Serotonin uptake by bovine pulmonary artery endothelial cells in culture. I. Characterization

Lee, Sheu-Ling; Fanburg, Barry L.

doi:10.1152/ajpcell.1986.250.5.c761

Cited by 59 publications

(34 citation statements)

References 13 publications

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“…Our findings that serotonylation is blocked by 5-HTT inhibitors support a role of 5-HTi in this process. 5-HT, at a concentration of 1 mmol/L, was used in these studies because our previous results showed that this concentration of 5-HT is actively transported by the 5-HTT, whereas higher concentrations enter the cell by diffusion (9,10). We also found that serotonylation of proteins is increased in the presence of a MAO inhibitor, which prevents degradation of internalized 5-HT.…”

Section: Discussionmentioning

confidence: 99%

“…We also have shown that 5-HT transport via the 5-HTT triggers activation of platelet-derived growth factor receptor-b and mitogen-activated protein kinase, which participate in proliferation of pulmonary artery SMCs (2,8). Furthermore, inhibition of intracellular monoamine oxidase (MAO), which degrades 5-HT, markedly enhances 5-HT-induced proliferation of SMCs (9)(10)(11). These data indicate that intracellular 5-HT (5-HTi) is an important signal for SMC proliferation; however, the mechanism underlying this activity is unknown.…”

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confidence: 97%

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Role of Protein Transamidation in Serotonin-Induced Proliferation and Migration of Pulmonary Artery Smooth Muscle Cells

Liu

Lin²,

Laskin³

et al. 2011

Am J Respir Cell Mol Biol

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Pulmonary hypertension is characterized by elevated pulmonary artery pressure and pulmonary artery smooth muscle cell (SMC) proliferation and migration. Clinical and experimental evidence suggests that serotonin (5-HT ) is important in these responses. We previously demonstrated the participation of the 5-HT transporter and intracellular 5-HT (5-HTi) in the pulmonary vascular SMCproliferative response to 5-HT. However, the mechanism underlying the intracellular actions of 5-HT is unknown. We speculated that 5-HTi activates SMC growth by post-translational transamidation of proteins via transglutaminase (TGase) activity, a process referred to as serotonylation. To test this hypothesis, serotonylation of pulmonary artery SMC proteins, and their role in 5-HT-induced proliferative and migratory responses, were assessed. 5-HT caused dose-and time-dependent increase in serotonylation of multiple proteins in both bovine and rat pulmonary artery SMCs. Inhibition of TGase with dansylcadaverin blocked this activity, as well as SMCproliferative and migratory responses to 5-HT. Serotonylation of proteins also was blocked by 5-HT transporter inhibitors, and was enhanced by inhibition of monoamine oxidase, an enzyme known to degrade 5-HTi, indicating that 5-HTi levels regulate serotonylation. Immunoprecipitation assays and HPLC-mass spectral peptide sequencing revealed that a major protein serotonylated by TGase was fibronectin (FN). 5-HT-stimulated SMC serotonylation and proliferation were blocked by FN small interfering (si) RNA. These findings, together with previous observations that FN expression in the lung strongly correlates with the progression of pulmonary hypertension in both experimental animals and humans, suggest an important role of FN serotonylation in the pathogenesis of this disease.Keywords: serotonin; transglutaminase; fibronectin; protein serotonylation; smooth muscle cell growth Serotonin (also known as 5-hydroxytryptamine ) is one of the most potent naturally occurring pulmonary vasoconstrictors (1). Clinical, experimental, and human genetic data support a relationship between 5-HT, pulmonary artery remodeling, and pulmonary hypertension. Our laboratories and others have demonstrated the importance of both the 5-HT transporter (5-HTT) and 5-HT receptors in pulmonary vascular smooth muscle cell (SMC)-proliferative and migratory responses to 5-HT (2-7). We also have shown that 5-HT transport via the 5-HTT triggers activation of platelet-derived growth factor receptor-b and mitogen-activated protein kinase, which participate in proliferation of pulmonary artery SMCs (2, 8). Furthermore, inhibition of intracellular monoamine oxidase (MAO), which degrades 5-HT, markedly enhances 5-HT-induced proliferation of SMCs (9-11). These data indicate that intracellular 5-HT (5-HTi) is an important signal for SMC proliferation; however, the mechanism underlying this activity is unknown.Transglutaminases (TGases), which are abundant in blood and vascular SMCs (12, 13), catalyze post-translational modifications of protei...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 97%

