• adenosine A2A receptor
• AMPA
• basal ganglia
• glutamate
• mGluRs
• phosphorylation
• signaling
• striatum

The striatum is the main input structure of the basal ganglia circuit and receives a dense glutamatergic innervation from several areas of cortex and thalamus. Glutamate acts on striatal neurons through two main classes of receptors, namely ionotropic and metabotropic receptors. Ionotropic receptors include AMPA (3-hydroxy-5-methylisoxazole-4-propionic acid), NMDA (N-methyl-D-aspartic acid) and kainate receptors, whereas metabotropic glutamate receptors (mGluRs) include three groups of receptors classified on the basis of their pharmacology and signaling properties. Group I mGluRs are widely expressed in striatum and enclose two receptor subtypes, mGlu1 and mGlu5.
Phosphorylation is an important mechanism for the post-translational modulation of ionotropic glutamate receptors, and in this study, I investigated the regulation of AMPA receptor GluR1 subunit phosphorylation by the stimulation of group I mGluRs in the mouse dorsal striatum. To this purpose, I used striatal slices as experimental model of investigation. Stimulation of group I mGluRs with DHPG (3,5-dihydroxyphenylglycine) was found to increase GluR1 phosphorylation at the cAMP-dependent protein kinase (PKA) site, Ser845, in a concentration-dependent manner. This effect was abolished by treating striatal slices with the selective mGlu5 antagonist MPEP (2-methyl-6-(phenylethynyl) pyridine hydrochloride) but not with the selective mGlu1 antagonist LY367385, thus suggesting a major contribution of mGlu5 receptors in the phosphorylation of GluR1 at Ser845.
Blockade of dopamine D1 receptors with SCH23390 did not affect phosphorylation of Ser845 evoked by stimulation of group I mGluRs. Conversely, blockade of adenosine A2A receptors with ZM241385, or treatment with adenosine deaminase (ADA) which converts endogenous adenosine to inosine, abolished the effect of DHPG. These results suggest that mGlu5 receptors require endogenous adenosine, and consequently the activation of A2A receptors, to promote phosphorylation of GluR1 at Ser845.
Direct stimulation of A2A receptors with the selective agonist CGS21680 did not produce any significant effect on phosphorylation of GluR1. However, co-treatment of striatal slices with DHPG restored the increase in phosphorylation of Ser845. These findings demonstrate that mGlu5 receptors enhance phosphorylation of GluR1 at Ser845 potentiating the effect of endogenous adenosine on A2A receptors.
Since adenosine A2A receptors in striatum are selectively expressed in striatopallidal neurons, the results presented here also indicate that the effect exerted by mGlu5 receptors occurs selectively in this neuronal subpopulation.
The increase in Ser845 phosphorylation was also found to be dependent on DARPP-32 (Dopamine and cAMP-regulated phosphoprotein of 32 kDa), a protein highly enriched in striatal neurons. Indeed, the effect produced by DHPG was abolished in mutant mice in which the PKA phosphorylation site on DARPP-32 was mutated into an alanin residue. When phosphorylated by PKA, DARPP-32 inhibits protein phosphatase-1 (PP-1), which dephosphorylates GluR1 at Ser845. Therefore, these results suggest that DARPP-32-mediated inhibition of PP-1 is necessary to preserve the effect of DHPG on AMPA receptors.
Collectively, these data demonstrate that stimulation of mGlu5 receptors enhances the phosphorylation of GluR1 subunit at Ser845 in striatopallidal neurons, through two pathways: a direct pathway, in which mGlu5 receptors activate PKA recruiting the A2A receptors signal transduction machinery thus potentiating the effect of endogenous adenosine, and an indirect pathway, in which the increment in phosphorylation is obtained through an inhibition of dephosphorylation mediated by DARPP-32.
These findings clarify the molecular mechanisms underlying the glutamatergic neurotransmission in striatum, which is involved in the striatal synaptic plasticity and consequently in behavioral and cognitive functions of this nucleus.