Phorbol ester-induced stress and myosin light chain phosphorylation in swine carotid medial smooth muscle.
The mechanisms by which activators of protein kinase C (PKC) stimulate contractile responses in arterial smooth muscle is not known. In this study, we assessed the relative contribution of CA(++)-dependent and independent pathways in mediating phorbol ester-induced 20 kdalton myosin light chain (MLC)-phosphorylation and force in medial smooth muscle strips from swine carotid artery. Phorbol 12,13-dibutyrate (PDB; 10(-7)M)-stimulated stress development was associated with a significant increase in the fraction of phosphorylated MLC, from 0.08 +/- 0.02 to 0.24 +/- 0.02 after 30 min of stimulation. Under conditions of Ca++ depletion, which normally do not support Ca++/calmodulin-dependent activation of myosin light chain kinase (MLCK) by physiological stimuli, PDB-induced contractile responses were reduced significantly. However, after Ca2++ depletion, PDB (10(-6) M; 30 min) still caused an increase in MLC-phosphorylation from 0.10 +/- 0.02 at rest to 0.19 +/- 0.03. Preincubation with nifedipine (10(-7) M) had no significant effect on contractile responses to PDB, indicating that Ca++ influx through nifedipine-sensitive voltage channels did not contribute significantly to the observed Ca++ dependency of the PDB responses. Staurosporine (0.1-0.3 microM), a putative PKC inhibitor, significantly inhibited PDB-induced contractile and MLC phosphorylation responses. Tonic histamine (3 microM)- and KCl-induced contractile and MLC-phosphorylation responses were inhibited by the same concentrations of staurosporine.(ABSTRACT TRUNCATED AT 250 WORDS)
Singer
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Weis Center for Research, Geisinger Clinic, Danville, Pennsylvania.
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D004347:Drug Interactions; D004789:Enzyme Activation / Q000187:drug effects; D006632:Histamine / Q000494:pharmacology; D066298:In Vitro Techniques; D009119:Muscle Contraction / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019311:Staurosporine; D013552:Swine
D000470:Alkaloids; D015240:Phorbol 12,13-Dibutyrate; D006632:Histamine; D011493:Protein Kinase C; D009219:Myosin-Light-Chain Kinase; D019311:Staurosporine; D009543:Nifedipine; D002118:Calcium