8-11-14-eicosatrienoic-acid and lucifer-yellow

Functional gap junctional communication between vascular cells has been implicated in ascending dilatation and the cytochrome P-450 (CYP) inhibitor-sensitive and NO- and prostacyclin-independent dilatation of many vascular beds. Here, we assessed the mechanisms by which the epoxyeicosatrienoic acids (EETs) generated by a CYP 2C enzyme control interendothelial gap junctional communication. In CYP 2C-expressing porcine coronary endothelial cells, bradykinin, which enhances EET formation, elicited a biphasic effect on the electrical coupling and transfer of Lucifer yellow between endothelial cells, consisting of a transient increase in coupling followed by a sustained uncoupling. The initial phase was sensitive to the CYP 2C9 inhibitor sulfaphenazole and the protein kinase A (PKA) inhibitors Rp-cAMPS and KT5720 and could be mimicked by forskolin and caged cAMP as well as by the PKA activators 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole 3',5'-cyclic monophosphorothioate sodium salt and Sp-cAMPS. Gap junction uncoupling in bradykinin-stimulated porcine coronary endothelial cells was prevented by inhibiting the activation of extracellular signal-regulated kinase (ERK)1/2. In human endothelial cells, which express little CYP 2C, bradykinin elicited only an ERK1/2-mediated inhibition of intercellular communication. The CYP 2C9 product, 11,12-EET, also exerted a dual effect on the electrical and dye coupling of human endothelial cells, which was sensitive to PKA inhibition. These results demonstrate that an agonist-activated CYP-dependent pathway as well as 11,12-EET can positively regulate interendothelial gap junctional communication, most probably via the activation of PKA, an effect that is curtailed by the subsequent activation of ERK1/2.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Aryl Hydrocarbon Hydroxylases; Biological Transport; Bradykinin; Cell Communication; Cells, Cultured; Colforsin; Connexin 43; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Endothelium, Vascular; Enzyme Activators; Enzyme Inhibitors; Fluorescent Dyes; Gap Junctions; Humans; Isoquinolines; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Patch-Clamp Techniques; RNA, Messenger; Signal Transduction; Steroid 16-alpha-Hydroxylase; Steroid Hydroxylases; Swine; Vasodilator Agents