Co-localization and functional interaction between adenosine A 2A and metabotropic group 5 receptors in glutamatergic nerve terminals of the rat striatum Ricardo J. Rodrigues, Tiago M. Alfaro, Nelson Rebola, Catarina R. Oliveira and Rodrigo A. Cunha Centre for Neuroscience of Coimbra, Institute of Biochemistry, Faculty of Medicine, University of Coimbra, Portugal Abstract The anti-Parkinsonian effect of glutamate metabotropic group 5 (mGluR5) and adenosine A 2A receptor antagonists is believed to result from their ability to postsynaptically control the responsiveness of the indirect pathway that is hyper- functioning in Parkinson’s disease. mGluR5 and A 2A antago- nists are also neuroprotective in brain injury models involving glutamate excitotoxicity. Thus, we hypothesized that the anti- Parkinsonian and neuroprotective effects of A 2A and mGluR5 receptors might be related to their control of striatal glutamate release that actually triggers the indirect pathway. The A 2A agonist, CGS21680 (1–30 nM) facilitated glutamate release from striatal nerve terminals up to 57%, an effect prevented by the A 2A antagonist, SCH58261 (50 nM). The mGluR5 agonist, CHPG (300–600 lM) also facilitated glutamate release up to 29%, an effect prevented by the mGluR5 antagonist, MPEP (10 lM). Both mGluR5 and A 2A receptors were located in the active zone and 57 ± 6% of striatal glutamatergic nerve ter- minals possessed both A 2A and mGluR5 receptors, suggest- ing a presynaptic functional interaction. Indeed, submaximal concentrations of CGS21680 (1 nM) and CHPG (100 lM) synergistically facilitated glutamate release and the facilitation of glutamate release by 10 nM CGS21680 was prevented by 10 lM MPEP, whereas facilitation by 300 lM CHPG was prevented by 10 nM SCH58261. These results provide the first direct evidence that A 2A and mGluR5 receptors are co-located in more than half of the striatal glutamatergic terminals where they facilitate glutamate release in a synergistic manner. This emphasizes the role of the modulation of glutamate release as a likely mechanism of action of these receptors both in striatal neuroprotection and in Parkinson’s disease. Keywords: A 2A receptor, adenosine, glutamate, mGluR5, release, striatum. J. Neurochem. (2005) 92, 433–441. The role of the striatum in the control of motor function is essentially viewed as a balanced activation of its two main pathways, the direct and indirect pathways that cause opposite control of thalamic relays, which then impinge on the motor cortex (Obeso et al. 2000). These two main striatal pathways are both constituted by medial spiny GABAergic neurons and are both triggered by cortico-thalamic glutamat- ergic fibres (Calabresi et al. 1996). These two pathways are modulated in an opposite manner by important modulators of locomotion, mainly dopamine and also adenosine. Thus, medial spiny neurons of the indirect pathway are endowed with inhibitory dopamine D 2 and facilitatory adenosine A 2A receptors, whereas the medial spiny neurons of the direct pathway are endowed with facilitatory D 1 receptors and inhibitory A 1 receptors (Svenningsson et al. 1999). Dopam- ine is considered the chief modulator of the balanced activation of striatal pathways. Thus, in Parkinson’s disease, the most common basal ganglia motor dysfunction, there is a severe decrease of striatal dopamine levels that causes a hyperactivation of the indirect pathway (Obeso et al. 2000). As facilitatory A 2A receptors are selectively located in neurons of the indirect pathway, antagonists of A 2A receptors have been developed as anti-parkinsonian drugs (Schwarzs- child et al. 2002). Furthermore, it is now recognised that the Received June 3, 2004; revised manuscript received July 28, 2004; accepted August 3, 2004. Address correspondence and reprint requests to Rodrigo A Cunha, Institute of Biochemistry, Faculty of Medicine, University of Coimbra, 3004–504 Coimbra, Portugal. E-mail: racunha@clix.pt Abbreviations used: CGS 21680, 2-[4-(2-p-carboxyethyl)phenylamino]- 5¢-N-ethylcarboxamidoadenosine; CHPG, (RS)-2-chloro-5-hydroxyphe- nylglycine; DPCPX, 1,3-dipropyl-8-cyclopentyladenosine; mGluR5, glutamate metabotropic group 5; MPEP, 2-methyl-6-phenylethyl- nyl)pyridine; SCH, 58261, 7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyr- azolo-[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine; vGluT, vesicular glutamate transporter. Journal of Neurochemistry , 2005, 92, 433–441 doi:10.1111/j.1471-4159.2004.02887.x Ó 2004 International Society for Neurochemistry, J. Neurochem. (2005) 92, 433–441 433