Cross-bridge phosphorylation and regulation of latch state in smooth muscle

1988 ◽  
Vol 254 (1) ◽  
pp. C99-C106 ◽  
Author(s):  
C. M. Hai ◽  
R. A. Murphy
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Experimental Observation ◽  
Rate Constants ◽  
Regulatory Mechanism ◽  
Myosin Phosphorylation ◽  
Model Simulations ◽  
Cross Bridge ◽  
Cross Bridges ◽  
Latch State

We have developed a minimum kinetic model for cross-bridge interactions with the thin filament in smooth muscle. The model hypothesizes two types of cross-bridge interactions: 1) cycling phosphorylated cross bridges and 2) noncycling dephosphorylated cross bridges ("latch bridges"). The major assumptions are that 1) Ca2+-dependent myosin phosphorylation is the only postulated regulatory mechanism, 2) each myosin head acts independently, and 3) latch bridges are formed by dephosphorylation of an attached cross bridge. Rate constants were resolved by fitting data on the time courses of myosin phosphorylation and stress development. Comparison of the rate constants indicates that latch-bridge detachment is the rate-limiting step. Model simulations predicted a hyperbolic dependence of steady-state stress on myosin phosphorylation, which corresponded with the experimental observation of high values of stress with low levels of phosphorylation in intact tissues. Model simulations also predicted the experimental observation that an initial phosphorylation transient only accelerates stress development, with no effect on the final steady-state levels of stress. Because the only Ca2+-dependent regulatory mechanism in this model was activation of myosin light chain kinase, these results are consistent with the hypothesis that myosin phosphorylation is both necessary and sufficient for the development of the latch state.

Regulation of shortening velocity by cross-bridge phosphorylation in smooth muscle

1988 ◽  
Vol 255 (1) ◽  
pp. C86-C94 ◽  
Author(s):  
C. M. Hai ◽  
R. A. Murphy
Keyword(s):  
Smooth Muscle ◽  
Shortening Velocity ◽  
Internal Load ◽  
Cross Bridge ◽  
Bridge Model ◽  
The Family ◽  
Cross Bridges ◽  
Latch State ◽  
The Relationship ◽  
State Stress

We have proposed a model that incorporates a dephosphorylated "latch bridge" to explain the mechanics and energetics of smooth muscle. Cross-bridge phosphorylation is proposed as a prerequisite for cross-bridge attachment and rapid cycling. Features of the model are 1) myosin light chain kinase and phosphatase can act on both free and attached cross bridges, 2) dephosphorylation of an attached phosphorylated cross bridge produces a noncycling "latch bridge," and 3) latch bridges have a slow detachment rate. This model quantitatively predicts the latch state: stress maintenance with reduced phosphorylation, cross-bridge cycling rates, and ATP consumption. In this study, we adapted A. F. Huxley's formulation of crossbridge cycling (A. F. Huxley, Progr. Biophys. Mol. Biol. 7: 255-318, 1957) to the latch-bridge model to predict the relationship between isotonic shortening velocity and phosphorylation. The model successfully predicted the linear dependence of maximum shortening velocity at zero external load (V0) on phosphorylation, as well as the family of stress-velocity curves determined at different times during a contraction when phosphorylation values varied. The model implies that it is unnecessary to invoke an internal load or multiple regulatory mechanisms to explain regulation of V0 in smooth muscle.

Dependence of stress on cross-bridge phosphorylation in vascular smooth muscle

1989 ◽  
Vol 256 (1) ◽  
pp. C96-C100 ◽  
Author(s):  
P. H. Ratz ◽  
C. M. Hai ◽  
R. A. Murphy
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Vascular Smooth Muscle ◽  
Light Chain ◽  
Phorbol Esters ◽  
Receptor Activation ◽  
Bay K 8644 ◽  
Cross Bridge ◽  
Cross Bridges ◽  
State Stress

Cross-bridge phosphorylation associated with agonist-stimulated contraction of vascular smooth muscle is often transiently elevated. Such observations led to the concept that phosphorylation of the 20-kDa myosin regulatory light chain (Mp) was required for initial activation and cross-bridge cycling but might not be necessary for steady-state maintenance of stress in the latch state. The possibility that stress maintenance is not regulated by phosphorylation has received some experimental support in contractions induced by phorbol esters and the calcium channel activator BAY K 8644 in which significant increases in Mp were not detected. Our aim was to test the hypothesis that phosphorylation is both necessary and sufficient for activation and for maintenance of steady-state stress. Activation of swine carotid media using agents that bypass receptor activation and elevate Ca2+ influx without mobilizing intracellular Ca2+ stores (BAY K 8644 and ionomycin) produced monotonic increases in both stress and Mp. Transient initial peaks in Mp were absent. Steady-state stress induced by both receptor- and nonreceptor-mediated activation was dependent on small increases in Mp. Increases in Mp greater than 0.3 mol Pi/mol myosin light chain had small effects on stress but produced large increases in the maximum rate of cross-bridge cycling at zero load (Vo). The experimentally determined dependence of stress on Mp was quantitatively predicted by our working hypothesis. This model proposes that Ca2+-stimulated cross-bridge phosphorylation is obligatory for cross-bridge attachment. However, dephosphorylation of attached cross bridges to form noncycling "latch bridges" allows stress maintenance with reduced Mp and cycling.

