Special Article
Quantal Mechanisms in Cardiac Contraction*
GERALD H.
S
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
ERENDIPITY and technology have combined
forces to reveal a phenomenon of striking proportions. What may prove to be the quantum
element underlying contraction was discovered by
chance, following the development of a high-resolution optical diffractometer designed to follow the dynamics of shortening in isolated cardiac muscle. Records showed a most unusual feature: periods of
sarcomere shortening were punctuated by periods of
pause during which there was little or no length
change, conferring a staircase-like character on the
shortening waveform.1
Could shortening really occur in discrete quanta? To
find precedent for quantal behavior one need not look
far. Nearby, at the myoneural junction, for example, a
detailed understanding of the mechanism of intercellular communication followed the discovery that acetylcholine was discharged in quantal packets.2 These
packets sum to form the end plate potential in much the
same way that the steps apparently sum to give the
shortening waveform. In the latter case, however, an
unfathomable implication has impeded acceptance.
The difficulty is that stepwise shortening implies a
colossal degree of synchrony. Why so? The optical
diffraction pattern reflects the behavior of a sizable
fraction of sarcomeres ranging over the field illuminated by the laser. This ensemble could pause in either of
two ways. In the complicated way, some sarcomeres
would shorten while others lengthened in such a way
that the average velocity remained zero for a period;
this situation would then recur fortuitously time and
again as contraction progressed to give the observed
cascade of pauses and steps. In fact, this possibility is
now ruled out; even when the sampled volume is diminished extraordinarily, the pauses remain in evidence (see below).
The alternative way is simpler. It implies that memFrom the Division of Bioengineering, University of Washington,
Seattle, Washington.
Address for reprints: Dr. Gerald H. Pollack, Division of Bioengineering, WD-12, University of Washington, Seattle, WA 98195.
*A critique of this article was written by Sir Andrew Huxley at
the invitation of the Editor. This critique is published herein as the
next article.
(Circulation Research 1986;59:l-8)
POLLACK
bers of the ensemble pause and shorten synchronously.
To those seriously engaged in thinking about molecular contractile mechanisms, this is disturbing. It implies that the elements underlying shortening can
somehow act in concert to produce the observed quantal behavior. The synchronous domain is vast. In the
experiments cited above, for example, the incident
beam encompassed some 100 million myofilaments,
a region which by molecular standards is essentially
infinite.
The synchrony implied by stepwise shortening,
then, questions the very foundation underlying current
ideas of how muscles shorten. The present theory, for
example, contains no provision to synchronize the action of bridges. 3 ' 4 Needless to say, the specter of Maxwell's demon calling the strokes has not served to
instill confidence in the phenomenon itself.5"7 On the
other hand, if steps do not arise out of some instrument
artifact, their discovery could prove seminal: Synchronized, quantal behavior implies a fundamental rethinking of how muscles shorten.
In the remainder of this article I will present the
evidence that has convinced me that muscle shortening
does, indeed, occur in synchronized steps. I will go on
to discuss some implications of the phenomenon, and
finally, I will briefly mention a possible molecular
mechanism that is, in fact, unrelated to cross-bridge
cycling. To those brought up on a steady diet of sliding filaments, the proposal may prove distinctly unpalatable.
Segment Length Dynamics
Following the pioneering work of Gordon, Huxley,
and Julian,8 a number of investigators have perfected
methods of measuring the dynamics of fiber segments.
Figure 1 shows an example. In this case each segment
was demarcated by a pair of thin hairs from the ear of a
black cat; segment length was tracked optically.9 The
steps apparent in Figure 1 were not a chance observation. Of 21 fibers studied, stepwise shortening was
absent in only 5; in the remaining 16, shortening steps
were punctuated either by brief hesitations or by more
prolonged pauses such as those of Figure 1. The patterns were clearly repeatable from cycle to cycle.
Circulation Research
Vol 59, No 1. July 1986
2%
FIGURE 1. Time course of segment length
change in single fibers of frog skeletal muscle. Segments were demarcated by thin,
dark hairs mounted on the fiber surface at
right angles to the fiber axis. Separation of
markers was tracked by ah optical scanner.
Unstimulated fibers subjected to ramp release and stretch. Initial segment length:
800 ixm, upper; 510 /xm, lower.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
20 msec
The prototype apparatus originated in Prof. K.A.P.
Edman's laboratory in Sweden.10 Black labrador dog
hairs were used. Although the apparatus was designed
for different types of experiment and the published
records are on a compressed time scale, several of the
traces (eg, Edman et al," Figure 8) show distinct
pauses.
Glass microelectrode tips have also been used as
segment markers. Housmans plunged these into the
belly of a papillary muscle and used an optical scanner
to follow the inter-marker spacing.12 Surprisingly
noise-free, the records showed both smooth and stepwise patterns. The stepwise patterns were particularly
apparent when the load was kept modest, and they
were repeatable from contraction to contraction.