Role of Protein Transamidation in Serotonin-Induced Proliferation and Migration of Pulmonary Artery Smooth Muscle Cells

Liu

Lin²,

Laskin³

et al. 2011

Am J Respir Cell Mol Biol

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“…Also, the three drugs having the greatest effect (fluvoxamine, fluoxetine and alaproclate) displayed the same order of potencies as in the synaptosomal [3H]-5-HT uptake inhibition (Wolf & Kuhn, 1991 (Catravas & Gillis, 1980;Bosin & Lahr, 1981;Robinson-White et al, 1981;Lee & Fanburg, 1986). To examine the contribution of endothelial uptake to the increases of extracellular 5-HT in rats, we tested in vivo the effects of a single dose of paroxetine.…”

Section: Discussionmentioning

confidence: 99%

Effects of monoamine uptake inhibitors on extracellular and platelet 5‐hydroxytryptamine in rat blood: different effects of clomipramine and fluoxetine

Ortiz

Artigas

1992

British J Pharmacology

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The concentration of 5‐hydroxytryptamine (5‐HT) in rat platelet‐free plasma increased significantly 30 min after a single i.p. injection (10 mg kg−1) of each of six inhibitors of the high‐affinity 5‐HT uptake (fluvoxamine, fluoxetine, alaproclate, paroxetine, sertraline and clomipramine). The increases ranged from 226% to 776% of control values. In contrast, imipramine, desipramine and femoxetine had no significant effect. The increase elicited by paroxetine was dependent on the dose (1, 5 and 10 mg kg−1) and returned to control values after 4 h. That observed after clomipramine was also transient and paralleled the plasma concentration of the drug (Spearman‐rank correlation r = 0.43). In vivo, the rat pulmonary vascular endothelium removed trace amounts (8.8 nmol in a bolus) of intravenously injected [14C]‐5‐HT. Paroxetine pretreatment (10 mg kg−1, 30 min before‐hand) reduced this uptake by 73%. Repeated fluoxetine treatments reduced rat whole blood 5‐HT concentration (ca. − 60% after daily 2× 5 mg kg−1, i.p. during 14 days). However, plasma (extracellular) 5‐HT was not increased. Various repeated treatments with clomipramine (i.p. injections or osmotic minipumps, up to 30 mg kg−1 day−1), failed to decrease rat whole blood 5‐HT concentrations. Platelet‐free plasma 5‐HT was also unchanged, even after treatments yielding plasma clomipramine levels 2.7 times higher than those that increased it acutely. These results indicate that the extracellular pool of 5‐HT in rat blood (measured in the platelet‐free plasma) is physiologically under the control of high‐affinity 5‐HT uptake systems. The sustained 5‐HT uptake inhibition does not result in an increase of 5‐HT in platelet‐free plasma, suggesting that adaptative mechanisms are triggered. The distinct long‐term effects of the two antidepressants clomipramine and fluoxetine on rat whole blood 5‐HT suggest a differential in vivo action on the rat 5‐HT uptake.

