Tonicity Effects on Intact Single Muscle Fibers: Relation Between Force and Cell Volume

Gulati, Jagdish; Babu, A.

doi:10.1126/science.6977845

Cited by 41 publications

(32 citation statements)

References 26 publications

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“…Although we observed a significant (-12%) increase in the diameter of fibers under rigor conditions (-7% in relaxing buffer) when the S-2 epitopes of the thick filament were saturated with anti-S-2 antibody, a similar force response also occurred when (rigor) fibers were treated with anti-S-2 Fab without a detectable change in the fiber diameter. In this connection, Gulati and Babu (17) [see also Edman and Reggiani (18) The simplest interpretation of the present results is that antibody binding to S-2 affects the ability of this segment to undergo a normal cross-bridge cycle. This might occur, for example, by altering the electrostatic charge along S-2 or by physically interfering with the bonding interactions between S-2 and neighboring regions of the thick-filament backbone.…”

Section: Discussionmentioning

confidence: 50%

Suppression of contractile force in muscle fibers by antibody to myosin subfragment 2.

Lovell

Karr

Harrington

1988

Proc. Natl. Acad. Sci. U.S.A.

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Polyclonal antibody directed against the subfragment-2 region of myosin was purified by affinity chromatography. Skinned muscle fibers that had been preincubated with antibody were able to sustain only 7% of the active isometric force generated by control fibers. The effect of antibody on force production could not be accounted for by inhibition of ATP turnover.The classical rotating-head sliding filament model for force generation in activated muscle fibers (1-3) proposes that the force-generating event results from a structural change in the subfragment 1 (S-1) region (the myosin head) while it is attached to actin. The helix-coil model (4, 5) for force generation proposes that melting and shortening in a section [the heavy meromyosin (HMM)/light meromyosin (LMM) hinge domain] of subfragment 2 (S-2) occurs as the actin-attached cross-bridge swings away from the thick filament surface in an active bridge cycle. It has proved difficult to decide conclusively between these two models; indeed, it seems possible that aspects of both processes are fundamental to the tension-generating mechanism in muscle. Evidence has been provided that beads coated with HMM (6) as well as soluble HMM fragments (7) can move along actin filaments energized by ATP at speeds approaching those obtained with muscle fibers under no-load conditions. Fluorescent actin filaments have also been shown to slide along single-headed myosin filaments bound to a glass support (8) and isolated S-1 subunits bound to a nitrocellulose film (9) (11,12). Although other interpretations are possible, this finding suggests that release of S-2 from the thick filament surface in a normal bridge cycle may be an essential element of the force-generating mechanism in muscle.In the work to be described below, we examine the effect of anti-S-2 polyclonal antibodies on the contractile force of activated, skinned psoas fibers. As in the cross-linking studies, our primary goal was to determine if modulation of the interaction between S-2 and the thick-filament backbone could suppress the isometric force in the absence of any direct effect on the ability of the S-1 subunit to cycle through actin. MATERIALS AND METHODSPreparation of Muscle Proteins and Immunogens. Chicken myosin was prepared by extraction of chicken breast muscle with Guba Straub solution (0.3 M KC1/0.09 M KH2PO4/ 0.06 M K2HPO4) followed by repeated washes in low-ionicstrength buffer (13). HMM and LMM were prepared by digestion of chicken myosin (10 mg/ml) with chymotrypsin (0.1 mg/ml) in 20 mM NaP,/3 mM MgC12/0.6 M NaCl, pH 6.9 for 6 hr at 40C. Digestion was quenched by the addition of phenylmethylsulfonyl fluoride to 1 mM. LMM and undigested myosin were precipitated by dialysis of the digest against 20 mM imidazole hydrochloride/i mM CaCl2, pH 6.2. LMM was further purified by ethanol fractionation. S-2 was prepared from HMM by the method of Sutoh et al. (14) and further purified by chromatography on Sepharose 4B in 20 mM imidazole hydrochloride (pH 6.2). Addition of CaCl2 (to 6 mM) to the c...

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Section: Discussionmentioning

confidence: 50%

Suppression of contractile force in muscle fibers by antibody to myosin subfragment 2.

Lovell

Karr

Harrington

1988

Proc. Natl. Acad. Sci. U.S.A.

