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MS 0813 Received 1 March 2000; accepted after revision 10 May 2000.
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ABSTRACT |
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1·7) and recovered to 0·86 ± 0·05 during the 250 ms period of after-stretch potentiation following the rapid decay of force at the end of lengthening (T 1·3); under the same conditions stiffness increased to 1·25 ± 0·02 and to 1·12 ± 0·03, respectively.
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INTRODUCTION |
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Steady lengthening imposed on a contracting muscle elicits an enhancement in force up to a steady value which, at high lengthening speed, is about twice the isometric tetanic force (T0) (Katz, 1939; Lombardi & Piazzesi, 1990). Some of the work done on the muscle during the stretch is mechanically evident after the end of lengthening as a residual potentiation of force (Edman et al. 1978) or shortening capability (Cavagna & Citterio, 1974), which follows the rapid force decay (rate constant, r = 50-250 s-1) to 
1·4T0 and vanishes at a much slower rate (r < 3 s-1) (Edman et al. 1978; Cavagna et al. 1986; Colomo et al. 1989).
Under conditions that preserve sarcomere homogeneity (Lombardi & Piazzesi, 1990), force enhancement during stretch is attributed to both increased strain and, to a lesser extent, increased number of cross-bridges (the myosin heads attached to actin). In terms of the mechanical-kinetic model of Huxley & Simmons (1971), the increased strain in cross-bridges promotes a redistribution towards an early state in the force-generating actin-myosin interaction (Piazzesi et al. 1992). Under these conditions the completion of the normal cross-bridge cycle is prevented, but, beyond a critical strain (3 nm), cross-bridges detach and rapidly reattach without commitment to ATP hydrolysis (Lombardi & Piazzesi 1990; Huxley, 1998). In view of this the rapid force decay to 1·4T0, which occurs when the lengthening is stopped, is related to both the substitution of isometric cross-bridges for cross-bridges which were strained before the stretch ended and a net decrease in number of attachments (Colomo et al. 1989). An alternative view (Cavagna et al. 1994) implies that cross-bridges attached during lengthening undergo a much larger strain: mechanical energy could be stored in the elastic element of the cross-bridges over 50-80 nm filament sliding and transferred, at the end of lengthening, to the damped elements of the cross-bridges without a change in number of attachments. According to this idea the rapid decay of force at the end of lengthening is due to the same process as occurs during the quick recovery following a step stretch of a few nanometres. This process consists, according to the model of Huxley & Simmons (1971), in the synchronised execution of the reversal of the working stroke elicited by the increased mechanical energy in the attached heads.
In this study the nature of force enhancement during and after stretch is reinvestigated in combined mechanical and X-ray diffraction experiments at the ELETTRA Synchrotron (Trieste, Italy) by measuring the stiffness of the half-sarcomere and the changes in the intensity of the bright third order myosin-based meridional X-ray reflection (M3), which arises from the 14·5 nm axial repeat of the myosin heads in the thick filament. The mechanical signal is an indicator of the fraction of attached myosin heads (Linari et al. 1998); the structural signal is sensitive to conformational changes of the heads which change the axial spread in mass density projection (Irving et al. 1992; Irving & Piazzesi, 1997) but is also influenced by changes in the number or axial dispersion of the heads. During both the elastic change of force in response to a step perturbation in length and the subsequent quick phase of force recovery, when no substantial detachment/attachment of cross-bridges has time to occur (Ford et al. 1977; Lombardi et al. 1992; Piazzesi et al. 1997), the changes in intensity of M3 reflection must reflect structural changes in the heads attached before the step. In fact it has been shown that following a step stretch the intensity of M3 decreases both during the elastic response and during the subsequent early phase of force recovery associated with the reversal of the working stroke (Lombardi et al. 1995). In terms of the tilting head model (Dobbie et al. 1998) the spread of mass density projection of the myosin heads onto the filament axis increases as the heads tilt away from the perpendicular to the filament axis and the actin binding domains of the heads move towards the barbed end of the actin filament (Fig. 2 in Lombardi et al. 1995). In a similar way the intensity of M3 reflection is expected to decrease during steady lengthening and, if the rapid force decay following the end of lengthening is driven by the reversal of the working stroke, as suggested by Cavagna et al. (1994), to further decrease when lengthening stops.