Finally, in a more detailed study, Tameyasu et al
used a high-speed video system to follow shortening in
single isolated frog heart cells.13 Segments were demarcated primarily by natural markers such as glycogen granules. Stepwise shortening was found regularly. The number of steps, however, was limited, as the
steps became progressively less distinct with shortening. Nevertheless, the step pattern was repeatable from
contraction to contraction as well as from preparation
to preparation. Catecholamines decreased the pause
duration, accounting for their positive inotropic
action.
The results are consistent: Fiber segments shorten in
steps. The studies include cardiac and skeletal muscle,
natural and artificial markers, and oriental and occidental investigators. They imply that segments as long
as 1 mm may shorten and pause synchronously.
A great advantage of the surface marker approach is
its ability to track a fixed region; the markers adhere to
the fiber surface and the surface adheres to the contractile proteins.' 4 - l5 This is not true of the various striation-based methods (see below), where fiber transla-
tion across the optical axis during contraction results in
a progressive shift in the sampled population, a shift
that could have the potential to wreak havoc6 (but see
rebuttal by Pollack,16 followed by Huxley's 17 rebuttal
of the rebuttal). In the segment method the tracking of
a consistent region averts such potential. Indeed, results have been compiled and submitted for publication
that confirm the method to be fully immune to the
effects of translation, not only in the axial direction but
in the plane normal to the fiber axis as well (Granzieret
al, unpublished data).
A disadvantage, on the other hand, is that the sampled region is perhaps too global to reveal shortening
dynamics on the molecular scale. For this, it is necessary to turn to the sarcomere length detection methods.
Sarcomere Length Dynamics
These approaches are based on the muscle's striation pattern. They fall into two classes — those in
which sarcomere length is inferred from the optical
diffraction pattern and those in which sarcomere length
is measured directly from the striation pattern.
The first to note that a transilluminated muscle fiber
produced a diffraction pattern was Ranvier. l8 By virtue
of the regularity of the pattern of striations, an incident
beam is bent (ie, diffracted) at an angle that is uniquely
dependent on the wavelength of the light and the spatial frequency of the striations. This permits the inference of sarcomere length. Because the sharpness of the
diffraction pattern improves with the purity of the incident beam, the advent of the laser has made the use of
diffraction as common, almost, as the use of the tension transducer.
It is no surprise, therefore, that stepwise shortening
was first identified by the laser diffraction method.1
Indeed, the very first mention of step-like phenomena
came a decade earlier from a Russian group19 who
Pollack Quantal Mechanisms in Contraction
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
reported "sawtooth-like" shortening patterns. In the
absence of modern optical technology, however, their
diffractions records had limited resolution and their
observations went unnoticed.
The widespread use of laser diffraction notwithstanding, this method has come under attack. 5 ' 7 Is it up
to the task of discerning features as fine as steps? The
basis for doubt stems from the question of the extent to
which a muscle fiber may be treated as "thin" diffraction grating. When treated as a thick grating, the influence of striation skew over the fiber thickness enters.20 2I If the skew angle happens to lie at the critical
"Bragg angle," an exceptionally strong reflection will
arise and will dominate the diffraction pattern.22 If
clusters of striations alternately satisfy and fail to satisfy the Bragg condition, it has been argued that apparent steps and pauses could arise.5
This hypothesis runs into several obstacles. First,
direct tests of the Bragg mechanism have proved unsupportive. Since the critical angle is wavelength dependent, each wavelength should reveal a different set
of Bragg planes. But they do not. The pattern of reflections remains wavelength insensitive.23
A second argument is that diffraction provides a
consistent result. When segment length and diffraction
methods are employed on the same fiber during the
same contraction, the records are generally superimposable. Steps and pauses occur with good consistency at
the same position on each of the two shortening waveforms.9 All and all, diffraction works!
The above-mentioned consistency extends also to
the striation imaging methods considered next. These
methods are independent of the thick grating/thin grating controversy; yet the shortening waveforms they
produce contain steps and pauses whose magnitudes
are similar to those found with optical diffraction.
In the first of these two methods a cine camera was
used to photograph the striation pattern at 4000 frames/
sec. The optical diffraction pattern was registered simultaneously. Frame-by-frame analysis of the film
gave shortening patterns that were noisy but clearly
stepwise, and they closely resembled the patterns obtained simultaneously by diffraction.24
Intrigued by the question of whether shortening was
stepwise, Prof. H. Sugi launched a reanalysis of his
own high-speed cine records obtained in a study of the
spread of activation, published well over a decade
ago.25 A micropipette placed near the fiber edge had
been used to inject depolarizing currents. Because contraction was thereby localized, the fiber did not translate and the cine recordings were able to track the
shortening of an unchanging cluster of striations. Although some shortening patterns were smooth, many
showed clear stepwise shortening (Sugi and Toride,
unpublished data).
