title: Ca2+ sensitization due to myosin light chain phosphatase inhibition and cytoskeletal reorganization in the myogenic response of skeletal muscle resistance arteries.
abstract: Abstract  The myogenic response of resistance arteries to intravascular pressure elevation is a fundamental physiological mechanism of crucial importance for blood pressure regulation and organ-specific control of blood flow. The importance of Ca(2+) entry via voltage-gated Ca(2+) channels leading to phosphorylation of the 20 kDa myosin regulatory light chains (LC20) in the myogenic response is well established. Recent studies, however, have suggested a role for Ca(2+) sensitization via activation of the RhoA/Rho-associated kinase (ROK) pathway in the myogenic response. The possibility that enhanced actin polymerization is also involved in myogenic vasoconstriction has been suggested. Here, we have used pressurized resistance arteries from rat gracilis and cremaster skeletal muscles to assess the contribution to myogenic constriction of Ca(2+) sensitization due to: (1) phosphorylation of the myosin targeting subunit of myosin light chain phosphatase (MYPT1) by ROK; (2) phosphorylation of the 17 kDa protein kinase C (PKC)-potentiated protein phosphatase 1 inhibitor protein (CPI-17) by PKC; and (3) dynamic reorganization of the actin cytoskeleton evoked by ROK and PKC. Arterial diameter, MYPT1, CPI-17 and LC20 phosphorylation, and G-actin content were determined at varied intraluminal pressures ± H1152, GF109203X or latrunculin B to suppress ROK, PKC and actin polymerization, respectively. The myogenic response was associated with an increase in MYPT1 and LC20 phosphorylation that was blocked by H1152. No change in phospho-CPI-17 content was detected although the PKC inhibitor, GF109203X, suppressed myogenic constriction. Basal LC20 phosphorylation at 10 mmHg was high at ∼40%, increased to a maximal level of ∼55% at 80 mmHg, and exhibited no additional change on further pressurization to 120 and 140 mmHg. Myogenic constriction at 80 mmHg was associated with a decline in G-actin content by ∼65% that was blocked by inhibition of ROK or PKC. Taken together, our findings indicate that two mechanisms of Ca(2+) sensitization (ROK-mediated phosphorylation of MYPT1-T855 with augmentation of LC20 phosphorylation, and a ROK- and PKC-evoked increase in actin polymerization) contribute to force generation in the myogenic response of skeletal muscle arterioles.
authors.affiliation: The Smooth Muscle Research Group, Department of Physiology and Pharmacology, Libin Cardiovascular Institute and Hotchkiss Brain Institute, Faculty of Medicine, University of Calgary, 3330 Hospital Drive N.W., Calgary, Alberta, Canada T2N 4N1.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Moreno-Domínguez
authors.forename: Olaia
authors.initials: O
authors.lastname: Colinas
authors.forename: Ahmed
authors.initials: A
authors.lastname: El-Yazbi
authors.forename: Emma J
authors.initials: EJ
authors.lastname: Walsh
authors.forename: Michael A
authors.initials: MA
authors.lastname: Hill
authors.forename: Michael P
authors.initials: MP
authors.lastname: Walsh
authors.forename: William C
authors.initials: WC
authors.lastname: Cole
mesh_terms: D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D062186:Arterial Pressure; D001158:Arteries / Q000201:enzymology; D020013:Calcium Signaling; D008297:Male; D040542:Mechanotransduction, Cellular; D009124:Muscle Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000098:blood supply*; D018994:Myosin Light Chains / Q000378:metabolism; D043263:Myosin-Light-Chain Phosphatase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D054645:Protein Phosphatase 1 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors; D014655:Vascular Resistance / Q000187:drug effects; D014661:Vasoconstriction / Q000187:drug effects; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D009124:Muscle Proteins; D018994:Myosin Light Chains; D010750:Phosphoproteins; C491519:Ppp1r14a protein, rat; D047428:Protein Kinase Inhibitors; D054460:rho-Associated Kinases; D011493:Protein Kinase C; C499850:Ppp1r12a protein, rat; D054645:Protein Phosphatase 1; D043263:Myosin-Light-Chain Phosphatase