F‐actin stabilization increases tension cost during contraction of permeabilized airway smooth muscle in dogs

Abstract

Dynamic actin reorganization involving actin polymerization and depolymerization may play an important functional role in smooth muscle. This study tested the hypothesis that F‐actin stabilization by phalloidin increases tension cost (i.e. ATP hydrolysis rate per unit of isometric force) during Ca2+‐induced activation of Triton X‐100‐permeabilized canine tracheal smooth muscle. Adenosine 5′‐triphosphate (ATP) hydrolysis rate was quantified using an enzyme‐coupled NADH fluorometric technique, regulatory myosin light chain (rMLC) phosphorylation was measured by Western blot analysis, and maximum unloaded shortening velocity (Vmax) was estimated by interpolation of the force‐velocity relationship to zero load during isotonic loading. Maximal activation with 10 μm free Ca2+ induced sustained increases in isometric force, stiffness, and rMLC phosphorylation. However, the increase in ATP hydrolysis rate initially reached peak values, but then declined to steady‐state levels above that of the unstimulated muscle. Thus, tension cost decreased throughout steady‐state isometric force. Following incubation of permeabilized strips with 50 μm phalloidin for 1 h, the increases in isometric force and stiffness were not sustained despite a sustained increase in rMLC phosphorylation. Also, after an initial decline, tension cost increased throughout activation. Phalloidin had no effect on Vmax during steady‐state isometric force or on rMLC phosphorylation. These findings suggest that dynamic reorganization of actin is necessary for optimal energy utilization during contraction of permeabilized airway smooth muscle.

Integrin-linked Kinase Regulates N-WASp-mediated Actin Polymerization and Tension Development in Tracheal Smooth Muscle

“…Previous studies of smooth muscle cells and tissues have shown that stimulation with contractile agonists initiates the polymerization of actin, and that this actin polymerization is required for tension development (6,7,9,12,16). Our studies suggest that this pool of actin is relatively small in differentiated muscle tracheal muscle tissues, probably less than 15% of the total pool of cellular actin (9,38).…”

Actin cytoskeletal dynamics in smooth muscle: a new paradigm for the regulation of smooth muscle contraction

“…The contraction of smooth muscle tissues and cells causes an increase in the pool of filamentous (F) actin and a decrease in the pool of monomeric (globular) (G) actin (12,27,48,65,68,72,84,120,130,143). These transitions in the state of actin have been documented in diverse smooth muscle cell and tissue types using a number of different approaches to estimate monomeric and filamentous pools of actin.…”

“…These transitions in the state of actin have been documented in diverse smooth muscle cell and tissue types using a number of different approaches to estimate monomeric and filamentous pools of actin. These include the use of DNase inhibition assays to measure the amount of globular (G) actin (84), measurements of soluble (G) actin and insoluble (F) actin by cell fractionation (104,120,127,143), fluorescence imaging to visualize G-and F-actin in isolated smooth muscle cells or tissues using G-and F-actin-specific stains (27,48,68,72,130), assessments of the rate of ADP-ATP actin monomer exchange (12), and electron microscopic studies that quantify actin filament density (65). All of these approaches have consistently shown that an increase in F-actin and a decrease in G-actin occurs when smooth muscle cells or tissues are activated by a contractile stimulus.…”

The Plastic Nature of the Vascular Wall: A Continuum of Remodeling Events Contributing to Control of Arteriolar Diameter and Structure

“…Studies performed primarily in tracheal smooth muscle indicate that remodeling of the actin cytoskeleton is required for the full development of force induced by stimulation with contractile agonists (94,113,183). The process involves increased polymerization of actin, tyrosine phosphorylation of paxillin, the activation of the small GTP-binding proteins Rho and Cdc42, and conformational changes within focal adhesion sites involving specific focal adhesion proteins (161,164,180,181).…”