endothelin-1 and ethylisopropylamiloride

We evaluated the effects of 5-(N-ethyl-N-isopropyl) amiloride (EIPA), a Na+/H+ exchanger (NHE) inhibitor, on ischemia/reperfusion (I/R)-induced acute renal failure (ARF) in mice. Ischemic ARF was induced by clamping the left renal artery and vein for 40 minutes followed by reperfusion 2 weeks after the contralateral nephrectomy. Preischemic treatment with EIPA attenuated the I/R-induced renal dysfunction. Histopathological examination of the kidney of ARF mice revealed severe renal damage such as tubular necrosis and proteinaceous casts in the tubuli. Histologically evident damage was also improved by preischemic treatment with EIPA. In addition, the I/R-induced increase in renal endothelin-1 (ET-1) content was suppressed by preischemic treatment with EIPA, reflecting the difference in immunohistochemical ET-1 localization in necrotic tubular epithelium. However, the postischemic treatment with EIPA failed to improve the I/R-induced renal dysfunction and ET-1 overproduction. These findings suggest that NHE activation, followed by renal ET-1 overproduction, plays an important role in the pathogenesis of I/R-induced renal injury. The inhibition of NHE by EIPA may be considered as a therapeutic approach to protect the postischemic ARF.

Topics: Acute Kidney Injury; Amiloride; Animals; Endothelin-1; Kidney; Kidney Tubular Necrosis, Acute; Kidney Tubules; Male; Mice; Protective Agents; Reperfusion Injury; Sodium-Hydrogen Exchangers

Endothelin (ET)-1 and norepinephrine (NE) are involved in myocardial ischemia/reperfusion injury. We investigated the role of ET-1 in ischemia/reperfusion-induced NE overflow and cardiac dysfunction using a selective ET(A) receptor antagonist (ABT-627), a selective ET(B) receptor antagonist (A-192621), and the spotting lethal (sl) rat, which carries a naturally occurring deletion in the ET(B) receptor gene.. According to the Langendorff technique, isolated hearts were subjected to 40-minute global ischemia followed by 30-minute reperfusion. In Sprague-Dawley rat hearts, ischemia/reperfusion-induced cardiac dysfunctions such as decreased left ventricular developed pressure and coronary flow and increased left ventricular end-diastolic pressure were worsened by treatment with A-192621. This agent enhanced excessive NE overflow in the coronary effluent from the postischemic heart. In contrast, treatment with ABT-627, in the absence or presence of A-192621, significantly improved postischemic cardiac dysfunction and markedly suppressed NE overflow to the same extent. Postischemic cardiac dysfunction and NE overflow in the heart of ET(B) receptor-deficient homozygous (sl/sl) rats were highly observed compared with cases in wild-type rats, and exaggerated responses to ischemia/reperfusion in sl/sl rats were abolished by ABT-627 treatment. Exogenously applied ET-1 produced severe cardiac dysfunction and a significant increase in NE overflow in a dose-dependent manner, but these responses were markedly suppressed in the presence of 5-N-ethyl-N-isopropyl-amiloride, an inhibitor of the Na+/H+ exchanger (NHE).. Pharmacological blockade or genetic deficiency of ET(B) receptors is detrimental to the postischemic heart, and exaggerated cardiac pathology under the above conditions is mediated by ET(A) receptor activation. ET(A)/NHE-mediated excessive NE overflow is contributive, at least in part, to postischemic cardiac dysfunction in rats.

Topics: Amiloride; Animals; Animals, Genetically Modified; Atrasentan; Endothelin A Receptor Antagonists; Endothelin B Receptor Antagonists; Endothelin-1; Gene Deletion; In Vitro Techniques; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Norepinephrine; Pyrrolidines; Rats; Rats, Sprague-Dawley; Receptor, Endothelin A; Receptor, Endothelin B; Regional Blood Flow; Ventricular Pressure

Chronic hypokalemia increases NHE3 activity in OKP cells. The aim of the present study was to determine whether an autocrine mechanism is involved in this activation.. After incubation of OKP cells in normal-K(+) and low-K(+) media for 24 h, the potassium concentration in the low-K(+) media was adjusted to a normal level. These conditioned media were then used as the normal-K(+) and low-K(+) supernatants. Other OKP cells were incubated in these normal-K(+) and low-K(+) supernatants and the mechanism of Na(+)/H(+) antiporter activation was examined.. The EIPA-resistant Na(+)/H(+) antiporter activity of OKP cells increased after 4 h incubation in the low-K(+) supernatant, and the amount of NHE3 protein increased at 24 h. Since both BQ788 and saralasin blocked this antiporter activation, the supernatant concentration of endothelin I (ET-I) and angiotensin II (Ang-II) were measured. The ET-I concentration was reduced, but the Ang-II concentration remained unchanged. There was a significant association between a reduction in the ET-I concentration and an increase in Na(+)/H(+) antiporter activity, but only when Ang-II was present in the supernatant.. An autocrine mechanism is involved in the activation of NHE3 in OKP cells. Both ET-I and Ang-II play a role in this activation.

