The efficiency of contraction in rabbit skeletal muscle fibres ...

J. Physiol. 517.3 Efficiency of muscle contraction
851
is initially faster than at 2·7 ìm, it becomes slower after
•0·5 s and the two traces cross over. The period of constant
ATPase rate is a feature of the conditions used here (2·7 ìm
and 12 °C). At the same sarcomere length, but at 15 °C, the
period of steady ATPase rate is not so clear (data not
shown). The ATPase rate in the first turnover is also
sensitive to temperature. At a sarcomere length of 2·4 ìm,
the ATPase rate increased from 15·2 ± 0·7 to 19·8 ± 2·2 s¢
(n = 5) when temperature was raised from 12 to 15 °C. At a
sarcomere length of 2·7ìm, the same temperature rise
caused the ATPase rate constant during the first turnover
to increase from 10·5 ± 1·0 to 15·4 ± 1·8 s¢ (n = 5). A
temperature rise of only 3 °C increases the rate constants by
factors of 1·3 and 1·47 for sarcomere lengths of 2·4 and
2·7 ìm, respectively.
DISCUSSION
The ATPase rate constant during the first turnover of
the ATPase
The experiments were carried out on muscle fibres which
had been treated with Triton X-100 as well as permeabilized
by treatment with glycerol. In consequence, the ATPase
activity remaining in the muscle fibre is attributed solely to
that of the actomyosin, as was shown previously (He et al.
1997) on the basis of its sarcomere length dependence, Ca¥
sensitivity and resistance to sarcoplasmic ATPase inhibitors.
At 12 °C, the mean ATPase rates during the first turnover
were 10·3 s¢, when referred to an active site concentration
of 0·14 mÒ, at a sarcomere length of 2·7 ìm and 15·2 s¢ at
2·4 ìm. This latter value is slightly lower than that reported
earlier (18·8 s¢) for psoas muscle fibres at 12 °C and a
sarcomere length of 2·4 ìm, referred to an active site
concentration of 0·15 mÒ (He et al. 1997). At 15 °C and a
sarcomere length of 2·4 ìm, the ATPase rate during the
first turnover (19·8 s¢) is lower than the value of 31·5 s¢
reported by He et al. (1998b). The high temperature
sensitivity of the ATPase in this range and possible
variations between fibres may explain this difference.
Decay of the ATPase rate constant following
photolysis of caged ATP
He et al. (1997, 1998b) reported a high initial ATPase rate
which gradually declines during the first few hundred
milliseconds following the photolytic release of ATP in
isometrically contracting muscle fibres. He et al. (1998b)
showed that the ATPase rate constant decayed gradually
with time for both soleus and psoas muscle fibres, and that
the decay was less marked in the presence of ADP. We show
here that the decay in the ATPase rate constant depends on
experimental conditions, in that, at a long sarcomere length
(2·7 ìm) and at 12 °C, a period is seen during which the
ATPase rate constant remains relatively constant, as
reported previously (Fig. 7ofHeet al. 1997). Here, we use
the period of steady ATPase to apply length changes, and
to study the corresponding changes in ATPase rate. The
conditions are therefore useful in that the observed changes
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in response to shortening are largely independent of the
time when the shortening was applied. The difference in
behaviour seen at short and long sarcomere lengths in rabbit
fibres may be related to the extent and the time course of
fibre shortening seen at different lengths.
Relationship between the ATPase rate constant
during the first turnover, in the isometric state and
that seen during filament sliding
We investigate below the effect of sarcomere shortening on
the ATPase rate in the early phases of contraction, when
the fibre length is constant. The high ATPase rate seen
previously during the first turnover (He et al. 1997, 1998b)
was not thought to be attributable to sarcomere shortening
in the initial phases of contraction, but in those experiments
sarcomere length measurements were not performed. Here,
we show sarcomere length measurements from the same
region of the fibre from which fluorescence signals are
obtained. Although the diffraction signal indicated that
sarcomere length changes occurred following the photolytic
release of ATP, the signal was variable, sometimes showing
either lengthening or shortening. In most cases, the velocity
of sarcomere shortening was slow and would not be
expected to cause a large acceleration in the ATPase rate,
unlike that seen in response to applied shortening. For
shortening to account for the ATPase rate in the first
turnover, it can be calculated from the hyperbola in Fig. 8
that the shortening velocity during the first turnover should
be 0·22 ML s¢. The mean of the data in Fig. 3 (0·09 ML s¢)
argues against this possibility. However, the hyperbola in
Fig. 8 shows that shortening at a velocity of 0·09 ML s¢
during the nominally isometric phase of contraction is
accompanied by an ATPase rate of 8 s¢, 22 % less than the
measured value of 10·3 s¢. The difference between the
observed ATPase rate constant and the ATPase rate
constant which can be explained by a period of shortening is
thereforesmallanditcannotbeexcludedthatoursarcomere
measurements fail to report precisely the nature of
sarcomere shortening in the field of view. For example, the
laser diffraction signal provides a mean shortening velocity,
but the presence of a small fraction of rapidly shortening
sarcomeres may increase the ATPase rate and not be
detected by our apparatus. It may also be that some
sarcomere jitter, or individual sarcomeres shortening rapidly
at the expense of their neighbours accounts for an increase
in ATPase rate. More recent experiments using videomicroscopy
of muscle fibres during experiments have
confirmed the accuracy of our laser diffraction method, but
do not improve the spatial resolution of the measurements
and do not exclude local shortening.
However, if the ATPase rate during the first turnover was
higher than in the steady state because of transient
shortening, the mean shortening velocity during the first
turnover (0·09 ML s¢, n = 29) would cause an increase in
the ATPase rate from 5·1 to 8·0 s¢ (calculated from the
hyperbola to the data shown in Fig. 8). This value is less