The seemingly endless number of look-alike frames
required for processing in the cine method prompted
the development of less cumbersome approaches. Following an idea put forth by Claes et al,26 an electronic
sensor was developed from which sarcomere length
could be computed on-line with high resolution.27 The
striation pattern is projected onto a linear photodiode
array. A scan, requiring only several hundred microseconds, produces a periodic signal corresponding to
the A- and I-bands. The frequency of this signal is
computed on the fly using a phase-locked loop, a circuit exquisitely suited to extract the frequency from a
periodic signal that may also contain substantial noise.
The method works well; when used properly, the effect
of fiber translation is insignificant and sarcomere
length resolution is better than 0.1%. 2 8
Records obtained using this method show stepwise
shortening at least as clearly as any of the other methods. An example is shown in Figure 2. Under conditions in which the load remained modest, records
showing cascades of steps similar to those of Figure 2
were found consistently.28
An advantage of this method is that it samples a
highly restricted volume. In Figure 2, for example, the
sampled volume was 30 (JL in length and several
microns either way in breadth. By contrast, the segment method samples a region some 10,000 times
more vast. The presence of pauses and steps in regions
so disparate in size implies that discrete behavior observed in large regions comprises similarly discrete
behavior in small regions, synchronized over space. In
other words, local behavior is stepwise.
To recapitulate: Four classes of methods have been
FIGURE 2. Time course of sarcomere
length change in single fibers of frog
skeletal muscle. Sarcomere length was
obtained using a phase locked-loop circuit to compute the spatial frequency of
striations projected onto a linear photodiode array. Left, unstimulated fiber
subjected to ramp shortening; right,
stimulated fiber contracting against
compliant ends.
20 ms
Circulation Research
brought to bear on the question of whether sarcomeres
shorten in synchronized steps. Stepwise shortening
was observed by all. Three of the four have been repeated and confirmed in more than one laboratory. On
the other hand, an insidious artifact may yet lurk in the
background, awaiting revelation. While such a possibility cannot be ruled out, it is not at all obvious how
the artifact might affect four so diverse methods equally, especially when pairs of methods used simultaneously give the same result.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Is Stepwise Shortening Obligatory?
This question has two parts. Is stepwise shortening
found in all kinds of muscle; and in a given muscle, is
contraction always stepwise?
To the first question, the answer is as yet unknown.
Because steps are found in cardiac and various types of
skeletal muscle, it is tempting to speculate that the
phenomenon may be universal. However, the mechanism underlying stepwise shortening has not yet been
elucidated, and until it is, it will not be possible to
establish which features of the shortening mechanism
are the critical ones for the appearance of steps and
which are not. Thus, the generality of the phenomenon
is difficult to predict.
On the other hand, stepwise phenomena have been
found in an altogether different kind of contractile system, implying that the features giving rise to steps may
indeed be fundamental. Using high-speed cinemicroscopy, Baba studied the time course of bending in the
giant abfrontal cilia of the mussel, Mytilus edulis.29
The bending motion was not continuous; rapid phases
were punctuated by periods of little or no bending.
Each bending step was attributed to a stroke of the
dynein arms, causing a discrete sliding of the axonemal microtubules. The parallelism of this phenomenon
to stepwise sarcomere shortening seems self-evident.
It lends credence to the possibility that the kind of
quantal action considered here may quite possibly constitute a fundamental basis underlying biological
motion.
The other side of the question is whether stepwise
shortening is an obligatory feature in a given specimen. The alternative is that it is not. The question
arises because records obtained under the very same
experimental conditions sometimes show steps and
sometimes do not.28 Can the stepping mechanism be
turned on and off?
Probably not. Under certain conditions stepwise
shortening is practically ubiquitous. For example, in
releases of unstimulated fibers30 and in lightly loaded
isotonic contractions (Tirosh et al, unpublished observations), stepwise shortening is observed in more than
95% of fiber regions. This implies that the failure to
observe steps in the few remaining regions is not likely
to be due to a vagary of the muscle's shortening
mechanism but, perhaps, to some technicality. A likely possibility is incomplete synchrony.
Synchrony is clearly less than infinite. If synchrony
extended the full length of the fiber, the steps would be
as distinct on the muscle shortening trace as on the
Vol 59, No 1, July 1986
sarcomere shortening trace. They are not. Following
release from isometric-to-isotonic contraction, muscle-shortening waveforms exhibit up to several inflections. But these are rounded and less distinct than the
quantitatively comparable inflections (ie, steps) seen
on the sarcomere length traces.31"33 Synchrony is substantial — but incomplete.