Stiffness and X-ray data from the experiments reported here give evidence that enhancement of force during steady lengthening is due to a large increase in the number of cross-bridges with a moderate degree of strain and that the rapid force decay at the end of lengthening is related to both rapid substitution of isometric cross-bridges for strained cross-bridges and partial reduction of the fraction of attached heads, so that the subsequent after-stretch potentiation period is related to a residual increased number of attached heads.
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METHODS |
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The experiments were done at the Small Angle X-ray Scattering (SAXS) beamline at ELETTRA (Trieste, Italy), providing a flux at the sample of up to 5 × 1012 photons s-1 at 0·154 nm wavelength.
Frogs (Rana temporaria), anaesthetised by cold (0°C), were killed by decapitation and pithing, following the official regulations of the European Community Council (Directive 86/609/EEC) assimilated into Italian Government Legislation. Single fibres, dissected from the lateral head of the tibialis anterior muscle, were mounted between the levers of a capacitance gauge force transducer (resonant frequency 40-50 kHz) and a loudspeaker motor, as has already been described elsewhere (Lombardi & Piazzesi, 1990). Two mica windows were moved close to the fibre to reduce the X-ray path in Ringer solution; stimulating electrodes were stuck on the top and bottom edges of the opposing windows. The system, with fibres oriented horizontally, was set on a plate mounted on the movable stage of an ACM Zeiss microscope for measurements of the fibre dimensions and resting sarcomere length and to monitor sarcomere length with a striation follower (Huxley et al. 1981) during stiffness experiments. For the X-ray experiments, the trough was sealed with a Perspex cover and the plate was mounted vertically on the beamline (transducer at the top) to maximize the spatial resolution along the meridional axis.
Mechanical parameters (force, stimulus frequency, fibre length change, sarcomere length change) were recorded on a PC via an A/D card (Computerscope, R.C. Electronics) and analysed using dedicated software. The same card was used to record the X-ray framing. Force and fibre length change were also continuously monitored on a chart recorder. Diffraction patterns were collected on a linear gas-filled detector (dimensions 100 mm × 8 mm) with a spatial resolution of 100 µm. A fast shutter (switching time < 0·25 ms) ensured that the fibre was exposed to X-rays only during the data collection period, to minimise radiation damage. To collect the axial distribution of the meridional reflections, the detector was mounted vertical, parallel to the fibre axis and centred on the beam axis. Alternatively, to record the radial distribution of M3 reflection, the detector was rotated by 90 deg and moved to the level of the third order myosin layer line. This is essential to correct for the changes in the radial width of the reflection related to changes in the three-dimensional lattice sampled by the X-ray beam (Huxley et al. 1982).
Experimental protocol
Tetanic contractions of 1·3 s duration were induced every 4 min at 4°C and 2·11-2·15 µm sarcomere length by electrical stimulation at a frequency of 15-25 Hz. X-ray and stiffness data were collected from a total of 22 fibres. Steady lengthening of 60 nm half-sarcomere-1 was imposed on the fibre by means of the loudspeaker motor at the isometric tetanus, 500 ms after the start of stimulation. Velocity of lengthening was limited to 160 nm s-1 half-sarcomere-1 to minimise the development of inhomogeneous distribution of lengthening (Lombardi & Piazzesi, 1990). At this velocity the transition between the initial rapid rise of force and the quasi-steady force value occurred 60 ms after the start of lengthening, and the steady force value, estimated after 185 ms of lengthening (midway through the quasi-steady tension response), was 1·7T0.
In X-ray experiments the intensity of the M3 reflection during each contraction cycle was recorded in 4 frames of 250 ms duration (Fig. 1): starting 250 ms before the start of stimulation; at the isometric tetanus plateau, 250 ms before the imposed lengthening; during the steady force response to lengthening, 60 ms after the start of lengthening; during the after-stretch potentiation, 50 ms after the end of lengthening, when the rapid phase of tension decay is complete.
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From top to bottom the traces represent: the change in length imposed at the end of the fibre connected to the motor, the change in the sarcomere length recorded by the striation follower (hs, half-sarcomere), force, the four periods when X-ray data were acquired, stimulus. The vertical bars below the force trace mark the times when, in stiffness experiments, length steps were imposed on the fibre. The times of the first (isometric plateau), second (during stretch) and fourth (after stretch) bar occur midway through the corresponding X-ray frames. | ||
X-ray experiments were terminated typically after 20 tetani (20 s total exposure), before development of radiation damage. Fibres with large cross-sectional area (CSA) were selected for X-ray experiments. This explains the difference in CSA between fibres used for X-ray and stiffness experiments (Tables 1 and 2).
