5-Hydroxytryptamine Evokes Endothelial Nitric Oxide Synthase Activation in Bovine Aortic Endothelial Cell Cultures

Abstract

Activation of endothelial nitric oxide synthase (eNOS) results in the production of nitric oxide (NO) that mediates the vasorelaxing properties of endothelial cells. The goal of this project was to address the possibility that 5-hydroxytryptamine (5-HT) stimulates eNOS activity in bovine aortic endothelial cell (BAEC) cultures. Here, we tested the hypothesis that 5-HT receptors mediate eNOS activation by measuring agonist-stimulated [3H]L-citrulline ([3H]L-Cit) formation in BAEC cultures. We found that 5-HT stimulated the conversion of [3H]L-arginine ([3H]L-Arg) to [3H]L-Cit, indicating eNOS activation. The high affinity 5-HTl receptor agonist, 5-nonyloxytryptamine (5-NOT)-stimulated [3H]L-Cit turnover responses were concentration-(0.01 nM to 100 pM) and time-dependent. Maximal responses were observed within 10 min following agonist exposures. These responses were effectively blocked by the 5-HT,, receptor antagonist, isamoltane, the 5-HT,$5-HT2 receptor antagonist, methiothepin, and the eNOS selective antagonists (0.01-10 pM): L-N"-monornethyl-L-arginine (L-NMMA) and L-Nu-iminoethyl-L-ornithine (L-NIO). Pretreatment of BAEC cultures with pertussis toxin (PTX; 1-100 ng/ml) for 16 hr resulted in significant inhibition of the agoniststimulated eNOS activity, indicating the involvement of Gi proteins. These findings lend evidence of a 5-HTlB receptor/eNOS pathway, accounting in part for the activation of eNOS by 5-HT. Further investigation is needed to determine the role of other vascular 5-HT receptors in the stimulation of eNOS activity. [P.S.E.B.M. 1999, Vol 2211everal vasoactive substances elicit the release of nitric oxide (NO) from endothelium following activation of S the calciumhalmodulin-dependent enzyme endothelial nitric oxide synthase (eNOS) (l), which has been cloned in both cultured and native bovine aortic endothelial cell (BAEC) cultures (2). The diffusion of NO into vascular smooth muscle cells (VSMCs) results in activation of soluble guanylyl cyclase (solGC) and subsequent vasorelaxation (1). 5-Hydroxytryptamine (5-HT) stimulates endothe-This project was supported by grants. NSF/RIMI 9550699 and NIH T32 HL077809 I To whom requests for reprints should be addressed

5-Hydroxytryptamine Lowers Blood Pressure in Normotensive and Hypertensive Rats

“…However, Western blot analyses of the endothelial cell marker Tie2 suggest this is not the case, although we recognize these blots are less than ideal. Alternatively, 5-HT may directly up-regulate or activate eNOS, an idea supported by work in bovine aortic endothelial cells (McDuffie et al, 1999;Richardson et al, 2003). We have initiated cultures of rat aortic endothelial cells to examine these ideas.…”

Protective effect of tryptophan against dextran sulfate sodium- induced experimental colitis

“…Another metabolite, 5-hydrotryptamine (5-HT), was reported to be involved in the pathogenesis of inflammation in experimental colitis (14). Moreover, 5-HT have been reported to accelerate NO production via endothelial NOS (eNOS) in vitro (15).…”

Serotonin induces the expression of tissue factor and plasminogen activator inhibitor-1 in cultured rat aortic endothelial cells

“…39 In the present study, the stimulation of RAECs to produce TF and PAI-1 required high concentration and prolonged incubation of 5-HT; however, these are in agreement with the earlier reports in other cell types and in other responses related to 5-HT. 7,27,40,41 On the basis of these observations, it is considered to be relevant that high 5-HT concentration levels persist at local sites until TF and PAI-1 proteins express.…”