Role of adenosine A(1) receptors in modulating extracellular adenosine levels.

The purpose of this investigation was to test the hypothesis that A(1) receptors modulate extracellular levels of adenosine in cardiovascular tissues. Rat cardiac fibroblasts and human aortic vascular smooth muscle cells were cultured to confluence and various pharmacological agents were applied to the cultures. The extracellular fluid was extracted and adenosine concentrations were measured by HPLC. Three selective A(1) receptor antagonists, namely 8-cyclopentyl-1,3-dipropylxanthine, xanthine amine congener, and N-0840, at a concentration of 10 nM significantly increased extracellular levels of adenosine in both rat cardiac fibroblasts and human aortic vascular smooth muscle cells. Further studies in rat cardiac fibroblasts revealed that the effects of A(1) receptor blockade on extracellular adenosine levels were concentration dependent and prevented by inhibition of G(i) proteins with pertussis toxin or blockade of ecto-5'-nucleotidase with alpha, beta-methyleneadenosine-5'-diphosphate. In cardiac fibroblasts in which the extracellular levels of endogenous adenosine were increased, the ability of A(1) receptor blockade to augment extracellular adenosine was attenuated. A time-course study revealed a time lag of several hours between blockade of A(1) receptors and increases in extracellular adenosine levels. These data suggest that A(1) receptors function to detect the long-term levels of extracellular adenosine, and appropriately adjust extracellular adenosine levels by a slow-onset mechanism involving G(i) proteins and ecto-5'nucleotidase.