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“…Functionally, this transporter is Na ϩ and Cl Ϫ dependent and is inhibited by SSRIs such as fluoxetine (20). SERT has also been characterized in specialized nonneuronal cells, including platelets (2,26), pulmonary endothelium (19), and placental syncytiotrophoblasts (26). At the intestinal level, previous studies have shown that rat and guinea pig mucosal epithelial cells express SERT mRNA and display functional 5-HT uptake through a Na ϩ /Cl Ϫ -dependent process (5,29,31).…”

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confidence: 99%

Function, expression, and characterization of the serotonin transporter in the native human intestine

Gill¹,

Pant²,

Saksena³

et al. 2008

American Journal of Physiology-Gastrointestinal and Liver Physi

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Gill RK, Pant N, Saksena S, Singla A, Nazir TM, Vohwinkel L, Turner JR, Goldstein J, Alrefai WA, Dudeja PK. Function, expression, and characterization of the serotonin transporter in the native human intestine. Am J Physiol Gastrointest Liver Physiol 294: G254-G262, 2008. First published November 8, 2007 doi:10.1152/ajpgi.00354.2007.-The enteric serotonin transporter (SERT) plays a critical role in modulating serotonin availability and thus has been implicated in the pathogenesis of various intestinal disorders. To date, SERT expression and function in the human intestine have not been investigated. Current studies were designed to characterize the function, expression, distribution, and membrane localization of SERT in the native human intestine. Real-time PCR studies showed relatively higher SERT mRNA expression in the human small intestine compared with colon (ileum ϾϾ duodenum ϾϾ jejunum). Northern blot analysis revealed three mRNA hybridizing species encoding SERT (3.0, 4.9, and 6.8 kb) in the human ileum. Consistent with SERT mRNA expression, SERT immunostaining was mainly detected in the epithelial cells of human duodenal and ileal resected tissues. Notably, SERT expression was localized predominantly to the apical and intracellular compartments and was distributed throughout the crypt-villus axis. Immunoblotting studies detected a prominent protein band (ϳ70 kDa) in the ileal apical plasma membrane vesicles (AMVs) isolated from mucosa obtained from organ-donor intestine. Functional studies showed that uptake of [ 3 H]serotonin (150 nM) in human ileal AMVs was 1) significantly increased in the presence of both Na ϩ and Cl Ϫ ; 2) inhibited (ϳ50%) by the neuronal SERT inhibitor, fluoxetine (10 M) and by unlabeled 5-HT; and 3) exhibited saturation kinetics indicating the presence of a carrier-mediated process. Our studies demonstrated differential expression of SERT across various regions of the human intestine and provide evidence for the existence of a functional SERT capable of removing intraluminal serotonin in human ileal epithelial cells. serotonin uptake; serotonin transporter expression SEROTONIN (5-hydroxytryptamine, 5-HT) is a neurotransmitter and hormone that influences diverse physiological functions. About 90% of the whole body content of 5-HT is present in the gastrointestinal (GI) tract, where it plays a critical role in modulation of gut motility and fluid and electrolyte transport (10,12,28,30). The majority of 5-HT in the GI tract is synthesized and stored in secretory granules of mucosal enterochromaffin (EC) cells. EC cells continuously release small amounts of 5-HT in response to a number of chemical and mechanical stimuli (13). Because 5-HT exerts its effects via specific 5-HT receptor subtypes, a mechanism for 5-HT deactivation is necessary to prevent the receptors from desensitization. The enzymes that catabolize 5-HT, including monoamine oxidases and glucuronyl transferases, are localized intracellularly and thus require transport of 5-HT across plasma membranes (5, 31). However, 5-...

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Serotonin uptake by bovine pulmonary artery endothelial cells in culture. I. Characterization

Cited by 59 publications

References 13 publications

Role of Protein Transamidation in Serotonin-Induced Proliferation and Migration of Pulmonary Artery Smooth Muscle Cells

Role of Protein Transamidation in Serotonin-Induced Proliferation and Migration of Pulmonary Artery Smooth Muscle Cells

Effects of monoamine uptake inhibitors on extracellular and platelet 5‐hydroxytryptamine in rat blood: different effects of clomipramine and fluoxetine

Function, expression, and characterization of the serotonin transporter in the native human intestine

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