The ACh-induced contraction in rat aortas is mediated by the Cys Lt1 receptor via intracellular calcium mobilization in smooth muscle cells.
1. Our previously published data indicate that an endogenously produced 5-lipoxygenase metabolite can strongly contract isolated endothelium-preserved rat aortic strips when cyclo-oxygenase isoenzymes are inhibited. Therefore, we decided to investigate if cysteinyl-containing leukotrienes (Cys Lts) are involved in this endothelium-dependent contraction. 2. The isometric contraction of endothelium-preserved rat aortic strips was recorded in preparations preincubated with 5 microM indomethacin and precontracted with phenylephrine, adjusting resting tension at 0.7 g. Acetylcholine (ACh) contracted control strips. Montelukast and MK-571, selective type 1 Cys Lts receptor (Cys Lt(1)) antagonists and the Cys Lt(1)/Cys Lt(2) (type 2 Cys Lts receptor) antagonist BAYu9773 dose-dependently prevented ACh-induced contraction, their IC(50)s being 2.2, 3.1 and 7.9 nM respectively. The leukotriene B4 receptor antagonist U75302 was far less potent (IC(50) 1.5 microM). 3. In rat aorta smooth muscle cells (RASMs), Western blot analysis showed the presence of Cys Lt(1) and Cys Lt(2) receptors, the Cys Lt(1) receptor being predominantly expressed. 4. In fura-2 loaded RASMs, LTD4 (0.01-100 nM) and LTC4 (200-800 nM) dose-dependently increased intracellular calcium concentration ([Ca(2+)](i)). Montelukast (1-100 nM) reduced LTD4-induced [Ca(2+)](i) increase, its IC(50) being approximately 10 nM. BAY u9773 exhibited significantly low effectiveness. 5. LTD4 (10 nM) induced a redistribution of smooth muscle actin fibres throughout the cytoplasm as visualized by confocal microscopy. 6. In conclusion, Cys Lt(1) activation by endogenously produced Cys Lts, can contract rat aortas, while Cys Lt(2) only marginally influences aortic tone. Intracellularly, this effect is mediated by an increase in [Ca(2+)](i). Therefore, Cys Lts, by inducing vascular contraction, can contribute to systemic hypertension.
Mazzetti
Luca
L
Department of Pharmacology, University of Florence, Viale Pieraccini, 6 50139 Florence, Italy.
Franchi-Micheli
Sergio
S
Nistri
Silvia
S
Quattrone
Silvia
S
Simone
Ruggero
R
Ciuffi
Mario
M
Zilletti
Lucilla
L
Failli
Paola
P
D000109:Acetylcholine / Q000494:pharmacology*; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D002118:Calcium / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D020024:Leukotriene Antagonists; D017998:Leukotriene D4 / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D018077:Receptors, Leukotriene / Q000502:physiology*; D014661:Vasoconstriction / Q000187:drug effects* / Q000502:physiology*
D020024:Leukotriene Antagonists; D008565:Membrane Proteins; D018077:Receptors, Leukotriene; D017998:Leukotriene D4; C412375:cysteinyl leukotriene receptor 2; C083169:leukotriene D4 receptor; D000109:Acetylcholine; D002118:Calcium