Sarcomere stiffness. In stiffness experiments, step length changes of different size (completed within 120 µs) were imposed on the active fibre by means of the loudspeaker motor at various times corresponding to the mid-times of the X-ray frames: 125 ms before and 185 ms after the start of lengthening, for the T0 frame and the stretch frame, respectively, and 175 ms after the end of lengthening for the after-stretch frame (Fig. 1, bars). Steps were also imposed at 50 and 300 ms after the end of lengthening, at the beginning and the end of the after-stretch frame. The actual change in sarcomere length during the step was recorded from a selected segment of the fibre (1-2 mm long) by means of a striation follower (Huxley et al. 1981) as already described (Cecchi et al. 1987). The stiffness was measured as the slope of the relation between the force attained at the end of the step and the corresponding change in average sarcomere length in the selected segment (T1 curve; Huxley & Simmons, 1971).
Data analysis
X-ray diffraction data were analysed with Sigmaplot and Peakfit (Jandel Scientific) software. X-ray patterns were corrected for the non-uniform sensitivity of the detector. The camera background, recorded with the fibre (but not the chamber and solution) moved out of the X-ray beam, was subtracted. The remaining background under each reflection was fitted by a straight line and subtracted. The M3 reflection spacing was estimated from the axial separation of the centres of the Gaussians fitted to the two M3 reflections, calibrated by assuming a spacing of 14·34 nm at rest (Haselgrove, 1975). The M3 intensity was measured as the area of the Gaussians. The M3 radial width was estimated as the width, 
, of the Gaussian fitted to the reflection recorded with the detector horizontal. The contribution of the beam width to the radial spread of the reflection was subtracted after estimating its FWHM (full width at half maximum, 1 mm) by fitting the beam profile at the detector with a Gaussian.
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RESULTS |
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X-ray changes
The axial distribution of the intensity of M3 reflection in the four conditions is shown in Fig. 2A. As already reported (Huxley & Brown, 1967; Bordas et al. 1993; Piazzesi et al. 1999), the spacing of M3 reflection, SM3, increased by 1·5 % from 14·340 nm at rest to 14·554 nm at the isometric tetanus plateau. During the force response to steady lengthening there was a further increase in SM3 to 14·575 nm (0·14 %), which was not reversed during force enhancement after stretch (Table 1).
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A, superimposed intensity distributions along the meridian in the region of the M3 reflection: | ||
Table 1. Mean values (± S.E.M.) for the relevant parameters of X-ray experiments
| Rest | Plateau | Stretch | After stretch 175 ms |
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| T/T0 | 0 | 1 | 1·65 ± 0·05 | 1·33 ± 0·02 |
| SM3 (nm) | 14·340 ± 0·003 | 14·554 ± 0·004 | 14·575 ± 0·006 | 14·572 ± 0·003 |
| IM3/IM3,0 | 1·19 ± 0·03 | 1 | 0·48 ± 0·02 | 0·65 ± 0·03 |
| W/Wr | 1 | 2·05 ± 0·06 | 2·85 ± 0·14 | 2·72 ± 0·11 |
| Corrected IM3/IM3,0 | 0·58 ± 0·02 | 1 | 0·67 ± 0·04 | 0·86 ± 0·05 |
of the Gaussian fitted to the intensity distribution relative to the isometric plateau value. Data from 18 fibres, of which 10 were used for measurement of W/Wr. Sarcomere length, 2·15 ± 0·01 µm; CSA, 23450 ± 1130 µm2; T0, 317 ± 13 kN m-2; lengthening velocity, 161 ± 3 nm s-1 half-sarcomere-1.
As reported for whole muscle (Matsubara & Yagi, 1985), the integrated intensity of the M3 reflection, IM3, decreased to less than 50 % of the isometric plateau value (IM3,0) during steady lengthening. IM3/IM3,0 recovered to 65 % during the after-stretch potentiation following rapid decay of force (Table 1).
