Effect of Docosahexaenoic Acid on Voltage-Independent Ca2+ Entry Pathways in Cultured Vascular Smooth Muscle Cells Stimulated with 5-Hydroxytryptamine

• Published in: Biological & pharmaceutical bulletin 2017/06/01, Vol.40(6), pp.916-921
• Main Authors: Machida, Takuji, Onoguchi, Akina, Iizuka, Kenji, Ishibashi, Sayuri, Yutani, Mikiko, Hirafuji, Masahiko
• Format: Article
• Language: English
• Subjects: 5-hydroxytryptamine; Calcium (extracellular); Calcium (intracellular); Calcium channels (voltage-gated); Calcium influx; Docosahexaenoic acid; Gene expression; Intracellular; Na+/Ca2+ exchanger; NCX1 protein; Polymerase chain reaction; Potassium channels (voltage-gated); receptor-operated Ca2+ entry; Rodents; Serotonin; Serotonin S2 receptors; Smooth muscle; store-operated Ca2+ entry; Transient receptor potential proteins; vascular smooth muscle cell
• ISSN: 0918-6158; 1347-5215
• DOI: 10.1248/bpb.b16-00788

We previously reported that docosahexaenoic acid (DHA) inhibits an increase in intracellular Ca2+ concentration ([Ca2+]i) in cultured rat vascular smooth muscle cells (VSMCs) through a mechanism involving mainly voltage-dependent Ca2+ channels; however, the effect of DHA on voltage-independent pathways, such as store-operated and receptor-operated Ca2+ entry, and Ca2+ entry through Na+/Ca2+ exchanger (NCX), has not been clarified. In the present study, we investigated the effect of DHA treatment on the expression of transient receptor potential canonical (TRPC) channels, capacitative Ca2+ entry, and Ca2+ entry through NCX in rat cultured VSMCs stimulated with 5-hydroxytryptamine (5-HT). RT-PCR analysis detected TRPC1, TRPC4, and TRPC6 mRNA in cultured VSMCs. DHA treatment for 2 d slightly but significantly decreased TRPC1, but not TRPC4 and TRPC6, mRNA expression. Sarpogrelate, a selective serotonin 5-HT2A receptor inhibitor, completely inhibited the 5-HT-induced increase in [Ca2+]i in cultured VSMCs. Ca2+ influx by adding extracellular Ca2+ (1.3 mM) to the Ca2+-free condition in the presence of 5-HT was partially but significantly inhibited by sarpogrelate. DHA treatment for 2 d had no effect on Ca2+ influx when extracellular Ca2+ was added to the Ca2+-free condition in the presence of either 5-HT alone or 5-HT with sarpogrelate. KB-R7943, a selective inhibitor of reverse mode NCX, significantly suppressed the 5-HT-induced increase of [Ca2+]i. Furthermore, DHA treatment for 2 d significantly decreased NCX1 mRNA expression. These results suggest that DHA seems to have little effect on capacitative Ca2+ entry. Through decreasing NCX1 expression, DHA may suppress the 5-HT-induced increase in [Ca2+]i.