Synchrony will be impaired by end effects. Clamping the specimen inevitably causes the load distribution over the fiber cross section to be nonuniform, an
end-effect that may penetrate well into the fiber. This
is especially true in cardiac specimens, where tendons
are in short supply and clamping transforms adjacent
tissue into what is perhaps best described as hamburger.34 Such nonuniformities impair step detectability.
Since the local stepping pattern is a function of load,35
variations in load over the cross section will cause the
steps to go out of phase with one another, thereby
smoothing the detected waveform. When the load is
low or zero, the load variation is small in absolute
terms and the impairment of synchrony will be minimal; but when the load is high, the effect will grow in
significance. This may explain why steps are regularly
detected when the load is low but less regularly detected when the load is high.28
All of this implies that the failure to detect steps on
occasion probably stems from purely technical considerations. The phenomenon may be locally obligatory,
but its detectability will necessarily depend on the extent to which synchrony is retained within the sampled
volume.
Steps and Sounds
A provocative implication of the discontinuous nature of shortening is that associated events may also
occur discontinuously. Consider the consumption of
fuel. If the ATP hydrolytic cycle is as intimately tied to
shortening as presently conceived, 36 ' 37 splitting may
occur as a linked series of explosions, not as a slow,
continuous burn. If so, shortening heat may also
evolve discontinuously. The challenge is to develop
sensors with enough resolution to test for such areas of
potential discreteness.
Discreteness has already been confirmed in the generation of sound. Sound? That contracting muscles
generate sound is easily demonstrated by stuffing your
thumbs into your ears and moving your ringers slowly,
as though waving. Or, for a more dramatic demonstration, place the business end of an electronic stethoscope (in which the frequency components of interest
are amplified) over the first dorsal interosseous muscle
between your thumb and forefinger and move your
forefinger to and from your middle finger. Steel yourself for a deafening roar!
Although it has long been known that muscles generate sound,38 the origin of the sound has remained
obscure. Clearly, sound is associated with movement.
If movement ceases during a pause, the fiber should
remain silent. In testing such an hypothesis, measurements must be confined to a region over which synchrony of steps and pauses is assured. This may have
Pollack Quantal Mechanisms in Contraction
been so in the experiments of Gordon and Holbourn,39
where sounds were detected from the orbicularis oculi,
an eyelid muscle. During a blink, all motor units of this
muscle contract in unison. The sounds came as a series
of distinct clicks, not as a continuous tone.
The discrete clicks have been confirmed in isolated
muscles.40 Spike-like sounds were observed when isolated frog sartorii were allowed to shorten under light
load, confirming that the clicks come from the muscle
itself, not the nerve. The discreteness of the sounds is
consistent with the discreteness of the contractile process; it remains to be demonstrated, however, that the
sounds are coincident with the steps.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Do Cross Bridges Cycle Synchronously?
The critical question regarding the steps must certainly be: What is the underlying mechanism? For what
reason has nature chosen to effect biological motion in
discrete, synchronous steps instead of in a smooth,
continuous manner? Is there some advantage, or is
synchrony merely an incidental by-product of the shortening mechanism?
A logical starting point is the presently accepted
theory, 34 in which contraction is mediated by the action of cross bridges. The theory presupposes that
bridges act independently, but one may imagine a molecular coxswain calling the strokes. A step could then
arise as cross bridges rotate in synchrony. The period
between synchronous rotations would correspond to
the pause. Could this be the underlying mechanism?
Probably not. Steps of similar size and character
appear as the fiber relengthens.41 If steps arise out of
synchronous bridge rotation, the bridges would have to
be endowed with the ability to cycle in reverse.
A more conclusive point is that the steps are observed in sarcomeres stretched to the point where thick
and thin filaments no longer overlap. Their presence
has been confirmed with both the laser diffraction and
phase-locked loop methods.30 Unless the mechanism
underlying steps beyond overlap is qualitatively different from the one operating with overlap, the source of
stepwise shortening cannot lie in synchronous crossbridge rotation.
On the other hand, if cross bridges do rotate, any
proposed mechanism must build in this feature. If
bridge action is stochastic, the task increases in formidability: How can discrete, synchronous behavior be
derived from a process that is random?
The point may be moot. The question of whether
cross bridges rotate is under intensive investigation in
several quarters, and the negativity of early results is
causing a stir (cf Goody42). Among the new probes, the
electron-spin-resonance method provides a "snapshot"
of the distribution of cross-bridge angles.43 The angles
were found to be broadly distributed in relaxed fibers
but restricted to a unique angle in rigor. During contraction, the signal comprised two components indistinguishable from relaxed and rigor. The authors quite
naturally assigned the relaxed component to the unattached bridges and the rigor component to the attached
bridges. The difficulty is that the attached bridges
should not have been caught at a unique angle; if they
had been rotating, the snapshot would have revealed a
species with a new angular distribution.