The radial distribution of the intensity of the M3 reflection is shown in Fig. 2B. The width of the reflection increased during activation, further increased during steady lengthening and remained large also during the recovery phase. The width W, defined as the of the Gaussian fitted to the intensity distribution corrected for the beam dimension, doubles during the isometric contraction and increases to 2·8 times during both the steady lengthening and the subsequent after-stretch potentiation period (Table 1). This width increase indicates a reduction in the three-dimensional lattice sampled by the X-ray beam due to loss of alignment of thick filaments (Huxley et al. 1982; Piazzesi et al. 1999). Here we show that the inter-filamentary coherence further reduces during lengthening and fails to recover during after-stretch potentiation. IM3 can be corrected for this effect by multiplying by W (Huxley et al. 1982). The corrected IM3/IM3,0 is 0·67 during steady lengthening and increases up to 0·86 during force enhancement after stretch (Table 1).
Stiffness changes
In agreement with previous experiments on Rana esculenta (Colomo et al. 1988, 1989), during steady lengthening at moderate velocity the stiffness of the half-sarcomere rises to a steady value and following the rapid force decay at the end of lengthening it recovers half-way towards the original isometric value. Force responses and T1 relations obtained with length steps of different size and direction (in the range ±2 nm half-sarcomere-1) imposed at the isometric tetanus plateau, during steady force response to lengthening and 175 ms after the end of lengthening are shown in Fig. 3A and B. The stiffness (e) measured as the slope of the first order regression lines fitted to the T1 relations was determined at various times. In four fibres (Fig. 4 and Table 2), stiffness relative to the isometric value (e/e0) was 1·25 during the steady force response to lengthening and dropped to 1·16 at 50 ms after the end of lengthening, when the phase of rapid force decay was complete. The corresponding drop in relative force (T/T0) was from 1·68 to 1·42.
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A, force response to either step stretch (upper row) or release (lower row) of ~1·3 nm half-sarcomere-1 imposed at the isometric tetanus plateau (left column), during steady lengthening (central column) and 175 ms after the end of lengthening (right column). Fibre length, 6·36 mm; segment length, 1·30 mm; sarcomere length at rest, 2·07 µm; CSA, 13700 µm2; T0, 250 kN m-2. B, T1 relations from another fibre in the three conditions in A. | ||
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Mean changes (4 fibres, the vertical bars showing ±S.E.M. are within the size of the symbols) of relative force (T/T0, | ||
Table 2. Mean values (± S.E.M.) for the relevant parameters of stiffness experiments
| Plateau | Stretch | After stretch | |||
| 50 ms | 175 ms | 300 ms | |||
| T/T0 | 1 | 1·684 ± 0·003 | 1·418 ± 0·006 | 1·315 ± 0·004 | 1·234 ± 0·004 |
| e (T0 nm-1) | 0·197 ± 0·003 | 0·246 ± 0·002 | 0·228 ± 0·003 | 0·220 ± 0·004 | 0·210 ± 0·003 |
| e/e0 | 1 | 1·25 ± 0·02 | 1·16 ± 0·02 | 1·12 ± 0·03 | 1·07 ± 0·02 |
1/  (nm T0-1) |
2·32 ± 0·09 | 1·31 ± 0·05 | 1·63 ± 0·07 | 1·79 ± 0·09 | 2·01 ± 0·08 |
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0·43 | 0·76 ± 0·03 | 0·61 ± 0·03 | 0·56 ± 0·03 | 0·50 ± 0·02 |
, cross-bridge equivalent compliance; , fraction of attached cross-bridges. Data from 4 fibres. Sarcomere length, 2·10 ± 0·03 µm; CSA, 17000 ± 3200 µm2; T0, 253 ± 34 kN m-2; lengthening velocity, 170 ± 10 nm s-1 half-sarcomere-1.
Force and stiffness continued to decrease in parallel with a time course which was much slower than that of the initial drop (Colomo et al. 1989). Both for force and stiffness the three after-stretch points can be interpolated with an exponential equation (dashed lines in Fig. 4) using as the fixed parameter the rate constant (2·5 s-1) estimated for the slow process in Colomo et al. (1989). The force and stiffness values extrapolated for zero time were respectively 1·47T0 and 1·19e0.
