Adenosine A(1) receptor-mediated inhibition of protein kinase A-induced calcitonin gene-related peptide release from rat trigeminal neurons

Calcitonin gene-related peptide (CGRP), a potent vasodilator, has been impl icated in the pathogenesis of migraine. Its release from adult rat trigemin al neurons in culture was shown to be markedly increased by the activation of adenylate cyclase with forskolin, Modulation of this secretion was inves tigated by a number of agents with known inhibitory effects on cAMP generat ion mediated via receptor coupling to G(i/o) proteins. Significantly, forsk olin-stimulated CGRP release could be closely correlated with the phosphory lation of the protein kinase A (PKA) substrate cyclic AMP response element- binding protein (CREB). Forskolin-stimulated CGRP release could be potently and effectively inhibited by the adenosine A(1) receptor-selective agonist GR79236X (pIC(50) = 7.7 +/- 0.1, maximal inhibition 65 +/- 2.5% at 300 nM) , whereas the A(2A) (CGS21680) and the A(3) (2-chloro-N-6-(3-iodobenzyl)- a denosine-5'-N-methyluronamide) receptor-selective agonists were without eff ect. GR79236X-mediated inhibition was abolished by the A(1) receptor antago nist 8-cyclopentyl-1,3-dipropylxanthine. Immunocytochemical studies and Wes tern analysis revealed the presence of adenosine A(1) receptors on trigemin al neurons. However, despite the additional detection of 5-hydroxytryptamin e (5-HT)(1B) receptors on these cells, the clinically effective antimigrain e 5-HT1B/1D agonist sumatriptan did not inhibit forskolin-stimulated CGRP r elease nor did it show any effect on the concomitant CREB phosphorylation. In contrast, the mu -opioid agonist fentanyl elicited a 74 +/- 4% reduction in CGRP levels. Forskolin-stimulated CGRP release and CREB phosphorylation could be mimicked by incubation of the cells with chlorophenylthio-cAMP an d blocked by pretreatment with the PKA inhibitor myrPKI(14-22). Taken toget her, the present data confirm the PKA-dependence of forskolin-stimulated CG RP release and suggest that A(1) adenosine agonists may warrant further inv estigation in models of migraine and neurogenic inflammation.