A more direct approach was taken by Giith,44 who
took advantage of the fact that the fluorescence of the
tryptophan contained within the S-l subunit of the
cross bridge was polarized. Giith examined rabbit
psoas fibers in relaxed, rigor, and contracting states.
Provided the level of substrate was adequate (a problem Giith identified in earlier studies of this type), no
difference of cross-bridge attitude could be found between relaxed and contracting fibers. This result has
recently been confirmed using an extrinsic fluorescence probe, eADP. 45 46
The observations are in concurrence: The expected
pattern of variation of cross-bridge angle is not found.
Similarly, when contracting or rigor fibers are quickstretched or quick-released44"47 the cross-bridge angle
remains stubbornly unchanged. All this notwithstanding, it is possible that bridge rotation does occur as
theorized but is restricted to a small, proximal segment
of the bridge that remains unsampled by any of the
several probes. An altogether different approach, fluctuation analysis, suggests not.
If across bridge cycles, its stroke will be reflected as
a miniscule fluctuation on an otherwise steady tension
signal. If the number of sampled bridges can be restricted, such fluctuations will become increasingly
prominent. (To prove the point, consider an extreme
condition with only two or three bridges: Fluctuations
will dominate the tension signal.) By studying single
myofibrils with diminished overlap and employing a
transducer that was sensitive enough to measure the
tension of a single cross-bridge, Iwazumi was able to
test for the presence of such tension fluctuations.48
Except for the noise induced by building vibrations and
other ambient sources, he found none; the tension signal was flat. Though enough ATP was present to preclude rigor, Iwazumi concluded that cross bridges
could not be cycling.
The inexorable consistency of these results is unsettling. It raises serious doubt about the most fundamental attribute of the accepted contractile mechanism.
Though cross-bridge rotation has been implicit in our
thinking for years, evidence to the contrary is now
accumulating rapidly. The implication for stepwise
shortening is profound: The mechanism need not rest
on asynchronous cross-bridge rotation.
How then might the steps originate?
Toward a Mechanism
An important clue is that the steps appear whether or
not the fiber has been activated. When unstimulated
fibers are stretched or released by a servo-motor, the
resultant sarcomere length changes are stepwise. 4 ' The
stepping mechanism is therefore independent of the
kinetics of activation; it is evidently tied up with the
basic structure.
The structural element responsible for the steps
could reside inside or outside the myofibril. If the
Circulation Research
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
element were outside the myofibril, it might be irrelevant to the contractile mechanism per se, whereas if it
were inside the myofibril, it could be associated with
an element that is integrally involved in contraction.
Outside the myofibril are four potential structures:
connective tissue, cell membrane, sarcoplasmic reticulum, and the recently identified intermediate filament
network.15 None of these elements, however, possess
the regularity that would be necessary to account for
synchrony. Irregular, random structures could not explain the synchronous occurrence of steps and pauses
among huge numbers of myofibrillar sarcomeres. Further, when these elements have been more or less
stripped away by skinning or by paring the preparation
to a small bundle of myofibrils, the steps persist.41-49
Inside the myofibril, on the other hand, the degree
of structural regularity is high; crystallinity is the hallmark of myofibrillar architecture, as any well-prepared electron micrograph or x-ray diffraction pattern
demonstrates. A shortening step could arise if some
axial dimension within the lattice changed discretely.
How might such a quantal change of length arise?
The obvious first choice is relative sliding of thin
over thick filaments. Regularly positioned resistance
points could give rise to brief shortening pauses. Simplicity notwithstanding, this hypothesis is probably inadequate, for the steps found in fibers stretched beyond
overlap would need to be explained by an entirely
different class of mechanism.
An alternative possibility within the myofibril is a
stepwise change of filament length. Which filament?
When a fiber is stretched beyond overlap, continuity
between Z-lines is not lost. It is retained through a set
of filaments that interconnect each end of the thick
filament with the nearest Z-line. When the fiber is
stretched, these "connecting filaments" are likewise
stretched. Probably they support much of the resting
tension.50
Connecting filaments are not well-known; they are
ignored in most textbooks. Yet they are well-studied
and apparently uncontroversial. Connecting filaments
were identified long ago by Huxley and Peachey,51
confirmed by Sjostrand52 and by Carisen et al,53 examined for structural detail by Locker and Leet,54 and
characterized biochemically in a series of experiments
from the laboratories of K. Maruyama and K. Wang
(cf Locker55 for review).
If the length of the thick filament remains constant
in an unactivated fiber, imposed length changes must
be conferred fully on the connecting filaments. This is
an inescapable feature of the filament arrangement. If
the sarcomere length change occurs stepwise, the
change of connecting filament length must also be
stepwise.