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DISCUSSION |
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Mechanical evidence for a large recruitment of cross-bridges during lengthening
During steady lengthening the half-sarcomere stiffness becomes 1·25 times the isometric value, similar to that found in Rana esculenta (Lombardi & Piazzesi, 1990; Piazzesi et al. 1992). The stiffness drops during the rapid force decay at the end of lengthening, but remains on average 12 % higher during the after-stretch potentiation period considered (Table 2).
From changes in the half-sarcomere stiffness it is possible to calculate changes in the number of attached myosin heads by using the procedure in Appendix A of Ford et al. (1981) (see also Linari et al. 1998), which allows isolation of the contribution of the mechanically relevant components of the half-sarcomere (myofilaments and cross-bridges) to the elasticity of the half-sarcomere. A prerequisite for the extension of the procedure of Ford et al. (1981) to the steady lengthening condition is that cross-bridge elasticity is linear at forces > T0. This assumption is supported by the finding that both at the tetanus plateau and during lengthening the slopes of instantaneous tension-extension relations are almost linear across the ordinate (Figs 8 and 17 in Piazzesi et al. 1992). At full overlap and assuming the compliance of Z-line to be zero, eqn A3 of Linari et al. (1998) can be reduced to:
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where all terms have dimensions in nm T0-1, Chs is the half-sarcomere compliance, Cf is the myofilament equivalent compliance (given by the sum of myosin filament and actin filament equivalent compliances) and 1/ is the cross-bridge equivalent compliance, with the fraction of attached myosin heads and the stiffness of cross-bridges when all are attached.
The myofilament compliance Cf is estimated from X-ray diffraction measurements (Dobbie et al. 1998; but see also Huxley et al. 1994 and Wakabayashi et al. 1994). At the isometric tetanus plateau, Cf is 0·543Chs. With Chs = (1/e0 =) 5·08 nm T0-1 (Table 2), Cf = (0·543 × 5·08 nm T0-1 =) 2·76 nm T0-1, thus 1/ = 2·32 nm T0-1. During the steady force response to lengthening (e/e0 = 1·25), Chs becomes (5·08 nm T0-1/1·25 =) 4·06 nm T0-1 and 1/ becomes 1·31 nm T0-1, that is cross-bridge compliance is almost halved with respect to the value at T0. Since the elasticity per cross-bridge is assumed to be linear, is a constant and the whole change depends on . If at T0 is 0·43 (Linari et al. 1998), 1/, the cross-bridge compliance with all myosin heads attached, is (2·32 nm T0-1 × 0·43 =) 1·00 nm T0-1. During lengthening 1/ is 1·31 nm T0-1 and becomes (1·00 nm T0-1/1·31 nm T0-1 =) 0·76, 77 % larger than the value at T0. Thus steady lengthening induces the recruitment of a new fraction of heads which represents a large proportion of those already attached in the isometric contraction. This conclusion implies that the high force developed during lengthening is redistributed over a proportionally large number of attachments, so that the strain on cross-bridges is similar to that generated by the working stroke in isometric conditions (1·31 nm T0-1 × 1·68T0 = 2·20 nm during lengthening versus 2·32 nm at T0). Considering that an increase in lengthening velocity produces only a moderate further increase in cross-bridge strain (Lombardi & Piazzesi, 1990), the process by which stretched cross-bridges detach during lengthening must become fast beyond 2-3 nm of cross-bridge strain.
The same analysis applied to stiffness data after stretch (Table 2) shows that, at 50 ms after the end of lengthening, when the rapid force decay is complete, is 0·61 (which is still 42 % larger than the original isometric value); at later times, force slowly recovers towards the original isometric value in parallel with the slow recovery of . Within the experimental error, also follows the exponential decay (rate constant 2·5 s-1) described in the results for force and stiffness. extrapolated to zero time is 0·65 ± 0·01 (-15 % with respect to the lengthening value). This drop is not significantly different from the corresponding drop in force (13 %). In fact, since passively stretched cross-bridges and actively force-generating cross-bridges have about the same average strain, the force per head is the same and it is not possible to discriminate a change in the relative proportion between strained and actively force-generating heads during the fast decay following the end of lengthening.
