Calcium dependence of myosin phosphorylation and airway smooth muscle contraction and relaxation

1986 ◽  
Vol 250 (4) ◽  
pp. C597-C604 ◽  
Author(s):  
W. T. Gerthoffer
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Dose Response ◽  
Time Course ◽  
Physiological Salt Solution ◽  
Shortening Velocity ◽  
Myosin Phosphorylation ◽  
Calcium Dependence ◽  
Contraction And Relaxation ◽  
Isotonic Shortening

The time course and the steady-state calcium dependence of myosin phosphorylation and isotonic shortening velocity were studied during contraction and relaxation of canine tracheal smooth muscle. Dephosphorylation of myosin coincided with the decay of isotonic shortening velocity during rapid relaxation following agonist washout. However, the decay of shortening velocity preceded dephosphorylation during a slow relaxation induced by Ca2+-free physiological salt solution (PSS). Carbachol dose-response curves for isometric stress development and myosin phosphorylation were superimposable but shifted to the left of the shortening velocity dose-response. The steady-state Ca2+ dependence of myosin phosphorylation was defined using carbachol and K+ as agonists. There was a significant dissociation of dephosphorylation and relaxation following a stepwise reduction of extracellular CaCl2 concentration. This result was related to muscarinic activation because the dissociation of relaxation and dephosphorylation was reduced by atropine in muscles stimulated with K+. Myosin phosphorylation was completely dissociated from contraction when muscles were stimulated with carbachol in Ca2+-free PSS and contracted by readmission of CaCl2. Mechanisms in addition to myosin phosphorylation appear to regulate airway muscle tone and shortening velocity, and two possibilities are discussed.

Myosin phosphorylation and contraction of feline esophageal smooth muscle

1985 ◽  
Vol 249 (1) ◽  
pp. C9-C14 ◽  
Author(s):  
N. W. Weisbrodt ◽  
R. A. Murphy
Keyword(s):  
Smooth Muscle ◽  
Mechanical Activation ◽  
Smooth Muscles ◽  
Circular Muscle ◽  
Electrical Field Stimulation ◽  
Muscle Layer ◽  
Field Stimulation ◽  
Shortening Velocity ◽  
Myosin Phosphorylation ◽  
Isotonic Shortening

We tested the hypothesis that phosphorylation of the 20,000-Da light chain of myosin (LC 20) is related to mechanical activation of esophageal smooth muscle. Circular muscle layer strips of cat esophagus were taken from the lower esophageal sphincter (LES) and the distal esophageal body (EB). The LES strips developed tone spontaneously, and the EB strips were tonically contracted with carbachol. Both tissues relaxed in response to electrical-field stimulation. Phosphorylation of the LC 20 was determined in tissues quick-frozen during relaxation and during stress redevelopment after cessation of field stimulation. Stress and phosphorylation levels were low after 30 s of field stimulation, and a rapid contraction followed field stimulation. Phosphorylation in the LES increased from 0.043 +/- 0.029 to 0.328 +/- 0.043 mol Pi/mol LC 20 within 10 s after stimulation of the inhibitory nerves was terminated, while stress was still rising rapidly. Phosphorylation in the LES then declined to a steady-state value of 0.162 +/- 0.034 mol Pi/mol LC 20 after 10 min. Isotonic shortening velocities at a constant afterload following a quick release showed changes with time that were proportional to the level of phosphorylation. This was also true for values of maximal shortening velocity estimated for zero external load and for the rate of stress redevelopment after a step shortening. Comparable measurements were made in the carbachol-contracted EB. These results indicate that visceral smooth muscles, which normally function tonically (LES) or phasically (EB), exhibit an initial rapid mechanical activation associated with myosin phosphorylation.(ABSTRACT TRUNCATED AT 250 WORDS)

Cross-bridge dephosphorylation and relaxation of vascular smooth muscle

1989 ◽  
Vol 256 (2) ◽  
pp. C282-C287 ◽  
Author(s):  
C. M. Hai ◽  
R. A. Murphy
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Time Course ◽  
Electrical Field Stimulation ◽  
Shortening Velocity ◽  
Electrical Field ◽  
Cross Bridge ◽  
Time Course Data ◽  
Necessary And Sufficient ◽  
Isotonic Shortening

We tested the hypothesis that relaxation in vascular smooth muscle is the result of inactivation of myosin light chain kinase and cross-bridge dephosphorylation. Fast neurally mediated contractions of swine carotid medial strips were induced by electrical field stimulation. Termination of the stimulus resulted in relaxation with a half time of 2 min. Nifedipine (0.1 microM) increased the relaxation rate without significant effects on the contractile response. Cross-bridge dephosphorylation was much faster than stress decay with basal levels reached within 1 min when 73% of the developed stress remained. The time-course data of dephosphorylation and stress were analyzed with a model that predicted the dependences of stress and isotonic shortening velocity on cross-bridge phosphorylation during contraction. Rate constants resolved from contraction data also fitted the relaxation data when the model's prediction was corrected for estimated errors in the phosphorylation measurements. Because Ca2+-dependent cross-bridge phosphorylation was the only postulated regulatory mechanism in the model, these results are consistent with the hypothesis that cross-bridge dephosphorylation is necessary and sufficient to explain relaxation in the swine carotid media.

