Mechanisms of vasoconstriction induced by endothelin-1 in smooth muscle of rabbit mesenteric artery.
1. The mechanism underlying the vasoconstriction induced by endothelin-1 (ET-1) was investigated by measuring the intracellular concentration of Ca2+ ([Ca2+]i), isometric force and phosphorylation of the myosin light chain (MLC) in endothelium-denuded unskinned and beta-escin-treated skinned smooth muscle from resistance vessels of the rabbit mesentery. The role of protein kinase C (PKC) in the action of ET-1 was studied in skinned smooth muscle using a synthetic peptide, PKC19-36, which corresponds to the autoinhibitory domain of PKC. 2. ET-1 (> 0.1 nM) induced slowly developing, maintained increases in [Ca2+]i and force. Nicardipine completely blocked the ET-1-induced increase in [Ca2+]i. BQ-123 (an inhibitor of the ETA receptor) blocked the ET-1-induced contraction but IRL 1620 (Suc-[Glu9,Ala11,15]-ET-1(8-21), an agonist of the ETB receptor) failed to induce contraction. 3. In ionomycin- and 70 mM K(+)-treated strips, ET-1 shifted the [Ca2+]i-force relationship to the left and enhanced the maximum amplitude of contraction induced by 2.6 mM Ca2+. In skinned smooth muscle treated with ionomycin, Ca2+ (0.1-3 microM) increased both force and MLC phosphorylation, in a concentration-dependent manner. ET-1 with GTP shifted both the Ca(2+)-force and Ca(2+)-MLC phosphorylation relationships to the left without significant changes in the maximum responses. ET-1 with GTP did not change the relationship between force and MLC phosphorylation. Similar effects were observed with phorbol 12,13-dibutyrate (PDBu, an activator of PKC). These results indicate that the sensitivity of MLC phosphorylation to Ca2+ is enhanced both by ET-1 with GTP and by PDBu. 4. PKC19-36 (an inhibitor of PKC) modified neither the contraction nor MLC phosphorylation induced by 0.3 microM Ca2+ but blocked the PDBu-induced enhancement of both these Ca(2+)-induced responses. However, PKC19-36 only partly inhibited the enhancement produced by ET-1 with GTP on the Ca(2+)-induced responses. PKC19-36 did not modify the relationship between force and MLC phosphorylation in the presence either of ET-1 with GTP or of PDBu. By contrast, BQ-123, neomycin and guanosine 5'-O-(2-thiodiphosphate) (GDP beta S) each abolished the ET-1-induced enhancement of the contraction induced by 0.3 microM Ca2+. 5. These results suggest that ET-1 acts on the ETA receptor and increases Ca2+ influxes through an activation of the dihydropyridine-sensitive Ca2+ channel, causing a long-lasting and maintained contraction in resistance vessels of the rabbit mesentery.(ABSTRACT TRUNCATED AT 400 WORDS)
Yoshida
M
M
Department of Pharmacology, Faculty of Medicine, Kyushu University, Fukuoka, Japan.
Suzuki
A
A
Itoh
T
T
D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D003285:Contractile Proteins / Q000378:metabolism; D004095:Dihydropyridines / Q000494:pharmacology; D016232:Endothelins / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism; D066298:In Vitro Techniques; D015640:Ion Channel Gating; D008297:Male; D008638:Mesenteric Arteries / Q000187:drug effects / Q000502:physiology; D009119:Muscle Contraction / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D009218:Myosins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism; D011817:Rabbits; D017466:Receptors, Endothelin / Q000502:physiology*; D014661:Vasoconstriction / Q000187:drug effects*
D015220:Calcium Channels; D003285:Contractile Proteins; D004095:Dihydropyridines; D016232:Endothelins; D017466:Receptors, Endothelin; C038806:1,4-dihydropyridine; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D009218:Myosins; D002118:Calcium