Endothelin-1 and endothelin-3 regulate differently vasoconstrictor responses of smooth muscle of the porcine coronary artery.
1. Using front-surface fluorometry of fura-2 and medial strips of the porcine coronary artery, we investigated mechanisms by which endothelin-1 (ET-1) and ET-3 function as vasoconstrictors. 2. In the presence of extracellular Ca2+(1.25 mM), ET-1 (10(-10)-10(-7) M) increased cytosolic Ca2+ concentrations ([Ca2+]i) and tension, in a concentration-dependent manner. ET-1, at concentrations greater than 10(-8) M, induced an abrupt elevation of [Ca2+]i which reached a transient peak (the first component, [Ca2+]i-rising phase) and subsequently declined ([Ca2+]i-declining phase) to reach a lower sustained phase (the second component, steady-state phase), while the tension rose monotonically to reach a peak and then slightly and gradually declined. ET-1, at concentrations lower than 10(-8) M, induced slowly developing and sustained increases in [Ca2+]i and tension ([Ca2+]i-rising phase followed by steady-state phase). All concentrations of ET-1 increased tension more slowly than [Ca2+]i. 3. In the presence of extracellular Ca2+, ET-3 (10(-8)-10(-5) M) induced concentration-dependent increases in [Ca2+]i and tension. However, the maximal elevations of [Ca2+]i and tension induced by ET-3 were substantially smaller than those induced by ET-1, indicating the involvement of an ETA receptor subtype. ET-3, at concentrations greater than 6 x 10(-7) M, caused biphasic slowly developing increases in [Ca2+]i and tension. At concentrations lower than 10(-6) M, ET-3 caused monophasic increases in [Ca2+]i and tension. At all concentrations of ET-3, the time courses of increases in [Ca2+]i and tension were similar. 4. The biphasic increases in [Ca2+]i and tension induced by 10-5 M ET-3 and by 1O-7M ET-1 were significantly inhibited by pretreatment with 10-5 M of the Ca2+ entry blocker, diltiazem, although the inhibition of the first component of ET-l-induced [Ca2+]i increase was partial.5. In the absence of extracellular Ca2+, ET-1 induced a concentration-dependent transient increase in[Ca2+]i, possibly due to release of Ca2+ from intracellular stores, and a sustained contraction. In contrast, ET-3 ( 10-6 M) caused little, if any, transient increase in [Ca2+]i and a small sustained contraction.6. Temporal changes in the relationships between [Ca2+]i and tension ([Ca2+]1-tension relationship)during contractions induced by ET-1 and ET-3 were compared with the [Ca2+]i-tension relationship of Ca2+-induced contractions (Ca2+-contractions) obtained by cumulative applications of extracellular Ca2+(0-7.5 mM) to tissues depolarized in the presence of 118 mMK+. In the [Ca2+]i-rising phase, ET-1 increased tension more slowly than [Ca2+]i, thereby shifting the [Ca2+]i-tension relation to the right from that for Ca2+-contractions. In the [Ca2+I-declining and the steady-state phases, ET-1, at concentrations higher than 10-9 M, produced greater tension development than that expected from a given change in[Ca2+ji, resulting in a leftward shift of the [Ca2+]i-tension relation. During ET-3-induced contractions,([Ca2+]i-rising, [Ca2+]i-declining and steady-state phases), the [Ca2+]i-tension relation was similar to that of Ca2+-contractions.7. BQ-123, a selective ETA receptor antagonist, completely inhibited the increases in [Ca2+1]i and tension induced by ET-1 and ET-3.8. These results suggest: (1) That ET-1 elicits vasoconstriction by increasing [Ca2+]i through the activation of Ca2+ influx from the extracellular space and Ca2+ release from intracellular storage sites,and by increasing the Ca2+ sensitivity of the contractile apparatus, whereas ET-3 induces vasoconstriction by increasing [Ca2+1] mainly through Ca2+ influx from the extracellular space. (2) Distinct mechanisms of time-dependent modulation of the Ca2+ sensitivity function in the vasoconstrictor responses to ET-1 and ET-3. (3) That both ET-1- and ET-3-induced contractions seem to be mediated via ETA-receptors in porcine coronary artery, and that the ETA-receptor-mediated effects of ET-1 and ET-3 can be dissociated at the sub-receptor levels of the signal transduction pathway.
Ushio-Fukai
M
M
Division of Molecular Cardiology, Faculty of Medicine, Kyushu University, Fukuoka, Japan.
Nishimura
J
J
Kobayashi
S
S
Kanaide
H
H
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D003331:Coronary Vessels / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D016232:Endothelins / Q000494:pharmacology*; D005260:Female; D005453:Fluorescence; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects*; D014661:Vasoconstriction / Q000187:drug effects*
D016232:Endothelins; D002118:Calcium