Aim: 

Urocortin II (ucn II) was shown in anesthetized pigs to increase coronary blood flow through a nitric oxide synthase (eNOS) related pathway and the involvement of the subtype 2 of CRF receptors (CRFR2). However information regarding the intracellular signalling induced by ucn II involving CRFR2 and nitric oxide (NO) is scarce. The present study was planned to better characterize the mechanism of action of ucn II leading to NO production.

Methods: 

In porcine aortic endothelial cells (PAE) the effects of ucn II on NO production and ERK, Akt, p38 and eNOS phosphorylation were examined in absence or presence of various agents such as the adenyl cyclase agonist and antagonist (forskolin and 2'5' dideoxyadenosine), the Ca²⁺ ionophore A23187, the Ca²⁺-calmodulin-kinase inhibitor KN93, the CRFR2 blocker astressin 2B and of the intracellular kinases specific inhibitors (UO126, wortmannin, SB203580). In some samples the experiments were performed in presence of EDTA.

Results: 

Ucn II caused an increase of NO production of about 36.4% (p < 0.05), which was amplified by forskolin (155.8%; p < 0.05) and A23187 (107.6%; p < 0.05). All effects of ucn II were prevented by l-NAME, 2'5' dideoxyadenosine, KN93, EDTA and astressin 2B (p > 0.05). Similarly, pre-treatment of cells with UO126, wortmannin and SB203580 abolished all effects of ucn II on NO production. Western Blot analysis confirmed the involvement of ERK, Akt and p38 in the eNOS activation.

Conclusion: 

In PAE ucn II interaction with CRFR2 causes a cAMP-dependent and Ca²⁺-related phoshorylation of ERK, Akt and p38 leading to eNOS activation.