The Role of Two-Pore Channels in Norepinephrine-Induced [Ca2+]i Rise in Rat Aortic Smooth Muscle Cells and Aorta Contraction

Cells

IrinaL. Zharkikh¹  AlexandraA. Tsitrina²  PavelV. Avdonin²  ElenaYu. Rybakova²  SergeiK. Trufanov²  PiotrP. Avdonin²  RichardO. Jenkins³  NikolayV. Goncharov⁴ 

[1] Institute of General Pathology and Pathophysiology RAMS, Moscow 125315, Russia;Koltsov Institute of Developmental Biology RAS, Moscow 119334, Russia;Leicester School of Allied Health Sciences, De Montfort University, The Gateway, Leicester LE1 9BH, UK;Sechenov Institute of Evolutionary Physiology and Biochemistry RAS, Saint Petersburg 194223, Russia;

关键词: norepinephrine;    calcium;    two-pore channels;    smooth muscle cells;    ned 19;    naadp;    sirna;    aorta;   

DOI  :  10.3390/cells8101144

来源: DOAJ

【 摘 要 】

Second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) triggers Ca²⁺ release via two-pore channels (TPCs) localized in endolysosomal vesicles. The aim of the present work is to evaluate the role of TPCs in the action of norepinephrine (NE), angiotensin II (AngII), vasopressin (AVP), and 5-hydroxytriptamine (5-HT) on free cytoplasmic calcium concentration ([Ca²⁺]_i) in smooth muscle cells (SMCs) isolated from rat aorta and on aorta contraction. To address this issue, the NAADP structural analogue and inhibitor of TPCs, NED 19, was applied. We have demonstrated a high degree of colocalization of the fluorescent signals of cis-NED 19 and endolysosmal probe LysoTracker in SMCs. Both cis- or trans-NED 19 inhibited the rise of [Ca²⁺]_i in SMCs induced by 100 μM NE by 50−60%. IC₅₀ for cis- and trans-NED 19 were 2.7 and 8.9 μM, respectively. The inhibition by NED 19 stereoisomers of the effects of AngII, AVP, and 5-HT was much weaker. Both forms of NED 19 caused relaxation of aortic rings preconstricted by NE, with relative potency of cis-NED 19 several times higher than that of trans-NED 19. Inhibition by cis-NED 19 of NE-induced contraction was maintained after intensive washing and slowly reversed within an hour of incubation. Cis- and trans-NED 19 did not cause decrease in the force of aorta contraction in response to Ang II and AVP, and only slightly relaxed aorta preconstricted by 5-HT and by KCl. Suppression of TPC1 in SMCs with siRNA caused a 40% decrease in [Ca²⁺]_i in response to NE, whereas siRNA against TPC2 did not change NE calcium signaling. These data suggest that TPC1 is involved in the NE-stimulated [Ca²⁺]_i rise in SMCs. Inhibition of TPC1 activity by NED 19 could be the reason for partial inhibition of aortic rings contraction in response to NE.

【 授权许可】

Unknown