ET_A receptors are the primary mediators of myofilament calcium sensitization induced by ET‐1 in rat pulmonary artery smooth muscle: a tyrosine kinase independent pathway

Abstract

1 We have investigated the possibility that ET-1 can induce an increase in myo®lament calcium sensitivity in pulmonary artery smooth muscle. Arterial rings were permeabilized using a-toxin (120 mg ml 71 ), in the presence of A23187 (10 mM) to`knock out' Ca 2+ stores, and pre-constricted with pCa 6.8 (bu ered with 10 mM EGTA). In the presence of this ®xed Ca 2+ concentration, 1 mM ET-1 induced a sustained, reversible constriction of 0.15 mN. 2 Pulmonary arterial rings were freeze-clamped at the peak of the induced constriction (time matched). Subsequent densitometric analysis revealed that ET-1 (1 mM) increased the level of phosphorylated myosin light chains by 34% compared to an 11% increase in the presence of pCa 6.8 alone.3 In contrast to ET-1, the selective ET B receptor agonist Sarafotoxin S6C (100 nM) failed to induce a signi®cant constriction. 4 The constriction induced by 1 mM ET-1 was reversibly inhibited when the preparation was preincubated (15 min) with the ET A receptor antagonist BQ 123 (100 mM). The constriction measured 0.13 mN in the absence and 0.07 mN in the presence of 100 mM BQ 123. 5 In contrast, the constriction induced by 1 mM ET-1 measured 0.19 mN in the absence and 0.175 mN following a 15 min pre-incubation with the ET B antagonist BQ 788 (100 mM). 6 The constriction induced by 1 mM ET-1 measured 0.14 mN in the presence and 0.13 mN following pre-incubation with the tyrosine kinase inhibitor Tyrphostin A23 (100 mM). 7 We conclude that ET-1 induced an increase in myo®lament calcium sensitivity in rat pulmonary arteries via the activation of ET A receptors and by a mechanism(s) independent of tyrosine kinase.

Thrombin-Induced Activation of RhoA in Platelet Shape Change

“…Precipitated proteins were washed with acetone, resuspended in sample buffer, and subjected to 2-dimensional electrophoresis as previously described (19). Resolved proteins were transferred on to nitrocellulose and the membrane stained with colloidal gold solution (Biorad).…”

Increased superoxide levels in ganglia and sympathoexcitation are involved in sarafotoxin 6c-induced hypertension

“…This was surprising in light of the fact that the two best described physiological responses to ET B R activation, i.e., release of vasodilators from endothelial cells, and increased renal sodium and water excretion, should lead to a fall in arterial pressure (42,48,50). We believe venoconstriction contributes to S6c-induced hypertension, because in vitro S6c has been shown to constrict veins from most vascular regions but to have little effect on most arteries (15,27,41,49).It is likely, however, that other mechanisms contribute to S6c-induced hypertension. Reactive oxygen species (ROS) are a variety of oxygen-containing molecules that are by-products of cellular metabolic processes under physiological condition, including superoxide anion (O 2 Ϫ ), hydrogen peroxide (H 2 O 2 ), and hydroxyl ion (OH⅐).…”

Mitogen-Activated Protein Kinases Partially Regulate Endothelin-1-Induced Contractions through a Myosin Light Chain Phosphorylation- Independent Patyway.

“…The increase in the sensitivity of the contractile apparatus to intracellular Ca 2+ ([Ca 2+ ] i ), as well as the increase of [Ca 2+ ] i concentration, may be involved in ET-1-induced vasoconstriction [8,26,28,29]. On the other hand, ET-1 evoked an increase of myofilament Ca 2+ sensitivity by a mechanism(s) independent of tyrosine kinase in the rat pulmonary artery [10]. Moreover, ET-1 increased the phosphorylation of p38 mitogen-activated protein kinase (MAPK) in the canine pulmonary artery [37].…”