Structural evidence for an increase in axial dispersion of cross-bridges during lengthening
Despite the increased number of cross-bridges during lengthening, IM3/IM3,0 decreases to 0·67 (Table 1). This discrepancy cannot be explained by the expected reduction in IM3 caused by the change in conformation of the heads during lengthening. In fact, according to Piazzesi et al. (1997), the reversal of the working stroke induced by stretch is at most 2·3 nm (the same extent as the working stroke responsible for isometric force generation). In terms of the structural model in Dobbie et al. (1998) this corresponds to a 16 deg tilt of the light chain region of the head (the lever arm, 9·5 nm long) further away from the perpendicular to the filament axis with respect to the isometric orientation. Taking into account the evidence from stiffness measurements that the elastic strain in the cross-bridges is similar to that in isometric conditions, it can be calculated, with the same model simulation as in Dobbie et al. (1998), that, depending solely on the form factor, IM3 reduces by 31 % going from the isometric configuration (Fig. 5A) to the steady stretch configuration (Fig. 5B). On the other hand, the recruitment of myosin heads, which add at random to the diffracting units and increase the attached fraction to 1·77, is expected to increase IM3 by a factor of (1·772 =) 3·13, so that IM3/IM3,0 becomes (3·13 × 0·69 =) 2·16 times larger. To explain the observed reduction of IM3/IM3,0 to 0·67 an increase in conformational or axial dispersion of the attached heads, which becomes the prominent factor in this case, must be taken into account. Due to the mismatch between actin and myosin periodicity and the dimension of the actin monomer (5·5 nm) the dispersion in isometric conditions should be 2·75 nm at most. Assuming at T0 a Gaussian dispersion with = 2 nm, the actual reduction of IM3/IM3,0 to 0·67 during lengthening can be reproduced with the structural simulation if the dispersion is increased by 1·9 times.
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A, the myosin head (light grey) is in the isometric tetanus plateau conformation: the axial coordinate (z) of the tip of the lever arm (residue 843 in Rayment et al. 1993), marked with a dot, is 7·2 nm away from the rigor value (Dobbie et al. 1998). B, the myosin head (light grey) is in the average conformation assumed during steady lengthening: the lever arm is further tilted by 16 deg away from rigor orientation (z shifted by further 2 nm away from Z-line). C, case with both heads attached during lengthening: it is assumed that the lever arm of the first head (light grey) is back tilted by 32 deg from the isometric configuration and has promoted the attachment of the second head (dark grey, same tilting of the lever arm as isometric) on the actin site farther from the Z-line. | ||
During the after-stretch potentiation period, IM3/IM3,0 recovers to 0·86, while the fraction of attached heads drops to 1·30 times the isometric value. IM3 recovery can be explained if the dispersion has reduced by half with respect to the lengthening value, as is expected if a fraction of heads from previous lengthening, presumably those less strained, are still attached.
The increase in axial dispersion of attached heads induced by lengthening can be attributed to the contribution of two mechanisms: (i) an increase of the range of strains of attached heads in the lengthening direction responsible for the average passive strain of 2 nm (see also Lombardi & Piazzesi, 1990), (ii) attachment of the second head of the same myosin molecule to the next actin monomer favoured by the changed strain or conformation of the first head (Huxley & Tideswell, 1997). Within the limits of the present investigation it is not possible to discriminate between these two hypotheses. The second hypothesis, supported by the strict relation between axial dispersion and number of cross-bridges during and after lengthening, is represented in Fig. 5C with the simplifying assumption that in each head couple the average back-tilting is 16 deg (as derived from mechanical data in Piazzesi et al. 1997) and the average strain is 2 nm (this study). It must be noted that in this case also, as in the case of head attachments adding at random, the doubling of mass on a large proportion (0·77) of the 14·5 nm periodicity originally occupied by one head is expected to increase IM3/IM3,0 by a factor of (1·772 =) 3·13 and thus all the considerations in the initial paragraph of this section remain valid. The idea of a two-head attachment during lengthening implies that each head of the same myosin molecule contributes individually to the overall cross-bridge stiffness. This is suggested by X-ray studies showing that the cross-bridge compliance resides in the myosin heads (Dobbie et al. 1998) and has been previously assumed in comparing stiffness in isometric contraction and rigor (Linari et al. 1998).