Effects of substrate and inhibition of oxidative metabolism on contraction and myosin phosphorylation in ASM

1993 ◽  
Vol 264 (6) ◽  
pp. L553-L559 ◽  
Author(s):  
C. M. Hai ◽  
C. Watson ◽  
S. J. Wallach ◽  
V. Reyes ◽  
E. Kim ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Airway Smooth Muscle ◽  
Oxidative Metabolism ◽  
Energy Utilization ◽  
Atp Content ◽  
Active Stress ◽  
Myosin Phosphorylation ◽  
Intracellular Calcium Store ◽  
Cross Bridge

Steady-state active stress in smooth muscle is maintained by cross bridges which undergo continuous cycling and myosin phosphorylation, and the two processes both consume ATP. In this study, we investigated whether energy utilization by cross-bridge cycling and myosin phosphorylation is compartmentalized and examined their relative affinities for ATP in airway smooth muscle. We measured active stress, myosin phosphorylation, O2 consumption, and tissue ATP content in bovine tracheal smooth muscle activated by K+ depolarization when glucose was replaced by pyruvate and when oxidative metabolism was inhibited by hypoxia or uncoupled by 2,4-dinitrophenol. The results indicate that ATP produced from both glycolysis and oxidative metabolism is available to both cross-bridge cycling and myosin phosphorylation. However, steady-state myosin phosphorylation was insensitive to the inhibition of oxidative metabolism by hypoxia and mitochondrial uncoupling when steady-state isometric stress and tissue ATP content were significantly reduced. These results suggest that, relative to actomyosin adenosine 5'-triphosphatase, myosin light chain kinase has a higher affinity for ATP in intact airway smooth muscle. However, peak myosin phosphorylation associated with the initial rapid stress development was sensitive to inhibition of oxidative metabolism, probably reflecting a lower content of intracellular calcium store as a result of metabolic inhibition.

Agonist activation modulates cross-bridge states in single vascular smooth muscle cells

1993 ◽  
Vol 264 (1) ◽  
pp. C103-C108 ◽  
Author(s):  
F. V. Brozovich ◽  
M. Yamakawa
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Relative Population ◽  
Cross Bridge ◽  
Cross Bridges ◽  
State Force

To determine cross-bridge properties during agonist-stimulated contractions, steady-state force and relative steady-state stiffness were recorded at rest (pCa 9) and during both full (pCa 4) and partial (pCa 7) Ca2+ activations of isolated single alpha-toxin permeabilized vascular smooth muscle cells. For pCa 4 and pCa 7, agonist (1 microM histamine) activation resulted in significant (P < 0.05) increases in both force and stiffness. The agonist-induced increase of steady-state force was significantly (P < 0.05) greater than that of stiffness; at pCa 4, there was a 48% increase for force vs. 17% for stiffness, and, at pCa 7, there was a 160% increase for force vs. 57% for stiffness. The increase in force and stiffness after agonist prestimulation implies that the number of attached cross bridges has increased. However, after agonist prestimulation, we found that the increase of force was greater (P < 0.05) than that of stiffness, resulting in a greater force at any given level of stiffness. Thus these data indicate that agonist activation, presumably via activation of a G protein, increases the relative force per attached cross bridge, possibly by modulating the kinetics of the actomyosin adenosinetriphosphatase to increase in the relative population of cross bridges in force-producing states [actinomyosin (AM) or AM.ADP].

Myosin phosphorylation and regulation of cross-bridge cycle in tracheal smooth muscle

1983 ◽  
Vol 244 (3) ◽  
pp. C182-C187 ◽  
Author(s):  
W. T. Gerthoffer ◽  
R. A. Murphy
Keyword(s):  
Smooth Muscle ◽  
Tracheal Smooth Muscle ◽  
Shortening Velocity ◽  
Active Stress ◽  
Myosin Phosphorylation ◽  
Cross Bridge ◽  
Rapid Changes ◽  
Cross Bridges ◽  
Resting Muscle ◽  
Isotonic Shortening

