s6c-sarafotoxin and Reperfusion-Injury

Endothelin-1 is a vasoactive peptide produced by the vascular endothelium. It is one of the most potent endogenous vascular smooth muscle constrictors. Two subtypes of the endothelin receptor have been cloned and sequenced and denoted endothelin-A and endothelin-B. The aim of this study was to define the influence of cold ischemia on the production of endothelin-1 and on the endothelin receptors. Two different preservation techniques (cold storage only and cold storage with microperfusion with University of Wisconsin solution) also were compared. The study was performed in an in vitro bone perfusion model to isolate the vascular endothelium from blood components. The production of endothelin-1 by the bone vasculature was not altered after 24 hours of cold ischemia. No contractions were observed with S6c, a selective endothelin-B agonist, and this effect was not influenced by cold ischemia. The response mediated by the endothelin-A receptor was increased significantly, an effect that was not influenced by preservation with University of Wisconsin solution. This latter finding was the only significant alteration in the vascular function detected in the in vitro model after 24 hours of cold ischemia. With regard to the pharmacologic properties of endothelin-1, this mediated response could be implicated in the pathogenesis of vasospasm.

Topics: Animals; Blood Flow Velocity; Bone and Bones; Bone Transplantation; Cryopreservation; Dogs; Dose-Response Relationship, Drug; Endothelins; Endothelium, Vascular; Female; In Vitro Techniques; Ischemia; Male; Norepinephrine; Perfusion; Receptors, Endothelin; Regional Blood Flow; Reperfusion Injury; Tibia; Vascular Resistance; Vasoconstrictor Agents; Viper Venoms

Coronary constrictor actions of endothelin-1 (ET-1) are enhanced after myocardial ischemia/reperfusion (I/R), possibly owing to enhanced ETA-receptor-mediated constriction and/or loss of the opposing ETB-receptor-mediated vasodilatation. We examined the actions of ET-1, ET-2, and ET-3 and the selective ETB-receptor agonist sarafotoxin 6c (Sx6c) after I/R in perfused rat heart. To examine the effects of a loss of ETB-receptor-mediated vasodilatation on coronary constrictor responses to ET-1, we used repeated doses of Sx6c to desensitize ETB receptors. After I/R, the coronary constrictor effects of all three ETs were enhanced, whereas their initial vasodilator effects were inhibited. The pure coronary dilator effect of Sx6c observed in control hearts was also inhibited after I/R. After desensitization of ETB receptors, the coronary constrictor action of ET-1 was enhanced by an amount equivalent to the vasodilatation that had been lost. This enhancement of constriction was not as marked as that noted after I/R, suggesting that the enhanced coronary constrictor action of ET-1 after I/R is not simply due to loss of opposing ETB-receptor-mediated vasodilatation and that other mechanisms are involved. The most likely explanation is upregulation of functional ETA receptors after I/R because ETB-receptor stimulation did not cause coronary constriction in this preparation. The vasoconstrictor enhancement therefore is likely to be the combined effect of receptor upregulation and vasodilator loss.

Topics: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; Analysis of Variance; Animals; Bradykinin; Endothelins; Heart; In Vitro Techniques; Male; Phenylephrine; Rats; Rats, Wistar; Receptors, Endothelin; Reperfusion Injury; Vasoconstrictor Agents; Vasodilation; Viper Venoms

We assessed the role of endogenous endothelin (ET) in reperfusion injury using a novel nonpeptidic ET antagonist of ETA and ETB receptors: bosentan (Ro 47-0203). In preliminary experiments in nonischemic rat hearts, we confirmed that bosentan 10(-5) M could block the changes in coronary flow (CF) and release of prostacyclin induced by ET-1 and the ETB-selective agonist sarafotoxin S6c (SRTX S6c). Thereafter, we induced global ischemia in paced hearts by closing the perfusion line for 20 min; this was followed by reperfusion. In one group of rats, bosentan (10(-5) M) was added to the perfusion medium before ischemia. In preischemic conditions, bosentan slightly increased CF by 14% (p = 0.094) in nonpaced hearts and by 35% (p = 0.001) in paced hearts, but did not influence cardiac contractility. Global ischemia produced severe ischemic damage, as shown by electromechanical dissociation (EMD) and decreased cardiac contractility. Bosentan did not influence recovery of cardiac function and did not ameliorate postischemic hemodynamic variables as compared with those of the control group. We conclude that endogenous ET does not play a major role in induction of reperfusion injury in isolated perfused rat heart.

Topics: Animals; Bosentan; Coronary Circulation; Drug Interactions; Endothelin Receptor Antagonists; Endothelins; Epoprostenol; Heart; Hemodynamics; Male; Myocardial Ischemia; Myocardium; Rats; Rats, Wistar; Reperfusion Injury; Sulfonamides; Vasoconstrictor Agents; Viper Venoms

Previous work indicated that endothelin (ET) may be involved in the pathogenesis of myocardial ischemia, although the relative importance of the ET receptor subtypes is presently not clear. The purpose of this study was to determine the role of myocardial ET-B receptors in mediating ischemic/reperfusion damage in isolated rat hearts. Saturation binding analyses were conducted with [125I]ET-1 and [125I]IRL-1620 to assess changes in ET-A and ET-B receptor binding. Total ET receptor density (Bmax) was greater in atrial versus ventricular tissue. ET-A Bmax was 8 to 10-fold greater than ET-B Bmax. In ischemic and ischemic/reperfused atrial tissue neither the equilibrium dissociation constant (Kd) nor Bmax for ET-B receptors was changed. The ET-B receptor Kd in ischemic or ischemic/reperfused ventricular tissue was also unchanged. In ischemic ventricular tissue there was a trend towards an increased ET-B Bmax, which was accentuated after ischemia/reperfusion. No changes were found in ET-A Bmax or Kd in ischemic ventricular or atrial tissue. The physiological importance of this receptor subtype in ischemic myocardium was determined using the selective ET-B agonist, sarafotoxin S6c. In non-ischemic tissue no effect on coronary flow or function were observed with sarafotoxin S6c. Furthermore, no changes were seen in ischemic time to contracture or any of the reperfusion indexes of myocardial damage. The sarafotoxin S6c utilized was active as it inhibited [125I]ET-3 binding to ET-B receptors (Ki = 0.1 nM). Thus, the pro-ischemic effect of ET-1 seems to be mediated by ET-A receptors. ET-B receptors do not appear to play a role in the pathogenesis of myocardial ischemia.

Topics: Animals; Endothelins; Hemodynamics; Male; Myocardial Ischemia; Rats; Rats, Sprague-Dawley; Receptor, Endothelin B; Receptors, Endothelin; Reperfusion Injury; Viper Venoms