We see, therefore, that the stepping mechanism
likely resides in the connecting filaments, at least in
the unstimulated fiber. No other element changes
length. This inference is consistent with the structure.
Isolated connecting filament proteins are threadlike
but often contain isolated nodules distributed along
their length, where the threadlike material appears to
Vol 59, No I, July 1986
have gathered up. 56 •" These nodules may represent
discrete, localized regions of shortening.
All well and good. But this cannot be the complete
answer. Stepwise behavior remains in evidence at sarcomere lengths far below those at which resting tension is present35 — lengths at which the connecting
filaments would presumably be slack. Another element must be responsible for these steps.
Since experiments have consistently shown that thin
filaments do not shorten, the remaining element is the
thick filament. The suggestion that thick filaments
shorten in steps, or for that matter shorten at all, is
obviously at odds with accepted views. However, the
classical observations of Huxley and Hanson58 and
Huxley and Niedergerke59 have not been broadly confirmed. On the contrary, a surprisingly large number of
papers published since the mid-fifties have reported Aband shortening or thick-filament shortening of substantial magnitude (for review, cf Pollack60). The issue
is far from settled, and I feel it would be premature to
discount this possibility out of hand.
Strongly against the thick filament shortening
hypothesis has been the observation that the axial reflections on the x-ray diffraction pattern do not shift as
the sarcomere shortens.61 Thick filaments cannot
"shrink" longitudinally. However, segmental shortening is by no means excluded. If each shortening step
were restricted to a small, distinct segment along the
filament and if the number of shortened segments remained modest, this phenomenon could well escape
detection by x-ray methods. The hypothesis remains
plausible.
By elimination, we have arrived at the hypothesis
that stepwise length changes are mediated by length
changes of two elements, connecting filaments and
thick filaments. Connecting filaments would account
for the steps when the load is increased or decreased in
fibers that are unactivated; in activated fibers, with thin
filaments bound to cross bridges, steps would be generated by the thick filaments. The division of labor is
thus based solely on whether the sarcomere has or has
not been activated.
Could steps of similar nature plausibly arise out of
length changes of two separate structures? Does this
hypothesis seem, perhaps, too complicated? Intuition
says yes. But modern electron micrographs lend credence to this proposition. The thick filament-connecting filament combination appears not as two distinct
filaments but as one long filament running from Z-line
to Z-line, a filament whose mid-region (thick filament)
is slightly thicker than its flanking regions (connecting
filaments). From this perspective, it is easy to envision
discrete length changes arising from any region along
the length of this compound filament. Two apparent
sources merge effectively into one — variations on a
single theme.
The proposed hypothesis obviously represents a departure from accepted views, one that is perhaps more
radical than may be apparent on the surface. If thick
filaments shorten and do work, is it necessary to invoke cross-bridge cycling as the sole work-producing
Pollack Quanta! Mechanisms in Contraction
mechanism? And in light of recent negative evidence
for angle changes of S-l during contraction (cited
above), is it reasonable to invoke cross-bridge cycling
at all? Could thick filament shortening accomplish the
entire task?
These are questions for the future; they have not
been settled here. What has been settled here, I hope,
is the question of whether the shortening process is
quantal. Stepwise shortening has now been confirmed
in several laboratories using four different classes of
methods. The phenomenon, unlike the mechanism
proposed to account for it, is therefore out of the speculative arena. The steps are waiting to be scaled.
References
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
1. Pollack GH, Iwazumi T, ter Keurs HEDJ, et al: Sarcomere
shortening in striated muscle occurs in stepwise fashion. Nature 1977;268:757-759
2. Katz B: Nerve. Muscle, and Synapse. New York, McGrawHill, 1966
3. Huxley AF: Muscle structure and theories of contraction. Prog
Biophys Biophys Chem 1957;7:255-318
4. Huxley HE: The mechanism of muscular contraction. Science
1969; 164:1356-1366
5. Riidel R, Zite-Frenczy F: Do laser diffraction studies on striated muscle indicate stepwise sarcomere shortening? Nature
1979;278:573-575
6. Altringham JD, Bottinelli R, Lacktis JW: Is stepwise sarcomere shortening an artifact? Nature 1984;307:653-655
7. Goldman YE, Simmons RM: Control of sarcomere length in
skinned muscle fibres of Rana Temporaria during mechanical
transients. J Physiol 1984;350:497-518
8. Gordon AM, Huxley AF, Julian FJ: Tension development in
highly stretched vertebrate muscle fibers. J Physiol (Lond)
1966; 184:143-169
9. Granzier HLM, Pollack GH: Simultaneous segment length and
sarcomere length measurements confirm stepwise shortening.
