title: Caffeine exposure alters adenosine system and neurochemical markers during retinal development.
abstract: Evidence points to beneficial properties of caffeine in the adult central nervous system, but teratogenic effects have also been reported. Caffeine exerts most of its effects by antagonizing adenosine receptors, especially A1 and A2A subtypes. In this study, we evaluated the role of caffeine on the expression of components of the adenosinergic system in the developing avian retina and the impact of caffeine exposure upon specific markers for classical neurotransmitter systems. Caffeine exposure (5-30 mg/kg by in ovo injection) to 14-day-old chick embryos increased the expression of A1 receptors and concomitantly decreased A2A adenosine receptors expression after 48 h. Accordingly, caffeine (30 mg/kg) increased [(3) H]-8-cyclopentyl-1,3-dipropylxanthine (A1 antagonist) binding and reduced [(3) H]-ZM241385 (A2A antagonist) binding. The caffeine time-response curve demonstrated a reduction in A1 receptors 6 h after injection, but an increase after 18 and 24 h. In contrast, caffeine exposure increased the expression of A2A receptors from 18 and 24 h. Kinetic assays of [(3) H]-S-(4-nitrobenzyl)-6-thioinosine binding to the equilibrative adenosine transporter ENT1 revealed an increase in Bmax with no changes in Kd , an effect accompanied by an increase in adenosine uptake. Immunohistochemical analysis showed a decrease in retinal content of tyrosine hydroxylase, calbindin and choline acetyltransferase, but not Brn3a, after 48 h of caffeine injection. Furthermore, retinas exposed to caffeine had increased levels of phosphorylated extracellular signal-regulated kinase and cAMP-response element binding protein. Overall, we show an in vivo regulation of the adenosine system, extracellular signal-regulated kinase and cAMP-response element binding protein function and protein expression of specific neurotransmitter systems by caffeine in the developing retina. The beneficial or maleficent effects of caffeine have been demonstrated by the work of different studies. It is known that during animal development, caffeine can exert harmful effects, impairing the correct formation of CNS structures. In this study, we demonstrated cellular and tissue effects of caffeine's administration on developing chick embryo retinas. Those effects include modulation of adenosine receptors (A1 , A2 ) content, increasing in cAMP response element-binding protein (pCREB) and extracellular signal-regulated kinase phosphorylation (pERK), augment of adenosine equilibrative transporter content/activity, and a reduction of some specific cell subpopulations. ENT1, Equilibrative nucleoside transporter 1.
authors.affiliation: Neurobiology of the Retina Laboratory, Department of Neurobiology and Program of Neurosciences, Institute of Biology, Fluminense Federal University, Niterói, Rio de Janeiro, Brazil.|Laboratory of Cellular Neurobiology, Department of Neurobiology and Program of Neurosciences, Institute of Biology, Fluminense Federal University, Niterói, Rio de Janeiro, Brazil.
authors.forename: Rafael
authors.initials: R
authors.lastname: Brito
authors.affiliation: Neurobiology of the Retina Laboratory, Department of Neurobiology and Program of Neurosciences, Institute of Biology, Fluminense Federal University, Niterói, Rio de Janeiro, Brazil.
authors.forename: Danniel
authors.initials: D
authors.lastname: Pereira-Figueiredo
authors.affiliation: Instituto de Investigação e Inovação em Saúde (i3S) and Instituto de Biologia Molecular e Celular (IBMC), Universidade do Porto, Porto, Portugal.
authors.forename: Renato
authors.initials: R
authors.lastname: Socodato
authors.affiliation: Laboratory of Cellular Neurobiology, Department of Neurobiology and Program of Neurosciences, Institute of Biology, Fluminense Federal University, Niterói, Rio de Janeiro, Brazil.
authors.forename: Roberto
authors.initials: R
authors.lastname: Paes-de-Carvalho
authors.affiliation: Neurobiology of the Retina Laboratory, Department of Neurobiology and Program of Neurosciences, Institute of Biology, Fluminense Federal University, Niterói, Rio de Janeiro, Brazil.
authors.forename: Karin C
authors.initials: KC
authors.lastname: Calaza
mesh_terms: D000241:Adenosine / Q000378:metabolism*; D058916:Adenosine A1 Receptor Antagonists / Q000494:pharmacology; D058908:Adenosine A2 Receptor Agonists / Q000494:pharmacology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D002642:Chick Embryo; D002645:Chickens; D000242:Cyclic AMP / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D058915:Purinergic P1 Receptor Antagonists; D043682:Receptor, Adenosine A1 / Q000378:metabolism; D043705:Receptor, Adenosine A2A / Q000187:drug effects / Q000378:metabolism; D012160:Retina / Q000187:drug effects / Q000254:growth & development*
chemical_list: D058916:Adenosine A1 Receptor Antagonists; D058908:Adenosine A2 Receptor Agonists; D017362:Cyclic AMP Response Element-Binding Protein; D058915:Purinergic P1 Receptor Antagonists; D043682:Receptor, Adenosine A1; D043705:Receptor, Adenosine A2A; D002110:Caffeine; D000242:Cyclic AMP; D000241:Adenosine
keywords: A1 and A2A receptors; CNS development; ENT 1; avian embryos; caffeine; inner retina