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The spontaneously hypertensive rat (SHR) is an excellent animal model that mimics the behavioral and neurochemical phenotype of attention-deficit/hyperactivity disorder (ADHD). Here, we characterized the striatal GABA transport of SHR and investigated whether caffeine, a non-selective antagonist of adenosine receptors, could influence GABAergic circuitry. For this purpose, ex vivo striatal slices of SHR and Wistar (control strain) on the 35th postnatal day were dissected and incubated with [3H]-GABA to quantify the basal levels of uptake and release. SHR exhibited a reduced [3H]-GABA uptake and release, suggesting a defective striatal GABAergic transport system. GAT-1 appears to be the primary transporter for [3H]-GABA uptake in SHR striatum, as GAT-1 selective blocker, NO-711, completely abolished it. We also verified that acute exposure of striatal slices to caffeine improved [3H]-GABA uptake and release in SHR, whereas Wistar rats were not affected. GABA-uptake increase and cAMP accumulation promoted by caffeine was reverted by A1R activation with N6-cyclohexyl adenosine (CHA). As expected, the pharmacological blockade of cAMP-PKA signaling by H-89 also prevented caffeine-mediated [3H]-GABA uptake increment. Interestingly, a single caffeine exposure did not affect GAT-1 or A1R protein density in SHR, which was not different from Wistar protein levels, suggesting that the GAT-1-dependent transport in SHR has a defective functional activity rather than lower protein expression. The current data support that caffeine regulates GAT-1 function and improves striatal GABA transport via A1R-cAMP-PKA signaling, specifically in SHR. These results reinforce that caffeine may have therapeutic use in disorders where the GABA transport system is impaired., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Caffeine is the most consumed psychostimulant drug in the world, acting as a non-selective antagonist of adenosine receptors A 1 R and A 2A R, which are widely expressed in retinal layers. We have previously shown that caffeine, when administered acutely, acts on A 1 R to potentiate the NMDA receptor-induced GABA release. Now we asked if long-term caffeine exposure also modifies GABA uptake in the avian retina and which mechanisms are involved in this process. Chicken embryos aged E11 were injected with a single dose of caffeine (30 mg/kg) in the air chamber. Retinas were dissected on E15 for ex vivo neurochemical assays. Our results showed that [ 3 H]-GABA uptake was dependent on Na + and blocked at 4 °C or by NO-711 and caffeine. This decrease was observed after 60 min of [ 3 H]-GABA uptake assay at E15, which is accompanied by an increase in [ 3 H]-GABA release. Caffeine increased the protein levels of A 1 R without altering ADORA1 mRNA and was devoid of effects on A 2A R density or ADORA2A mRNA levels. The decrease of GABA uptake promoted by caffeine was reverted by A 1 R activation with N6-cyclohexyl adenosine (CHA) but not by A 2A R activation with CGS 21680. Caffeine exposure increased cAMP levels and GAT-1 protein levels, which was evenly expressed between E11-E15. As expected, we observed an increase of GABA containing amacrine cells and processes in the IPL, also, cAMP pathway blockage by H-89 decreased caffeine mediated [ 3 H]-GABA uptake. Our data support the idea that chronic injection of caffeine alters GABA transport via A 1 R during retinal development and that the cAMP/PKA pathway plays an important role in the regulation of GAT-1 function., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

l-Glutamate and l-aspartate are the main excitatory amino acids (EAAs) in the Central Nervous System (CNS) and their uptake regulation is critical for the maintenance of the excitatory balance. Excitatory amino acid transporters (EAATs) are widely distributed among central neurons and glial cells. GLAST and GLT1 are expressed in glial cells, whereas excitatory amino acid transporter 3/excitatory amino acid carrier 1 (EAAT3/EAAC1) is neuronal. Different signaling pathways regulate glutamate uptake by modifying the activity and expression of EAATs. In the present work we show that immature postnatal day 3 (PN3) rat retinas challenged by l-glutamate release [ 3 H]-d-Aspartate linked to the reverse transport, with participation of NMDA, but not of non-NMDA receptors. The amount of [ 3 H]-d-Aspartate released by l-glutamate is reduced during retinal development. Moreover, immature retinae at PN3 and PN7, but not PN14, exposed to a single dose of 200 or 500μM caffeine or the selective A2A receptor (A2AR) antagonist 100nM ZM241385 decreased [ 3 H]-d-Aspartate uptake. Caffeine also selectively increased total expression of EAAT3 at PN7 and its expression in membrane fractions. However, both EAAT1 and EAAT2 were reduced after caffeine treatment in P2 fraction. Addition of 100nM DPCPX, an A1 receptor (A1R) antagonist, had no effect on the [ 3 H]-d-Aspartate uptake. [ 3 H]-d-Aspartate release was dependent on both extracellular sodium and Dl-TBOA, but not calcium, implying a transporter-mediated mechanism. Our results suggest that in the developing rat retina caffeine modulates [ 3 H]-d-Aspartate uptake by blocking adenosine A2AR., (Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.)