Cyclic AMP-Adenosine Pathway Induces Nitric Oxide Synthesis in Aortic Smooth Muscle Cells

Abstract

The main purpose of this investigation was to evaluate whether the cyclic AMP-adenosine pathway, ie, the conversion of cAMP to AMP and, hence, to adenosine, is involved in the regulation of nitric oxide (NO) synthesis by vascular smooth muscle cells (SMCs). Treatment of confluent monolayers of SMCs with adenosine, 2-chloroadenosine (stable analog of adenosine), and agents that elevate endogenous (SMC-derived) adenosine (EHNA and iodotubericidin) increased nitrite/nitrate (stable metabolites of NO) levels in the medium and enhanced the conversion of 3 H-L-arginine to 3 H-L-citrulline by cytosolic extracts obtained from the pretreated SMCs. The stimulatory effects of adenosine were not mimicked by low (1 to 100 nmol/L) concentrations of CGS21680 , an A 2A receptor agonist, or CPA, a selective A 1 receptor agonist. The stimulatory effects of 2-chloroadenosine and EHNA plus iodotubericidin were significantly inhibited by KF17837, a selective A 2 receptor antagonist, and by DPSPX, and A 1 /A 2 receptor antagonist, but not by DPCPX, a selective A 1 receptor antagonist. DDA (adenylyl cyclase inhibitor) and Rp-cyclic AMP (protein kinase A inhibitor) did not block the effects of adenosine on NO synthesis. Incubation of SMCs with exogenous cyclic AMP, at concentrations previously shown to elevate levels of adenosine in the medium, also increased nitrite/nitrate levels and 3 H-L-citrulline formation, and the effects of cyclic AMP on NO synthesis were blocked by DPSPX and KF17837, but not by DPCPX. These findings provide evidence that exogenous and SMC-derived adenosine induce NO synthesis via A 2B receptors linked to a pathway not involving adenylyl cyclase/protein kinase A. Moreover, extracellular cyclic AMP induces NO synthesis via conversion to adenosine and activation of A 2B adenosine receptors. The cyclic AMP-adenosine pathway may be importantly involved in the vascular production of NO.