Involvement of Organic Cation Transporter-3 and Plasma Membrane Monoamine Transporter in Serotonin Uptake in Human Brain Vascular Smooth Muscle Cells

Rachel W S Li,S W Chan,Y W Kwan,Simon M Y Lee,George P H Leung,Cui Yang

doi:10.3389/fphar.2013.00014

Abstract

The serotonin (5-HT) uptake system is supposed to play a crucial part in vascular functions by “fine-tuning” the local concentration of 5-HT in the vicinity of 5-HT2 receptors in vascular smooth muscle cells. In this study, the mechanism of 5-HT uptake in human brain vascular smooth muscle cells (HBVSMCs) was investigated. [3H]5-HT uptake in HBVSMCs was Na+-independent. Kinetic analyses of [3H]5-HT uptake in HBVSMCs revealed a Km of 50.36 ± 10.2 mM and a Vmax of 1033.61 ± 98.86 pmol/mg protein/min. The specific serotonin re-uptake transporter (SERT) inhibitor citalopram, the specific norepinephrine transporter (NET) inhibitor desipramine, and the dopamine transporter (DAT) inhibitor GBR12935 inhibited 5-HT uptake in HBVSMCs with IC50 values of 97.03 ± 40.10, 10.49 ± 5.98, and 2.80 ± 1.04 μM, respectively. These IC50 values were 100-fold higher than data reported by other authors, suggesting that those inhibitors were not blocking their corresponding transporters. Reverse transcription-polymerase chain reaction results demonstrated the presence of mRNA for organic cation transporter (OCT)-3 and plasma membrane monoamine transporter (PMAT), but the absence of OCT-1, OCT-2, SERT, NET, and DAT. siRNA knockdown of OCT-3 and PMAT specifically attenuated 5-HT uptake in HBVSMCs. It is concluded that 5-HT uptake in HBVSMCs was mediated predominantly by a low-affinity and Na+-independent mechanism. The most probable candidates are OCT-3 and PMAT, but not the SERT.

Highlights

Serotonin [5-hydroxytryptamine (5-HT)] is a neurotransmitter in the central nervous system and digestive tract, and a potent vasoconstrictor
In the present study, we characterized the 5-HT uptake system in human brain vascular smooth muscle cells (HBVSMCs)
Similar to the findings in certain vascular beds such as mesenteric arteries, vena cava, and jugular vein (Wanstall et al, 2003; Linder et al, 2008a,b), serotonin re-uptake transporter (SERT) does not have a significant contribution to 5-HT uptake in HBVSMCs

Summary

Introduction

Serotonin [5-hydroxytryptamine (5-HT)] is a neurotransmitter in the central nervous system and digestive tract, and a potent vasoconstrictor. 5-HT is mainly stored in platelets, thereby maintaining a low level of free-circulating 5-HT. The released 5-HT feeds back on the platelets to amplify the aggregation process and causes the contraction of vascular smooth muscle cells through the stimulation of 5-HT2 receptors (Vanhoutte, 1990). Several studies have suggested that 5-HT may be involved in vascular diseases such as hypertension. Arterial contraction to 5-HT is profoundly enhanced in hypertension in animals and humans (Wyse, 1984; Dohi and Lüscher, 1991; HutriKähönen et al, 1999). An increased plasma level of 5-HT has been measured in various models of hypertension (Soares-da-Silva et al, 1995; Krygicz et al, 1996)

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Conclusion