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REFERENCES |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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| Bordas, J., Diakun, G. P., Diaz, F. G., Harries, J. E., Lewis, R. A., Lowy, J., Mant, G. R., Martin-Fernandez, M. L. & Towns-Andrews, E. (1993). Two-dimensional time-resolved X-ray diffraction studies of live isometrically contracting frog sartorius muscle. Journal of Muscle Research and Cell Motility 14, 311-324 | [Medline] |
| Cavagna, G. A. & Citterio, G. (1974). Effect of stretching on the elastic characteristics and contractile component of frog striated muscle. The Journal of Physiology 239, 1-14 | [Medline] |
| Cavagna, G. A., Heglund, N. C., Harry, J. D. & Mantovani, M. (1994). Storage and release of mechanical energy by contracting frog muscle fibres. The Journal of Physiology 481, 689-708 | [Abstract] |
| Cavagna, G. A., Mazzanti, M., Heglund, N. C. & Citterio, G. (1986). Mechanical transients initiated by ramp stretch and release to P0 in frog muscle fibers. American Journal of Physiology 251, C571-579 | [Medline] |
| Cecchi, G., Colomo, F., Lombardi, V. & Piazzesi, G. (1987). Stiffness of frog muscle fibres during rise of tension and relaxation in fixed-end or length-clamped tetani. Pflügers Archiv 409, 39-46 | [Medline] |
| Colomo, F., Lombardi, V., Menchetti, G. & Piazzesi, G. (1989). The recovery of isometric tension after steady lengthening in tetanized fibres isolated from frog muscle. The Journal of Physiology 415, 130P. | |
| Colomo, F., Lombardi, V. & Piazzesi, G. (1988). The mechanism of force enhancement during constant velocity lengthening in tetanized single fibres of frog muscle. In Molecular Mechanism of Muscle Contraction, ed. Sugi, H. & Pollack, G. H., Advances in Experimental Medicine and Biology, vol. 226, pp. 489-502. Plenum Press, New York. | |
| Dobbie, I., Linari, M., Piazzesi, G., Reconditi, M., Koubassova, N., Ferenczi, M. A., Lombardi, V. & Irving, M. (1998). Elastic bending and active tilting of myosin heads during muscle contraction. Nature 396, 383-387 | [Medline] |
| Edman, K. A. P., Elzinga, G. & Noble, M. I. M. (1978). Enhancement of mechanical performance by stretch during tetanic contractions of vertebrate skeletal muscle fibres. The Journal of Physiology 281, 139-155 | [Abstract] |
| Ford, L. E., Huxley, A. F. & Simmons, R. M. (1977). Tension responses to sudden length change in stimulated frog muscle fibres near slack length. The Journal of Physiology 269, 441-515 | [Medline] |
| Ford, L. E., Huxley, A. F. & Simmons, R. M. (1981). The relation between stiffness and filament overlap in stimulated frog muscle fibres. The Journal of Physiology 311, 219-249 | [Abstract] |
| Haselgrove, J. C. (1975). X-ray evidence for conformational changes in the myosin filaments of vertebrate striated muscle. Journal of Molecular Biology 92, 113-143 | [Medline] |
| Huxley, A. F. (1998). Biological motors: Energy storage in myosin molecules. Current Biology 8, R485-488 | [Medline] |
| Huxley, A. F., Lombardi, V. & Peachey, L. D. (1981). A system for fast recording of longitudinal displacement of a striated muscle fibre. The Journal of Physiology 317, 12-13P. | |
| Huxley, A. F. & Simmons, R. M. (1971). Proposed mechanism of force generation in striated muscle. Nature 233, 533-538 | [Medline] |
| Huxley, A. F. & Tideswell, S. (1997). Rapid regeneration of power stroke in contracting muscle by attachment of second myosin head. Journal of Muscle Research and Cell Motility 18, 111-114 | [Medline] |
| Huxley, H. E. & Brown, W. (1967). The low-angle X-ray diagram of vertebrate striated muscle and its behaviour during contraction and rigor. Journal of Molecular Biology 30, 383-434 | [Medline] |
| Huxley, H. E., Faruqi, A. R., Kress, M., Bordas, J. & Koch, M. H. J. (1982). Time-resolved X-ray diffraction studies of the myosin layer-line reflections during muscle contraction. Journal of Molecular Biology 158, 637-684 | [Medline] |
