Signaling Pathways Linked to Serotonin-Induced Superoxide Anion Production: A Physiological Role for Mitochondria in Pulmonary Arteries.

Serotonin (5-HT) is a potent vasoconstrictor agonist and contributes to several vascular diseases including systemic or pulmonary hypertension and atherosclerosis. Although superoxide anion (O₂^•-) is commonly associated to cellular damages due to O₂^•- overproduction, we previously demonstrated that, in physiological conditions, O₂^•- also participates to the 5-HT contraction in intrapulmonary arteries (IPA). Here, we focused on the signaling pathways leading to O₂^•- production in response to 5-HT in rat IPA. Using electron paramagnetic resonance on rat IPA, we showed that 5-HT (100μM)-induced O₂^•- production was inhibited by ketanserin (1μM-an inhibitor of the 5-HT₂ receptor), absence of extracellular calcium, two blockers of voltage-independent calcium permeable channels (RHC80267 50μM and LOE-908 10μM) and a blocker of the mitochondrial complex I (rotenone-100 nM). Depletion of calcium from the sarcoplasmic reticulum or nicardipine (1μM-an inhibitor of the L-type voltage-dependent calcium channel) had no effect on the 5-HT-induced O₂^•- production. O₂^•- levels were also increased by a-methyl-5-HT (10μM-a 5-HT₂ receptors agonist) whereas GR127935 (1μM-an antagonist of the 5-HT 1B/D receptor) and citalopram (1μM-a 5-HT transporter inhibitor) had no effect on the 5-HT-induced O₂^•- production. Peroxynitrites were increased in response to 5-HT (100μM). In isolated pulmonary arterial smooth muscle cells loaded with rhod-2 or mitosox probes, we respectively showed that 5-HT increased both mitochondrial calcium and O₂^•- levels, which were both abrogated in absence of extracellular calcium. Mitochondrial O₂^•- levels were also abolished in the presence of rotenone (100 nM). In pulmonary arterial smooth muscle cells loaded with TMRM, we showed that 5-HT transiently depolarized the mitochondrial membrane whereas in the absence of extracellular calcium the mitochondrial membrane depolarisation was delayed and sustained in response to 5-HT. 5-HT decreased the mitochondrial respiratory rate measured with a Clark oxygen electrode. Altogether, in physiological conditions, 5-HT acts on 5-HT₂ receptors and induces an O₂^•- production dependent on extracellular calcium and mitochondria. [ABSTRACT FROM AUTHOR]