Biophys J 1985 ;47:289a
10. Edman KAP, Hoglund O: A technique for measuring length
changes of individual segments of an isolated muscle fiber. J
Physiol 1981;317:8-9p
11. Edman KAP, Elzinga G, Noble MIM: Residual force enhancement after stretch of contracting frog single muscle fibers. J
Gen Physiol 1982;80:769-784
12. Housmans P: Discussion, in Pollack GH, Sugi H (eds): Contractile Mechanisms in Muscle. New York, Plenum Press,
1984, pp 782-784
13. Tameyasu T, Toyoki T, Sugi H: Non-steady motion in unloaded contractions of single frog cardiac cells. Biophys J
1985;48:461-465
14. Street S: Lateral transmission of tension in frog myofibers: A
myofibrillar network and transverse cytoskeletal connections
arc possible transmitters. J Cell Physiol 1983;114:346-363
15. Wang K, Ramirez-Mitchell R: A network of transverse and
longitudinal intermediate filaments is associated with sarcomeres of adult vertebrate skeletal muscle. J Cell Biol 1983;96:
562-570
16. Pollack GH: Response to "Is stepwise sarcomere shortening an
artifact?" Nature 1984;309(5970):7l2-714
17. Huxley AF: Response to "Is stepwise sarcomere shortening an
artifact?" Nature 1984;309(5970):713-714
18. Ranvier L: Du spectra produit par les muscles stries. Arch de
Physiol 1874;774-780
19. Emel'yanov VB, Efimov VN, Frank GM: A diffraction investigation of "sliding" during muscle contraction. Dokl Akad
NaukSSR 1966;167(4):923-924
20. Yeh Y, Baskin RJ, Lieber RL, et al: Theory of light diffraction
by single skeletal muscle fibers. Biophys J 1980;29:509-522
21. Judy MM, Summerour V, LeConey T, etal: Muscle diffraction
theory: Relationship between diffraction subpeaks and discrete
sarcomere length distributions. Biophys J 1982:37:475-487
22. Riidel R, Zite-Ferenczy F: Interpretation of light diffraction by
cross-striated muscle as Bragg reflexion of light by the lattice
of contractile proteins. J Physiol 1979;290:317-330
23. Leung AF: Light diffractometry for determining the sarcomere
length of striated muscle: An evaluation. J Muscle Res Cell
Motil 1983;4:473-483
24. Delay MJ, Ishide N, Jacobson RC, et al: Stepwise sarcomere
shortening: Analysis by high-speed cinemicrography. Science
1981;213:1523-1525
25. Sugi H: Local activation of frog muscle fibers with linearly
rising currents. J Physiol 1968;199:549-567
26. Claes VA, deClerck NM, VanOcken ER, et al: New method of
sarcomere measurements. Fed Proc 1980;39:1729
27. Myers J, Tirosh R, Jacobson RC, et al: Phase locked loop
measurement of sarcomere length with high time resolution.
IEEE Trans Biomed Eng 1982;29(6):463-466
28. Jacobson RC, Tirosh R, Delay MJ, et al: Quantitized nature of
sarcomere shortening steps. J Muscle Res Cell Motil 1983;4:
529-542
29. Baba S: Regular steps in bending cilia during the effective
stroke. Nature 1979;282:717-720
30. Granzier HLM, Pollack GH: Stepwise shortening in unstimulated skeletal muscle fibers. J Physiol 1985;362:173-188
31. Sugi H, Tsuchiya T: Isotonic velocity transients in frog muscle
fibres following quick changes in load. J Physiol 1981;319:
219-238
32. Civan MM, Podolsky RJ: Contraction kinetics of striated muscle fibers following quick changes in load. J Physiol 1966;
184:511-534
33. Gulati J, Podolsky RJ: Contraction transients of skinned muscle fibers: effects of calcium and ionic strength. J Gen Physiol
1978,72:701-716
34. Krueger JW, Pollack GH: Myocardial sarcomere dynamics
during isometric contraction. J Physiol (Lond) 1974;251:
627-643
35. Vassallo DV, Pollack GH: The force-velocity relation and
stepwise sarcomere shortening in cardiac muscle. Circ Res
1982;51(l):37-42
36. Lymn RW, Taylor EW: Mechanism of adenosine triphosphate
hydrolysis by actomyosin. Biochemistry 1971; 10:4617-4624
37. Eisenberg E, Greene LE: The relation of muscle biochemistry
to muscle physiology. Ann Rev Physiol 1980;42:293-309
38. Wollaston WH: On the duration of muscle action. Phil Trans
Rev Soc Lond Biol Soc 1810; 1-5
39. Gordon G, Holbourn AHS: The sounds from single units in a
contracting muscle. J Physiol 1948;107:456-464
40. Brozovich F, Pollack GH: Muscle contraction generates discrete sound bursts. Biophys J 1983;41:35-40
41. Pollack GH, Tirosh R, Brozovich FV, et al: Stepwise shortening: Evidence and implications, in Pollack GH, Sugi H (eds):
Contractile Mechanisms in Muscle. New York, Plenum Press,
1984, pp 765-786
42. Goody RS: The European Molecular Biology Organization/
Muscular Dystrophy Association Workshop on the structure of
