Coaggregation, Cointernalization, and Codesensitization of Adenosine A2A Receptors and Dopamine D2Receptors

Abstract

Adenosine and dopamine signaling exert opposite effects in the basal ganglia, a brain region involved in sensory-motor integration. Thus, adenosine agonists induce motor depression and adenosine antagonists, such as caffeine, produce motor activation (1). These opposite effects result from specific antagonistic interactions between subtypes of adenosine and dopamine receptors in the striatum, the main input structure of the basal ganglia. In fact, striatal dopamine receptors and, to some extent, adenosine receptors are segregated in the two main populations of ␥-aminobutyric acid (GABA) efferent neurons. EXPERIMENTAL PROCEDURESCell Cultures-Maintenance of SH-SY5Y cells (parental and D 2 Rtransfected cells) as well as the pharmacological characterization and maintenance of D 2 R-and D 1 R-transfected mouse fibroblast Ltk Ϫ cells are described in detail elsewhere (7-9). For primary cultures, striata were removed from 16-day-old Sprague-Dawley rat embryos (B&K Universal) in Ca 2ϩ /Mg 2ϩ -free PBS supplemented with 20 units/ml penicillin and 20 g/ml streptomycin (Invitrogen). The tissue fragments were pooled and mechanically dissociated in SFM Neurobasal serum-free medium (Invitrogen), supplemented with B27 (Invitrogen), glutamine (2 mM; Invitrogen), penicillin/streptomycin (20 units/ml/20 g/ml; Invitrogen), and ␤-mercaptoethanol (25 M) (Invitrogen). Cells were collected by centrifugation at 100 ϫ g for 5 min and resuspended in fresh medium. The resulting single-cell suspension was seeded on 24-well plates coated with gelatin (Sigma) and poly-L-lysine (Sigma), and cells were grown at 37°C in saturation humidity with 5% CO 2 .Immunolabeling Experiments-Neuroblastoma cells were grown on glass coverslips coated with poly-L-lysine (Sigma) and exposed to vari-* This work was

Stimulation of Adenosine Receptors Selectively Activates Gene Expression in Striatal Enkephalinergic Neurons

“…Stimulation of A 2A receptors decreases the affinity of D 2 receptors for agonists by means of an intramembrane interaction (Ferré et al, 1991), while stimulation of D 2 receptors inhibits A 2A receptor-induced activation of adenylyl-cyclase (Kull et al, 1999;Hillion et al, 2002). By means of the strong antagonistic D 2 -A 2A receptor interaction at the adenylyl cyclase level, and due to the existence of a tonic effect of dopamine on D 2 receptors, under normal conditions A 2A receptors are practically unable to activate the cAMP-PKA signaling pathway (Agnati et al, 2003;Ferré et al, 2003Ferré et al, , 2004.…”

“…A 2A and D 2 receptors exert reciprocal antagonistic interactions that modulate the function of GABAergic enkephalinergic neurons (Ferré et al, 1993(Ferré et al, , 1997Agnati et al, 2003) and recent studies have demonstrated the ability of A 2A and D 2 receptors to heteromerize Kamiya et al, 2003;Ciruela et al, 2004;Woods and Ferré, 2005). A 2A and D 2 receptors are prototypical G s -and G i -coupled receptors, respectively, which play opposite effects on adenylyl-cyclase (Kull et al, 1999;Hillion et al, 2002;Lee et al, 2002). Stimulation of A 2A receptors can potentially stimulate striatal adenylyl-cyclase, with the consequent activation of the cAMP-PKA signaling pathway and the induction of the expression of different genes, such as c-fos and preproenkephalin (Agnati et al, 2003;Ferré et al, 2003Ferré et al, , 2004.…”

Simultaneous Blockade of Adenosine A2A and Metabotropic Glutamate mGlu5 Receptors Increase their Efficacy in Reversing Parkinsonian Deficits in Rats

“…These results also demonstrate for the first time that a chronic A 2A blockade, via the administration of its selective antagonist CSC, is effective against the 6-OHDA-induced akinetic deficits. In view of the recent demonstration that A 2A and mGlu 5 receptors can form heteromeric complexes (Ferré et al, 2002) as well as coaggregate and cointernalize with the DA D 2 receptors after their prolonged stimulation (Hillion et al, 2002), the idea of a reciprocal adaptation of these receptors subtypes appear to be further corroborated. It seems therefore conceivable that intramembrane interactions implying common final pathways between striatal A 2A and mGlu 5 receptors could have been involved in the functional synergism underlying the efficient recovery of the akinesia observed in the present study.…”

Striatal Adenosine A2A and Cannabinoid CB1 Receptors Form Functional Heteromeric Complexes that Mediate the Motor Effects of Cannabinoids

“…Animals were maintained in accordance with guidelines of the Institutional Care and Use Committee of the Intramural Research Program, National Institute on Drug Abuse, NIH. Rats were (Hillion et al, 2001) and anti-CB 1 receptor rabbit polyclonal IgG (PA1-745; Affinity BioReagents, Golden, CO, USA). The selectivity of both antibodies has been previously characterized (Hillion et al, 2001;Twitchell et al, 1997 Immunocytochemistry SH-SY5Y cells were grown on glass coverslips coated with poly-L-lysine (Sigma).…”

“…Rats were (Hillion et al, 2001) and anti-CB 1 receptor rabbit polyclonal IgG (PA1-745; Affinity BioReagents, Golden, CO, USA). The selectivity of both antibodies has been previously characterized (Hillion et al, 2001;Twitchell et al, 1997 Immunocytochemistry SH-SY5Y cells were grown on glass coverslips coated with poly-L-lysine (Sigma). At 60% confluence, cells were rinsed with PBS, fixed in 4% paraformaldehyde for 15 min, and washed with PBS containing 20 mM glycine.…”