Topics: Acidosis; Amiloride; Angiotensin II; Animals; Autocrine Communication; Benzoquinones; Cell Line; Culture Media, Conditioned; Dose-Response Relationship, Drug; Endothelin B Receptor Antagonists; Endothelin-1; Hydrogen-Ion Concentration; Kidney; Lactams, Macrocyclic; Oligopeptides; Opossums; Piperidines; Potassium; Protein-Tyrosine Kinases; Quinones; Receptor, Endothelin B; Rifabutin; Sodium-Hydrogen Exchanger 3; Sodium-Hydrogen Exchangers; Time Factors

The positive inotropic effect of endothelin-1 (ET-1) on normal myocardial contraction may be altered in pathological states. The purpose of this study was to assess the direct effect of ET-1 on cardiomyocyte performance and its cellular mechanism in congestive heart failure (CHF).. We measured the plasma levels of ET-1 and compared the effects of ET-1 (10(-10)-10(-8) M) on contractile performance and the [Ca2+]i transient in the myocytes of left ventricles (LV) from 15 age-matched normal adult rats and 15 rats with isoproterenol (ISO)-induced CHF.. With CHF, the plasma levels of ET-1 (19.7 +/- 6.3 vs. 4.1 +/- 0.5 fmol/ml, p < 0.05) were markedly elevated. In normal myocytes, superfusion of ET-1 caused significant increases in the systolic amplitude (SA, 8-16%) and the peak velocity of shortening (dL/dtmax, 20-35%; p < 0.01) without causing a change in the peak [Ca2+]i transient. In contrast, in myocytes from CHF rats, ET-1 produced significant reductions in SA (9-13%) and in the velocity of relengthening, dR/dtmax (10-14%; p < 0.05). The myocytes' dR/dtmax also decreased by 8-10% (p < 0.05). These changes were associated with a significant decrease in the peak [Ca2+]i transient (20-23%, p < 0.01). These responses to ET-1 were abolished by the incubation of myocytes with an ETA receptor antagonist (BQ123) or a protein kinase C (PKC) inhibitor (H-7 or staurosporine).. ISO-induced CHF is associated with elevated plasma ET-1 and an altered cardiomyocyte response to ET-1. After CHF, ET-1 produces a direct depression of cardiomyocyte contractile performance that is associated with a significant decrease in the peak [Ca2+]i transient. These effects are likely to be mediated through ETA receptors and involve the PKC pathway.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Amiloride; Animals; Anti-Arrhythmia Agents; Calcium; Cardiotonic Agents; Cell Size; Cells, Cultured; Endothelin Receptor Antagonists; Endothelin-1; Enzyme Inhibitors; Heart Failure; Hemodynamics; Isoproterenol; Male; Myocardial Contraction; Myocardium; Peptides, Cyclic; Protein Kinase C; Rats; Rats, Sprague-Dawley; Receptor, Endothelin A; Receptors, Endothelin; Sodium-Hydrogen Exchangers; Staurosporine

This report describes the fractional separation of microvessels from human brain for establishment of segmentally derived endothelial cell (EC) cultures. The investigation comprised evaluation of media constituents and purity of the cell culture and focused on functional biochemical characterization of endothelium derived from large microvessels (EC) Cells contained endothelial marker factor VIII (von Willebrand antigen), secreted endothelin-1 (ET-1) and prostaglandins, and took up 86Rb+ as a measure of K+. Exogenous ET-1 stimulated phosphatidylinositol hydrolysis and K+ uptake; BQ-123 (selective ETA receptor antagonist) but not IRL-1038 or BQ-788 (selective ETB receptor antagonists) inhibited both. Ouabain (inhibitor of Na(+)-K(+)-ATPase) and bumetanide (inhibitor of Na(+)-K(+)-Cl- cotransport) reduced (74-80 and 20-40%, respectively) the ET-1-stimulated K+ uptake. Staurosporine [protein kinase C (PKC) inhibitor] selectively reduced Na(+)-K(+)-Cl- cotransport, whereas verapamil but not nifedipine (L-type voltage-dependent Ca2+ channel blockers) decreased Na(+)-K(+)-ATPase activity induced by ET-1. Phorbol 12-myristate 13-acetate (PMA; activator of PKC) stimulated K+ uptake, which was only decreased with bumetanide. N-ethylisopropylamiloride (inhibitor of Na+/H+ exchange) reduced the ET-1-stimulated but not the PMA-induced K+ uptake. Results indicate that phosphatidylinositol hydrolysis and ion transport systems in large microvascular EC are stimulated by ET-1 through activation of ETA receptors. The findings also suggest that the ET-1-stimulated Na(+)-K(+)-ATPase activity, in contrast to Na(+)-K(+)-Cl- cotransport, is not mediated by PKC. In addition, the data suggest a linkage between Na(+)-K(+)-ATPase activity and Na+/H+ exchange.

Topics: Amiloride; Carrier Proteins; Cell Division; Cells, Cultured; Cerebrovascular Circulation; Endothelin-1; Endothelium, Vascular; Enzyme Inhibitors; Humans; Inositol 1,4,5-Trisphosphate; Microcirculation; Ouabain; Potassium; Sodium-Potassium-Chloride Symporters; Sodium-Potassium-Exchanging ATPase; Staurosporine; Tetradecanoylphorbol Acetate; Thymidine