Myosin light chain phosphorylation and contraction of guinea pig gallbladder smooth muscle

1991 ◽  
Vol 261 (6) ◽  
pp. G952-G957
Author(s):  
R. J. Washabau ◽  
M. B. Wang ◽  
J. P. Ryan
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Light Chain ◽  
Myosin Light Chain ◽  
State Level ◽  
Myosin Light Chain Phosphorylation ◽  
Shortening Velocity ◽  
Cross Bridges ◽  
Contraction And Relaxation ◽  
Isotonic Shortening

These experiments were designed to determine 1) whether acetylcholine (ACh) stimulation is accompanied by changes in myosin light chain phosphorylation in gallbladder smooth muscle and 2) whether dephosphorylated noncycling cross bridges (latch bridges) exist in gallbladder smooth muscle. Isometric stress, isotonic shortening velocity, and myosin light chain phosphorylation were determined under conditions of contraction and relaxation in ACh-stimulated guinea pig gallbladder smooth muscle. Unstimulated muscle contained 6.8 +/- 2.0% phosphorylated myosin light chain. ACh stimulation (5 x 10(-5) or 10(-4) M) was associated with a rapid increase in myosin light chain phosphorylation to a value that was maintained throughout the tonic contraction. In contrast, isotonic shortening velocity was maximal at 30 s of stimulation and then declined over time to a steady-state level that was 25-30% of the peak velocity. Upon agonist washout (relaxation), dephosphorylation of the myosin light chain occurred at about the same rate as the decline in shortening velocity and preceded the decline in isometric stress. These data suggest that ACh stimulation is accompanied by changes in myosin light chain phosphorylation but that dephosphorylation of cross bridges is not necessary for the slowing of cross-bridge cycling rates in gallbladder smooth muscle.

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.

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.

Role of myosin light-chain phosphorylation in guinea pig gallbladder smooth muscle contraction

1994 ◽  
Vol 266 (3) ◽  
pp. G469-G474 ◽  
Author(s):  
R. J. Washabau ◽  
M. B. Wang ◽  
C. Dorst ◽  
J. P. Ryan
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Smooth Muscle Contraction ◽  
Myosin Light Chain Phosphorylation ◽  
Shortening Velocity ◽  
Cross Bridges ◽  
Isotonic Shortening

In acetylcholine (ACh)-stimulated gallbladder smooth muscle, we have previously shown that phosphorylation of the 20,000-Da myosin light chains is necessary for the initiation of contraction, that myosin is stably phosphorylated at steady state, and that dephosphorylation of cross bridges is not necessary for the slowing of cross-bridge cycling rates during the period of steady-state isometric stress. The present studies were undertaken to determine whether 1) K+ (60 or 80 mM) or cholecystokinin (CCK, 10(-8) M) stimulation is accompanied by changes in myosin light-chain phosphorylation in gallbladder smooth muscle and 2) dephosphorylated noncycling cross bridges exist in K(+)- or CCK-stimulated gallbladder smooth muscle. Isometric stress, isotonic shortening velocity, and myosin light-chain phosphorylation were determined during contraction with K+ or CCK. Steady-state isometric stress was reached within 2.5 min of stimulation with K+ or CCK and was maintained for the duration of the stimulation. Stimulation with K+ or CCK was associated with rapid increases in myosin light-chain phosphorylation and maintenance of myosin light-chain phosphorylation during the stimulation. In contrast, isotonic shortening velocity was maximal at 1 min of stimulation with either K+ or CCK and then declined significantly to values that were only 26-32% of the peak velocity. These data, along with data from previous experiments with ACh, suggest that myosin light-chain phosphorylation is essential in the initiation of contraction in gallbladder smooth muscle, regardless of the source of Ca2+ or of the contractile agonist.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of glycerinated smooth muscle contraction and relaxation by myosin phosphorylation

1988 ◽  
Vol 255 (1) ◽  
pp. C34-C42 ◽  
Author(s):  
J. A. Tanner ◽  
J. R. Haeberle ◽  
R. A. Meiss
Keyword(s):  
Smooth Muscle ◽  
Isometric Force ◽  
Smooth Muscle Contraction ◽  
Muscle Stiffness ◽  
Bathing Solution ◽  
Shortening Velocity ◽  
Myosin Phosphorylation ◽  
Uterine Muscle ◽  
Contraction And Relaxation ◽  
Gamma S