| Huxley, H. E., Stewart, A., Sosa, H. & Irving, T. (1994). X-ray diffraction measurements of the extensibility of actin and myosin filaments in contracting muscle. Biophysical Journal 67, 2411-2421 | [Abstract] |
| Irving, M., Lombardi, V., Piazzesi, G. & Ferenczi, M. A. (1992). Myosin head movements are synchronous with the elementary force-generating process in muscle. Nature 357, 156-158 | [Medline] |
| Irving, M. & Piazzesi, G. (1997). The motions of myosin heads that drive muscle contraction. News in Physiological Sciences 12, 249-254. | |
| Katz, B. (1939). The relation between force and speed in muscular contraction. The Journal of Physiology 96, 45-64. | |
| Linari, M., Dobbie, I., Reconditi, M., Koubassova, N., Irving, M. Piazzesi, G. & Lombardi, V. (1998). The stiffness of skeletal muscle in isometric contraction and rigor: the fraction of myosin heads bound to actin. Biophysical Journal 74, 2459-2473 | [Abstract/Full Text] |
| Lombardi, V. & Piazzesi, G. (1990). The contractile response during steady lengthening of stimulated frog muscle fibres. The Journal of Physiology 431, 141-171 | [Abstract] |
| Lombardi, V., Piazzesi, G., Ferenczi, M. A., Thirlwell, H., Dobbie, I. & Irving, M. (1995). Elastic distortion of myosin heads and repriming of the working stroke in muscle. Nature 374, 553-555 | [Medline] |
| Lombardi, V., Piazzesi, G. & Linari, M. (1992). Rapid regeneration of the actin-myosin power stroke in contracting muscle. Nature 355, 638-641 | [Medline] |
| Matsubara, I. & Yagi, N. (1985). Movements of cross-bridges during and after slow length changes in active frog skeletal muscle. The Journal of Physiology 361, 151-163 | [Abstract] |
| Piazzesi, G., Francini, F., Linari, M. & Lombardi, V. (1992). Tension transients during steady lengthening of tetanized muscle fibres of the frog. The Journal of Physiology 445, 659-711 | [Abstract] |
| Piazzesi, G., Linari, M., Reconditi, M., Vanzi, F. & Lombardi, V. (1997). Cross-bridge detachment and attachment following a step stretch imposed on active single frog muscle fibres. The Journal of Physiology 498, 3-15 | [Abstract] |
| Piazzesi, G., Reconditi, M., Dobbie, I., Linari, M., Boesecke, P., Diat, O., Irving, M. & Lombardi, V. (1999). Changes in conformation of myosin heads during the development of isometric contraction and rapid shortening in single frog muscle fibres. The Journal of Physiology 514, 305-312 | [Abstract/Full Text] |
| Rayment, I., Holden, H. M., Whittaker, M., Yohn, C. B., Lorenz, M., Holmes, K. C. & Milligan, R. A. (1993). Structure of the actin-myosin complex and its implications for muscle contraction. Science 261, 58-65 | [Medline] |
| Wakabayashi, K., Sugimoto, Y., Tanaka, H., Ueno, Y., Takezawa, Y. & Amemiya, Y. (1994). X-ray diffraction evidence for the extensibility of actin and myosin filaments during muscle contraction. Biophysical Journal 67, 2422-2435 | [Abstract] |
This work has been supported by grants from MURST (Cofinanziamento 1998), Telethon (n. 945), INFM and CNR. We thank Professor Malcolm Irving for discussion and suggestions during the preparation of the manuscript, Dr Natalia Koubassova for substantial help with the structural model and Mr Alessandro Aiazzi and Mr Mario Dolfi for mechanical and electronics support.
Corresponding author
V. Lombardi: Dipartimento di Scienze Fisiologiche, Viale G.B. Morgagni 63, I-50134 Firenze, Italy.
Email: vincenzo.lombardi{at}unifi.it
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, rest; 
, isometric tetanus plateau; 
, lengthening; 
, after lengthening. The integrated area is ±1/85 nm-1 across the meridian, according to the width of the linear detector. Two fibres from March 1999 visit. B, intensity distribution of the M3 reflection in the radial direction. The integrated area is from 1/12·5 nm-1 to 1/17·5 nm-1. In these fibres (6 fibres from March 1999 visit) values for W/Wr were 1·96 for the isometric tetanus plateau, 2·54 during stretch and 2·54 after stretch.
) and stiffness (e/e0,