Evidence that the substance P-induced enhancement of pacemaking in lymphatics of the guinea-pig mesentery occurs through endothelial release of thromboxane A2 |
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Authors: | Sharyn E Rayner Dirk F Van Helden |
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Institution: | The Neuroscience Group, Discipline of Human Physiology, Faculty of Medicine and Health Sciences, University of Newcastle, Callaghan, NSW 2308, Australia |
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Abstract: | - In vitro studies were performed to examine the mechanisms underlying substance P-induced enhancement of constriction rate in guinea-pig mesenteric lymphatic vessels.
- Substance P caused an endothelium-dependent increase in lymphatic constriction frequency which was first significant at a concentration of 1 nM (115±3% of control, n=11) with 1 μM, the highest concentration tested, increasing the rate to 153±4% of control (n=9).
- Repetitive 5 min applications of substance P (1 μM) caused tachyphylaxis with tissue responsiveness tending to decrease (by an average of 23%) and significantly decreasing (by 72%) for application at intervals of 30 and 10 min, respectively.
- The competitive antagonist of tachykinin receptors, spantide (5 μM) and the specific NK1 receptor antagonist, WIN51708 (10 μM) both prevented the enhancement of constriction rate induced by 1 μM substance P.
- Endothelial cells loaded with the Ca2+ sensing fluophore, fluo 3/AM did not display a detectable change in [Ca2+]i upon application of 1 μM substance P.
- Inhibition of nitric oxide synthase by NG nitro-L-arginine (L-NOARG; 100 μM) had no significant effect on the response induced by 1 μM substance P.
- The enhancement of constriction rate induced by 1 μM substance P was prevented by the cyclo-oxygenase inhibitor, indomethacin (3 μM), the thromboxane A2 synthase inhibitor, imidazole (50 μM), and the thromboxane A2 receptor antagonist, SQ29548 (0.3 μM).
- The stable analogue of thromboxane A2, U46619 (0.1 μM) significantly increased the constriction rate of lymphangions with or without endothelium, an effect which was prevented by SQ29548 (0.3 μM).
- Treatment with pertussis toxin (PTx; 100 ng ml−1) completely abolished the response to 1 μM substance P without inhibiting either the perfusion-induced constriction or the U46619-induced enhancement of constriction rate.
- Application of the phospholipase A2 inhibitor, antiflammin-1 (1 nM) prevented the enhancement of lymphatic pumping induced by substance P (1 μM), without inhibiting the response to either U46619 (0.1 μM) or acetylcholine (10 μM).
- The data support the hypothesis that the substance P-induced increase in pumping rate is mediated via the endothelium through NK1 receptors coupled by a PTx sensitive G-protein to phospholipase A2 and resulting in generation of the arachidonic acid metabolite, thromboxane A2, this serving as the diffusible activator.
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Keywords: | Lymphatic vessels endothelium constriction rate pacemaking substance P thromboxane A2 arachidonic acid calcium pertussis toxin NK1 receptor |
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