We have tested the hypothesis that phosphorylation of the 20,000-dalton myosin light chains (LC 20) in rabbit tracheal smooth muscle modulates cross-bridge kinetics and isotonic shortening velocity. The thin muscle [190 +/- 10 (SE) microns] allowed detection of rapid changes in carbachol-induced active stress development, LC 20 phosphorylation, and isotonic shortening velocities. Phosphorylation of the LC 20 in resting muscle was 0.12 +/- 0.04 mol Pi/mol LC 20. Carbachol (10(-5) M) increased the level of phosphorylation to 0.46 +/- 0.03 mol Pi/mol LC 20 within 30 s. Phosphorylation then declined significantly as steady-state active stress was reached. A positive correlation was always found between LC 20 phosphorylation and shortening velocity. This result supports the hypothesis that the level of myosin phosphorylation was related to the mean cross-bridge cycling rate rather than the number of cross bridges contributing to the developed stress. Dephosphorylation of LC 20 occurred at about the same rate as the decline in shortening velocity and stress upon stimulus washout.

An expanded latch-bridge model of protein kinase C-mediated smooth muscle contraction

2005 ◽  
Vol 98 (4) ◽  
pp. 1356-1365 ◽  
Author(s):  
Chi-Ming Hai ◽  
Hak Rim Kim
Keyword(s):  
Smooth Muscle ◽  
Atpase Activity ◽  
Thin Filament ◽  
Muscle Mechanics ◽  
Smooth Muscle Contraction ◽  
Myosin Phosphorylation ◽  
Cross Bridge ◽  
Bridge Model ◽  
Cross Bridges ◽  
Smooth Muscle Mechanics

A thin-filament-regulated latch-bridge model of smooth muscle contraction is proposed to integrate thin-filament-based inhibition of actomyosin ATPase activity with myosin phosphorylation in the regulation of smooth muscle mechanics. The model included two latch-bridge cycles, one of which was identical to the four-state model as proposed by Hai and Murphy ( Am J Physiol Cell Physiol 255: C86–C94, 1988), whereas the ultraslow cross-bridge cycle has lower cross-bridge cycling rates. The model-fitted phorbol ester induced slow contractions at constant myosin phosphorylation and predicted steeper dependence of force on myosin phosphorylation in phorbol ester-stimulated smooth muscle. By shifting cross bridges between the two latch-bridge cycles, the model predicts that a smooth muscle cell can either maintain force at extremely low-energy cost or change its contractile state rapidly, if necessary. Depending on the fraction of cross bridges engaged in the ultraslow latch-bridge cycle, the model predicted biphasic kinetics of smooth muscle mechanics and variable steady-state dependencies of force and shortening velocity on myosin phosphorylation. These results suggest that thin-filament-based regulatory proteins may function as tuners of actomyosin ATPase activity, thus allowing a smooth muscle cell to have two discrete cross-bridge cycles with different cross-bridge cycling rates.

Time dependence of shortening velocity in tracheal smooth muscle

1986 ◽  
Vol 251 (3) ◽  
pp. C435-C442 ◽  
Author(s):  
N. L. Stephens ◽  
M. L. Kagan ◽  
C. S. Packer
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Contraction ◽  
Tracheal Smooth Muscle ◽  
Shortening Velocity ◽  
Time Intervals ◽  
Activity Maximum ◽  
Cross Bridge ◽  
Cross Bridges ◽  
Latch State ◽  
Stimulation Of

It seems fairly well established that in the early phase of smooth muscle contraction cross bridges cycle at a relatively rapid rate. Later on these are replaced by very slowly cycling cross bridges or "latch bridges," operating with high economy. We describe a method to identify the time at which the transition occurs. By abruptly applying a light afterload at varying time intervals after stimulation of a canine tracheal smooth muscle, a point in time could be identified when cross-bridge cycling slowed. This was called the transition time. Because this transition was load dependent, the study was repeated with the preload abruptly reduced to zero. This permitted analysis of data in terms of cross-bridge activity. Maximum zero load velocity (Vo) of the contractile machinery was plotted against time and yielded a biphasic curve. The descending limb of the curve was fitted by a curve of the form Vo(t) = alpha e-K1t + beta e-K2t; K1 was almost three times greater than K2. We speculate that the faster rate constant represented activity of the early rapidly cycling cross bridges, and the slower constant reflected cycling rates in the latch state. These results are consistent with the latch bridge hypothesis put forward by Dillon et al. and enable us to provide a first approximation of the relative velocities of the two types of cross bridges.

