxiethyl]-4-methoxybenzenesulfonamide (KN93) or the myristoylated autocamtide-2-related inhibitory peptide, suggesting that the enzyme modulates the nucleoside transporter. The distribution of intracellular purines was not affected by KN62. These results indicate that activation of glutamate receptors triggers the release of purines from retinal cells by a mechanism involving calcium influx, CAMK II and the nitrobenzylthioinosine-sensitive nucleoside transporter. The regulation of adenosine release by glutamate receptors and CAMK II could have important consequences in the presynaptic control of glutamate release.
authors.affiliation: Program of Neuroimmunology and Department of Neurobiology, Institute of Biology, Federal Fluminense University, Caixa Postal 100180, Niterói, Rio de Janeiro 24001-970, Brazil. robpaes@vm.uff.br
authors.forename: Roberto
authors.initials: R
authors.lastname: Paes-de-Carvalho
authors.forename: Bruno V
authors.initials: BV
authors.lastname: Dias
authors.forename: Rochele A
authors.initials: RA
authors.lastname: Martins
authors.forename: Mariana R
authors.initials: MR
authors.lastname: Pereira
authors.forename: Camila C
authors.initials: CC
authors.lastname: Portugal
authors.forename: Claudia
authors.initials: C
authors.lastname: Lanfredi
mesh_terms: D000241:Adenosine / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002478:Cells, Cultured; D002642:Chick Embryo; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018698:Glutamic Acid / Q000378:metabolism; D019271:Hypoxanthine / Q000378:metabolism; D007288:Inosine / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D033703:Nucleoside Transport Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017729:Presynaptic Terminals / Q000187:drug effects / Q000378:metabolism; D011687:Purines / Q000378:metabolism*; D018091:Receptors, AMPA / Q000819:agonists / Q000378:metabolism; D017470:Receptors, Glutamate / Q000187:drug effects / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000819:agonists / Q000378:metabolism; D012160:Retina / Q000187:drug effects / Q000378:metabolism*; D009435:Synaptic Transmission / Q000187:drug effects / Q000502:physiology
chemical_list: D004791:Enzyme Inhibitors; D033703:Nucleoside Transport Proteins; D011687:Purines; D018091:Receptors, AMPA; D017470:Receptors, Glutamate; D016194:Receptors, N-Methyl-D-Aspartate; D019271:Hypoxanthine; D018698:Glutamic Acid; D007288:Inosine; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D000241:Adenosine; D002118:Calcium