Regulation of isometric force maintenance, isotonic shortening velocity, and muscle stiffness by myosin phosphorylation was examined during both contraction and relaxation of chemically permeabilized (glycerinated) rat uterine smooth muscle. Phosphorylation of the 20,000-Da light chain of myosin (LC20) was manipulated by varying the calcium activity of the bathing solution or by thiophosphorylation of LC20 in the presence of ATP gamma S. With saturating calcium and calmodulin, LC20 phosphorylation was 0.43 mol PO4/mol LC20. This increased to 0.92-0.96 mol PO4/mol LC20 on addition of ATP gamma S. Over the entire range of phosphorylation, there was a significant (P less than 0.001) linear correlation between force and phosphorylation. Stiffness increased monotonically with increasing force; however, the relationship was nonlinear, with stiffness increasing faster at lower levels of activation. Force, stiffness, shortening velocity, and LC20 phosphorylation were compared at identical calcium activities during steady-state conditions of partial contraction and partial relaxation. The ratio of the value of each parameter measured during relaxation to that measured during contraction was 1.11 for force, 1.09 for stiffness, 1.01 for shortening velocity, and 0.83 for LC20 phosphorylation. These results support the hypothesis that contraction and relaxation in glycerinated rat uterine muscle are regulated primarily by phosphorylation and dephosphorylation of LC20.

scholarly journals Rat uterine microbiochemistry: metabolic enzyme activities stimulated by 17-beta-estradiol are localized in epithelial cells.

1987 ◽  
Vol 35 (6) ◽  
pp. 657-662 ◽  
Author(s):  
J P Holt ◽  
E Rhe
Keyword(s):  
Enzyme Activity ◽  
Smooth Muscle ◽  
Epithelial Cells ◽  
Dose Response ◽  
Enzyme Activities ◽  
Time Course ◽  
Citrate Synthase ◽  
Ovariectomized Rats ◽  
Similar Response ◽  
Estradiol Treatment

Lactate dehydrogenase (LDH; EC 1.1.1.27), citrate synthase (CS; EC 4.1.3.7), and beta-hydroxyacyl-CoA-dehydrogenase (beta-OH-acyl-CoA-DH; EC 1.1.1.35) activities were determined in each of the three major cell types of rat uterus, i.e., epithelial, stromal, and smooth muscle, using quantitative microanalytical techniques. Adult ovariectomized rats were treated with 17-beta-estradiol to determine the time course and dose response (0.025-50 micrograms/300-g rat) effect of estrogen on enzyme activity of each type of uterine cell. The use of "oil well" and enzyme-cycling microtechniques to determine the time course and the dose responses of enzyme activity changes required microassays involving 1595 microdissected single cell specimens. Estradiol treatment increased epithelial LDH, CS and beta-OH-acyl-CoA-DH activity but had no effect on these enzymes in the stroma or in smooth muscle cells. The estradiol-stimulated peak enzyme activities on Day 4 in the intervention group are compared with those in the ovariectomized rat controls as follows: LDH, 44.5 +/- 3.5 vs 22.3 +/- 3.9; CS, 3.5 +/- 0.2 vs 1.5 +/- 0.6; beta-OH-acyl-CoA-H, 3.5 +/- 0.32 vs 2.2 +/- 0.2 (mean +/- standard deviation; mol/kg/hr). Stromal cell activities (LDH, 7.4 +/- 1.0; CS, 1.2 +/- 0.2; beta-OH-acyl-CoA-DH, 0.9 +/- 0.1) were significantly lower than epithelial cell levels and were similar to smooth muscle levels. Therefore, even in the ovariectomized animal epithelial cells have markedly higher metabolic activity compared with adjacent cells. The enzyme activities are expressed as moles of substrate reacting per kilogram of dry weight per hour. All three enzymes exhibited a 17-beta-estradiol-induced dose response between 0.025-0.15 micrograms/300-g rat. The three enzymes studied all had similar response patterns to estrogen. The effect of estradiol was restricted to epithelial cells, with enzyme activities increasing to maximal levels after approximately 96 hr of hormone treatment. This study therefore not only confirms the specific and differential metabolic responses of uterine cells to estradiol treatment, but clearly demonstrates that marked metabolic differences exist between epithelial cells and stromal or smooth muscle uterine cells.

Mechanical strain memory in airway smooth muscle

2000 ◽  
Vol 278 (5) ◽  
pp. C895-C904 ◽  
Author(s):  
Wah-Lun Chan ◽  
Jeanette Silberstein ◽  
Chi-Ming Hai
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Mechanical Strain ◽  
Control Level ◽  
High Stress ◽  
Peak Stress ◽  
Airway Smooth Muscle Cells ◽  
Initial Length ◽  
Myosin Phosphorylation ◽  
Contraction And Relaxation

We investigated the effect of a single rapid stretch on poststretch force and myosin phosphorylation in bovine tracheal smooth muscle. When unstimulated muscle strips were stretched from suboptimal length to optimal length ( L o), poststretch steady-state force was not significantly different from that of unstretched control at L o. However, when carbachol-activated muscle strips were stretched from suboptimal length to L o, poststretch force and myosin phosphorylation were lower than control and significantly correlated with initial length. When poststretch muscle strips were allowed to relax for 1 h and then activated by K+ depolarization, the developed force remained significantly correlated with initial length. When the same strain was applied in 23 increments to minimize peak stress, poststretch force and myosin phosphorylation increased significantly, approaching the levels expected at L o. Furthermore, poststretch force development increased after each cycle of contraction and relaxation, approaching the control level after four cycles. These results suggest that activated airway smooth muscle cells can retain relatively precise memory of past strain when they are stretched rapidly with high stress.

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