5-hydroxytryptamine stimulates phosphorylation of p44/p42 mitogen-activated protein kinase activation in bovine aortic endothelial cell cultures.
5-Hydroxytryptamine (5-HT) is sequestered and released by endothelial cells, acts as an endothelial cell mitogen, promotes the release of nitric oxide (NO), and has been associated with the p44/p42 mitogen-activated protein kinase (MAPK) cascade. NO also acts as a cell mitogen and promotes signals that culminate in the phosphorylation of MAPK. The aim of this study was to test whether endothelial 5-HT receptors stimulate dual (tyrosyl- and threonyl-) phosphorylation of MAPK through a mitogen-activated protein kinase kinase-1 (MEK-1) and eNOS-dependent pathway in bovine aortic endothelial cells (BAECs). As shown by Western blot analysis, 5-HT and the 5-HT1B-selective agonist 5-nonyloxytryptamine (5-NOT) stimulate time- and concentration-dependent (0.001-10 microM) phosphorylation of MAPK in these cells. The agonist-stimulated phosphorylation of MAPK was blocked by the 5-HT1b-receptor antagonist isamoltane (0.01-10 p3M) and the MEK-1 inhibitor PD 098059 ([2-(2'-amino-3'-methoxy-phenyl)-oxanaphthalen-4-one]; 0.01-10 microM¿. The eNOS inhibitor L-N(omega)-iminoethyl-L-ornithine (L-NIO; 0.01-10 microM) failed to block the 1 microM 5-NOT-stimulated responses. Our findings suggest that the 5-HT receptors (specifically 5-HT1B) mediate signals to MEK-1 and subsequently to MAPK through an eNOS-independent pathway in BAECs.
McDuffie
J E
JE
Department of Pathology, University of Michigan Medical School, Ann Arbor 48109, USA. mcdphd@path.med.umich.edu
Motley
E D
ED
Limbird
L E
LE
Maleque
M A
MA
D000818:Animals; D001345:Autoradiography; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005419:Flavonoids / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism; D009952:Ornithine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011985:Receptors, Serotonin / Q000187:drug effects; D012701:Serotonin / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D005419:Flavonoids; D011985:Receptors, Serotonin; D012701:Serotonin; C065027:N(G)-iminoethylornithine; D009952:Ornithine; D019950:Mitogen-Activated Protein Kinase 1; D020928:Mitogen-Activated Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one