actin and myosin and the mechanism of their interaction. Alpbach, March 1983. J Muscle Res Cell Motil 1983;4:391-394
43. Cooke R, Crowder MS, Thomas DD: Orientation of spin labels
attached to cross-bridges in contracting muscle fibers. Nature
1982;300:776-778
44. Giith K: Polarization of tryptophan fluorescence measurements
in muscle: a reevaluation. Biophys Struct Mech 1980;6:81-93
45. Yanagida T: Angles of fluorescently labelled myosin heads and
actin monomers in contracting and rigor strained muscle fiber,
in Pollack GH, Sugi H (eds): Contractile Mechanisms in Muscle. New York, Plenum Press, 1984, pp 397-411
46. Yanagida T: Angle of active site of myosin heads in contracting
muscle during sudden length changes. J Muscle Res Cell Motil
1985;6:43-52
47. Cooke R: Stress does not alter the conformation of a domain of
the myosin cross-bridge in rigor muscle fibers. Nature 1981;
294:570-571
48. Iwazumi T: No tension fluctuation in normal and healthy single
myofibrils. Biophys J 1984;45:158a
49. Gordon AM, Pollack GH: Effects of calcium on the sarcomere
Circulation Research
50.
51.
52.
53.
54.
length-tension relation in rat cardiac muscle: Implications for
the Frank-Starling mechanism. Cire Res 1980;47:610-619
Magid A, Ting-Beall HP, Carvell M, et al: Connecting filaments, core filaments, and side-struts: A proposal to add three
new load-bearing structures to the sliding filament model, in
Pollack GH, Sugi H (eds): Contractile Mechanisms in Muscle.
New York, Plenum Press, 1984, pp 307-328
Huxley AF, Peachey LD: The maximum length for contraction
in vertebrate striated muscle. J Physiol 1961 ;156:150—165
Sjostrand FS: The connections between A- and I-band filaments in striated frog muscle. J Ultrastruct Res 1962;7:
225-246
Carlsen F, Fuchs F, Knappels GG: Contractility and ultrastructure in glycerol-extracted muscle fibers. J Cell Biol 1965;27:
35-46
Locker RH, Leet NG: Histology of highly stretched beef muscle. II. Further evidence on the location and nature of gap
filaments. J Ultrastruct Res 1976;55:157-172
KEY WORDS
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
diffraction
Vol 59, No I, July 1986
55. Locker RH: The role of gap filaments in muscle and meat.
Food Microstructure 1984;3:17-32
56. Trinick J, Knight P, Whiting A: Purification and properties of
native titin. J Mol Biol 1984;180:331-356
57. Wang K, Ramirez-Mitchell R, Palter D: Titin is an extraordinarily long, flexible, and slender myofibrillar protein. Proc
Natl Acad Sci USA 1984;81:3685-3689
58. Huxley HE, Hanson J: Changes in the cross-striations of muscle during contraction and stretch and their structural interpretation. Nature 1954; 173:973-976
59. Huxley AF, Niedergerke R: Structural changes in muscle during contraction. Interference microscopy of living muscle fibers. Nature 1954; 173:971-973
60. Pollack GH: The slidingfilament/cross-bridgetheory: A critical review. Physiol Rev 1983;63:1049-1113
61. Huxley HE, Brown W: The low-angle x-ray diagram of vertebrate striated muscle and its behavior during contraction and
rigor. J Mol Biol 1967;30:383-434
• stepwise shortening • cross-bridge cycling
• thick-filament shortening
synchrony
• optical
Quantal mechanisms in cardiac contraction.
G H Pollack
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Circ Res. 1986;59:1-8
doi: 10.1161/01.RES.59.1.1
Circulation Research is published by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231
Copyright © 1986 American Heart Association, Inc. All rights reserved.
Print ISSN: 0009-7330. Online ISSN: 1524-4571
The online version of this article, along with updated information and services, is located on the
World Wide Web at:
http://circres.ahajournals.org/content/59/1/1.citation
Permissions: Requests for permissions to reproduce figures, tables, or portions of articles originally published in
Circulation Research can be obtained via RightsLink, a service of the Copyright Clearance Center, not the
Editorial Office. Once the online version of the published article for which permission is being requested is
located, click Request Permissions in the middle column of the Web page under Services. Further information
about this process is available in the Permissions and Rights Question and Answer document.
Reprints: Information about reprints can be found online at:
http://www.lww.com/reprints
Subscriptions: Information about subscribing to Circulation Research is online at:
http://circres.ahajournals.org//subscriptions/