Adenosine2A receptor vasodilation of rat preglomerular microvessels is mediated by EETs that activate the cAMP/PKA pathway

Dilation of rat preglomerular microvessels (PGMV) by activation of adenosine A(2A) receptors (A(2A)R) is coupled to epoxyeicosatrienoic acid (EET) release. We have investigated the commonality of this signal transduction pathway, i.e., sequential inhibition of G(s alpha), adenylyl cyclase, PKA, and Ca2+-activated K+ (K-Ca) channel activity, to the vasoactive responses to A(2A)R activation by a selective A(2A) agonist, CGS-21680, compared with those of 11,12-EET. Male Sprague-Dawley rats were anesthetized, and microdissected arcuate arteries (110-130 mu m) were cannulated and pressurized to 80 mmHg. Vessels were superfused with Krebs solution containing N-G-nitro-L-arginine methyl ester (L-NAME) and indomethacin and preconstricted with phenylephrine. We assessed the effect of 3-aminobenzamide (10 mu M), an inhibitor of mono-ADP-ribosyltranferases, on responses to 11,12-EET (3 nM) and CGS-21680 (10 mu M) and found that both were inhibited by similar to 70% (P < 0.05), whereas the response to SNP (10 mu M) was unaffected. Furthermore, 11,12-EET (100 nM), like cholera toxin (100 ng/ml), stimulated ADP-ribose formation in homogenates of arcuate arteries compared with control. SQ-22536 (10 mu M), an inhibitor of adenylyl cyclase activity, and myristolated PKI (14-22) amide (5 mu M), an inhibitor of PKA, decreased activity of 11,12-EET and CGS-21680. Incubation of 11,12- EET (3 nM- 3 mu M) with PGMV resulted in an increase in cAMP levels (P < 0.05). The responses to both 11,12-EET and CGS-21680 were significantly reduced by superfusion of iberiotoxin ( 100 nM), an inhibitor of KCa channel activity. Thus in rat PGMV activation of A(2A)R is coupled to EET release upstream of adenylyl cyclase activation and EETs stimulate mono-ADP-ribosyltransferase, resulting in Gs(alpha) protein activation.