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“…Decreasing myofilament lattice spacing beyond a certain point could interfere with normal crossbridge interaction and any contribution crossbridges might make to the activation of the thin filaments. This is indicated by the observations that fibre compression, with greater than 10% T-500, decreases maximal force (Gulati & Babu, 1982 and increases resting stiffness in intact and skinned fibres (Berman & Maughan, 1982;Goldman & Simmons, 1986;Umazume et al, 1986). These results, along with the decrease in pK oberved with osmotic compression at 3.4 ~m, suggest that there may be a critical lattice spacing below which myosin interaction with actin is altered, resulting in not only altered force, but a decrease in filament calcium sensitivity.…”

Section: The Effect Of Lattice Spacing Changes On Pkmentioning

confidence: 55%

Length and myofilament spacing-dependent changes in calcium sensitivity of skeletal fibres: effects of pH and ionic strength

Martyn

Gordon

1988

J Muscle Res Cell Motil

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The calcium sensitivity of force was measured in glycerinated rabbit psoas fibres at sarcomere lengths (SL) from 2.3 to 3.4 micron. Increased SL caused calcium sensitivity to increase and the slope of force-calcium relations to decrease. We have hypothesized that length-dependent changes in myofilament lattice spacing and the presence of fixed charge on the myofilaments are important in determining calcium sensitivity. Lattice spacing changes were monitored by measuring fibre diameter (D). D was decreased by increasing SL, decreasing bathing solution pH and by osmotic compression with 3% PVP. 3% PVP caused D to decrease by about 15% at all SLs and pH values tested. Force-calcium relations were measured at different SLs and pH values, with and without 3% PVP in the bathing solutions. At all pH values D at SL 2.3 micron with 3% PVP was comparable to the value at 3.4 micron, without PVP. At pH 7.5 and 7.0 calcium sensitivity was about the same at both SL, although the slope of the force-calcium relation was less at longer SL. The similarity of the calcium sensitivity at the same D, but much different SL, indicates that lattice spacing is important in determining calcium sensitivity, while SL and the degree of myofilament overlap are important in determining the slope of force-calcium relations. In order to test for the role of myofilament charge in determining calcium sensitivity, pH and ionic strength were varied. Decreasing pH caused decreased maximum force and calcium sensitivity. In addition, the influence of SL on calcium sensitivity decreased as pH was lowered, with minimal SL dependence at pH 5.5; even though lattice spacing still decreased with increasing SL. When D was decreased with PVP, calcium sensitivity increased at all SLs in pH 7.5 and 7.0 while the same lattice spacing changes at pH 6.0 and 5.5 resulted in greatly reduced shifts in calcium sensitivity. These results indicate that the effect of lattice spacing on calcium sensitivity depends on myofilament charge. At pH 6.0, even though osmotic compression of the lattice has no effect, increasing SL causes about half the shift in calcium sensitivity seen at pH 7.0. Lowering ionic strength from 200 to 110 mM caused an increase in both the magnitude and length dependence of calcium sensitivity at pH 7.0, while at pH 5.5 both decreased.(ABSTRACT TRUNCATED AT 400 WORDS)

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“…This resultant increase in extracellular osmolarity is large and rapid and is known to result in shrinkage of erythrocytes [2,35] and non-contracting skeletal muscle [3]. It is well known that changes in skeletal muscle cell volume affect contractile function [36][37][38] and cellular metabolism [39,40], suggesting that it may be important for skeletal muscle cells to regulate cell volume.…”

Section: Role Of the Nkcc In Skeletal Muscle Cell Volume Regulationmentioning

confidence: 99%

K<sup>+</sup> Transport and Volume Regulatory Response by NKCC in Resting Rat Hindlimb Skeletal Muscle

Lindinger

Hawke

Lipskie

et al. 2002

Cell Physiol Biochem

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This study tested the hypothesis that the NKCC is involved in volume regulation, specifically regulatory volume increase (RVI), in resting skeletal muscle. Neurally and vascularly isolated rat hindlimbs were perfused with a bovine erythrocyte perfusate containing ⁴²K or ⁸⁶Rb as markers of unidirectional K⁺ flux across the sarcolemma. Compared to controls, perfusion with 120 µM bumetanide (a specific inhibitor of the NKCC) decreased J_inK by 15±2%, indicating the functional presence of the NKCC. Experiments with ouabain (to block active K⁺ transport by the Na,K ATPase) showed that the bumetanide-sensitive component of J_inK comprised 35% of the total ouabain-sensitive J_inK. Inhibition of NKCC resulted in a net loss of water by muscle. When hindlimbs were perfused with hypertonic (380 mOsm/L by addition of sucrose) perfusate for 20 min, after initially blocking K⁺ channels with 1 mM barium, J_inK rapidly (2-3 min) increased 2-fold followed by a rapid decline. This rapid, transient increase in J_inK was abolished with bumetanide, confirming that perfusion with hypertonic perfusate stimulated NKCC activity and RVI. The hypertonic perfusate also resulted in temporally associated decreases in net water uptake by muscle. It is concluded that a functional NKCC is present in mammalian skeletal muscle and that it is involved in cell volume regulation.

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Tonicity Effects on Intact Single Muscle Fibers: Relation Between Force and Cell Volume

Cited by 41 publications

References 26 publications

Suppression of contractile force in muscle fibers by antibody to myosin subfragment 2.

Suppression of contractile force in muscle fibers by antibody to myosin subfragment 2.

Length and myofilament spacing-dependent changes in calcium sensitivity of skeletal fibres: effects of pH and ionic strength

K<sup>+</sup> Transport and Volume Regulatory Response by NKCC in Resting Rat Hindlimb Skeletal Muscle

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