Specific subclass of adenosine receptors modulate release of EETs and superoxide in brain tissues

Three adenosine receptor types (A1, A2 and A3) are distributed in neurons, glia, endothelium and vascular smooth muscle. Two A2 receptor subclasses A2a and A2b are associated with vascular function. The adenosine receptor type mediating the formation of EETs or superoxide in the brain is unknown. We examined the role of adenosine receptor subtypes in EETs release and generation of superoxide in cultured astrocytes and in brains of A2a receptor knock mouse. Stimulation of astrocytes with the A2a agonist CGS-21680 (1 μM) induced a greater production of superoxide than that induced by the non-selective A2a and A2b agonist NECA (1 μM). The A2a receptor antagonist ZM 241385 (1 μM) reduced the CGS 21680-induced but not the NECA-induced superoxide generation. A2a receptor knockout reduced basal level of superoxide, whereas this A2a knock down had no effect on the increase in superoxide stimulated by CGS-21680 or NECA. Results of LC-MS analysis revealed that A2a receptor knock out markedly reduced basal release of EETs compared to that in the wild type. Stimulation of astrocytes with the A2a agonist CGS-21680 (1 μM) or with adenosine (1 μM) increased the release of EETs that was attenuated by the A2a receptor blockade but not by A2a knock out. These results indicate that A2a adenosine receptor is not the only subtype that is important for basal or adenosine evoked release of EETs or superoxide in the brain.