Cross-bridge kinetics and shortening in smooth muscle

1994 ◽  
Vol 72 (11) ◽  
pp. 1334-1337 ◽  
Author(s):  
Per Hellstrand
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Isometric Force ◽  
Muscle Mechanics ◽  
Cross Bridge ◽  
Intracellular Ca ◽  
Cross Bridges ◽  
Latch State ◽  
Smooth Muscle Mechanics ◽  
Low Rate

Stiffness measurements were performed on smooth muscle preparations from guinea-pig taenia coli to obtain information on the number of attached cross bridges under varying contractile conditions. The normalized stiffness of the cross-bridge system in smooth muscle may be of a magnitude similar to that assumed in skeletal muscle. Transition from isometric contraction to unloaded shortening was associated with a decrease in stiffness to 50% or less of the isometric value, slightly higher than that found in skeletal muscle fibers. Comparison of phasic (5 s) and tonic (5 min) contractions showed lower Vmax, intracellular [Ca2+], and myosin 20 kDa light chain phosphorylation at 5 min, indicating development of a latch state. Isometric force and stiffness were identical in the two types of contraction. However, stiffness during unloaded shortening was greater in the latch state, which may be the result of the presence of a population of cross bridges with a low rate constant for detachment.Key words: smooth muscle mechanics, cross bridges, latch, myosin phosphorylation.

Sr2+ activates cross-bridge phosphorylation and latch state in smooth muscle

1988 ◽  
Vol 255 (3) ◽  
pp. C401-C407 ◽  
Author(s):  
C. M. Hai ◽  
R. A. Murphy
Keyword(s):  
Steady State ◽  
Induced Responses ◽  
Shortening Velocity ◽  
Myosin Phosphorylation ◽  
Stress Development ◽  
Cross Bridge ◽  
Induced Stress ◽  
Latch State ◽  
State Stress ◽  
Isotonic Shortening

Sr2+ induced myosin phosphorylation and stress development in both skinned and K+-depolarized, Ca2+-depleted, intact swine carotid media. Although higher concentrations of Sr2+ than Ca2+ were required for phosphorylation and stress development, the dependence of stress on phosphorylation was the same in intact tissues. K+ depolarization in the presence of 5 mM Sr2+ produced a transient in phosphorylation (53.2 +/- 5.1% at 1 min, falling to a steady-state value of 21.7 +/- 2.0%) in Ca2+-depleted tissues in which intracellular stores were refilled with Sr2+. Stress developed without a transient (T1/2 = 0.70 min) to a steady state of 89.7 +/- 2.0% of the stress induced by K+ depolarization in the presence of 1.6 mM Ca2+ (K-PSS). Cross-bridge cycling rate as measured by isotonic shortening velocity was proportional to myosin phosphorylation throughout the contraction. When intracellular stores were not refilled with Sr2+, phosphorylation rose to a sustained value of 28.8 +/- 2.7% and stress developed slowly (T1/2 = 2.9 min) to a steady state of 95.9 +/- 1.5% K-PSS-induced stress. Therefore, an initial phosphorylation transient induced by intracellular Sr2+ release only accelerated stress development without significant effects on steady-state stress or phosphorylation (as was true for Ca2+- induced responses). We concluded that Sr2+ substitutes for Ca2+ in phosphorylation and regulation of the latch state in the swine carotid media.

Myosin cross-bridge kinetics in airway smooth muscle: a comparative study of humans, rats, and rabbits

2002 ◽  
Vol 282 (1) ◽  
pp. L83-L90 ◽  
Author(s):  
Y. Lecarpentier ◽  
F.-X. Blanc ◽  
S. Salmeron ◽  
J.-C. Pourny ◽  
D. Chemla ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Comparative Study ◽  
Mechanical Behavior ◽  
Airway Smooth Muscle ◽  
Rate Constants ◽  
Species Differences ◽  
Isometric Tension ◽  
New Formalism ◽  
Cross Bridge ◽  
Cross Bridges

To analyze the kinetics and unitary force of cross bridges (CBs) in airway smooth muscle (ASM), we proposed a new formalism of Huxley's equations adapted to nonsarcomeric muscles (Huxley AF. Prog Biophys Biophys Chem7: 255–318, 1957). These equations were applied to ASM from rabbits, rats, and humans ( n = 12/group). We tested the hypothesis that species differences in whole ASM mechanics were related to differences in CB mechanics. We calculated the total CB number per square millimeter at peak isometric tension (Ψ ×109), CB unitary force (Π), and the rate constants for CB attachment ( f 1) and detachment ( g 1 and g 2). Total tension, Ψ, and Π were significantly higher in rabbits than in humans and rats. Values of Π were 8.6 ± 0.1 pN in rabbits, 7.6 ± 0.3 pN in humans, and 7.7 ± 0.2 pN in rats. Values of Ψ were 4.0 ± 0.5 in rabbits, 1.2 ± 0.1 in humans, and 1.9 ± 0.2 in rats; f 1 was lower in humans than in rabbits and rats; g 2 was higher in rabbits than in rats and in rats than in humans. In conclusion, ASM mechanical behavior of different species was characterized by specific CB kinetics and CB unitary force.

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