Contractile Responses and Myosin Phosphorylation in Reconstituted Fibers of Smooth Muscle Cells From the Rat Cerebral Artery

• Published in: Japanese Journal of Pharmacology 2002, Vol.90(1), pp.36-50
• Main Authors: Oishi, Kazuhiko, Takatoh, Yoshitaka, Bao, Jianjun, Uchida, Masaatsu K.
• Format: Article
• Language: English
• Subjects: Animals; Cell Culture Techniques - methods; Collagen gel fiber; Contractility; Cultured smooth muscle cell; Isometric force; Male; Middle cerebral artery; Middle Cerebral Artery - cytology; Middle Cerebral Artery - drug effects; Middle Cerebral Artery - metabolism; Muscle Contraction - drug effects; Muscle Contraction - physiology; Muscle Fibers, Skeletal - cytology; Muscle Fibers, Skeletal - drug effects; Muscle Fibers, Skeletal - metabolism; Muscle Proteins - biosynthesis; Myocytes, Smooth Muscle - cytology; Myocytes, Smooth Muscle - drug effects; Myocytes, Smooth Muscle - metabolism; Myosins - metabolism; Phosphorylation; Rats; Rats, Inbred WKY; Vasoconstriction - drug effects; Vasoconstriction - physiology
• ISSN: 0021-5198; 1347-3506
• DOI: 10.1254/jjp.90.36

String-shaped reconstituted smooth muscle fibers were prepared in rectangular wells by thermal gelation of a mixed solution of collagen and cultured smooth muscle cells derived from the rat cerebral artery. The fibers contracted in response to KCl, 5-hydroxytryptamine (5-HT), noradrenaline, endothelin-1, endothelin-2, angiotensin II, prostaglandin F2α and prostaglandin E2. 5-HT-induced contraction was partially inhibited by the L-type voltage-dependent Ca2+ channel inhibitor nifedipine, putative non-selective cationic channel inhibitor SKF96365 and intracellular Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N>′-tetraacetic acid acetoxymethyl ester (BAPTA-AM), and completely abolished by the myosin light chain kinase inhibitor ML-9. The fibers pre-contracted by 5-HT were completely relaxed by the Rho kinase inhibitor Y-27632, serine/threonine kinase inhibitor staurosporine, 8-bromo cyclic GMP and papaverine, and partially relaxed by dibutyryl cyclic AMP. Moreover, 5-HT as well as endothelin-1 and KCl enhanced 20-kDa myosin light chain phosphorylation in the fibers. These results suggested that the characteristics of contraction of the fibers reflect typical contractilities of vascular smooth muscle tissues. This technique will allow us to directly address questions relating to heterogeneity of receptor mechanisms and intracellular pathways of vascular smooth muscle contraction as a function of vessel type.