Endothelin increases myofilament Ca2+ sensitivity in alpha-toxin-permeabilized rabbit mesenteric artery.
This study was designed to investigate the mechanism of endothelin-1 (ET-1) contractions in Staphylococcus alpha-toxin-permeabilized vascular smooth muscle. Rabbit small mesenteric arteries permeabilized with alpha-toxin were mounted for isometric or isotonic force recording or were processed for determination of myosin light chain (MLC) phosphorylation levels. Addition of 100 nM ET-1 plus 10 microM GTP significantly enhanced myofilament Ca2+ sensitivity as compared with the addition of Ca2+ alone (EC50, 0.47 microM Ca2+ for Ca2+ alone and 0.13 microM Ca2+ for ET-1 plus (GTP). This enhanced sensitivity was reversed by GDP beta S. ET-1-induced contractions were relaxed at a constant [Ca2+] by the addition of 30 microM cAMP or cGMP, demonstrating a direct effect of the cyclic nucleotides on contractile regulation. Inhibition of protein kinase C activity by 100 nM staurosporine relaxed ET-1 plus GTP-induced contractions, and pretreatment with 40 microM chelerythrine inhibited the ET-1 plus GTP increase in force. At 0.32 microM Ca2+, steady-state levels of shortening velocity were not increased by ET-1 plus GTP, although steady-state levels of MLC phosphorylation were significantly enhanced. The ET-1-induced increase in MLC phosphorylation was not altered by changes in [Ca2+], whereas the shortening velocity was Ca2+ dependent, suggesting that the increase MLC phosphorylation level may be the result of protein kinase C, rather than MLC kinase, activation. These results are consistent with the hypothesis that ET-1 increases myofilament Ca2+ sensitivity by a G protein-dependent pathway and subsequent activation of protein kinase C.(ABSTRACT TRUNCATED AT 250 WORDS)
Nishimura
J
J
Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine.
Moreland
S
S
Ahn
H Y
HY
Kawase
T
T
Moreland
R S
RS
van Breemen
C
C
D008841:Actin Cytoskeleton / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000242:Cyclic AMP / Q000494:pharmacology; D006152:Cyclic GMP / Q000494:pharmacology; D016232:Endothelins / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D008638:Mesenteric Arteries / Q000378:metabolism*; D009119:Muscle Contraction / Q000187:drug effects*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D010539:Permeability; D011493:Protein Kinase C / Q000378:metabolism; D011817:Rabbits; D010738:Type C Phospholipases / Q000494:pharmacology*
D016232:Endothelins; D006160:Guanosine Triphosphate; D000242:Cyclic AMP; D011493:Protein Kinase C; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D006152:Cyclic GMP; D002118:Calcium