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G H Pollack

Publications and source records attributed to G H Pollack.

At least 55 records · Page 3Linked to original sources
PubMed1993

Spontaneous sarcomeric oscillations at intermediate activation levels in single isolated cardiac myofibrils.

Spontaneous oscillations observed in various heart muscle preparations are widely thought to be triggered by spontaneous release of Ca2+ from the sarcoplasmic reticulum (SR). Here, we report undamped propagated oscillations that occur in the absence of SR. In single cardiac myofibrils treated with Triton X-100 to remove SR and held isometrically, partial activation initiated periodic fluctuations of sarcomere length persisting up to 1 hour. Oscillation characteristics could be readily quantitated by virtue of the small size of the preparation. In an individual sarcomere, the oscillation cycle generally consisted of a slow shortening phase, followed by a phase of rapid lengthening. Oscillations usually propagated along the myofibril--frequently along the entire specimen--in a wavelike fashion (average velocity, 12.3 microns/s at 10 degrees C; Q10, approximately 1.3). The oscillation period was 2.30 and 1.72 seconds at 10 degrees and 20 degrees C, respectively, and was insensitive to stretch. The average oscillation amplitude, which was temperature independent, decreased with stretch from more than 20% of the mean sarcomere length at lengths below 2 microns to zero beyond a sarcomere length of 3 microns. Stiffening of the Z line by labeling with anti-alpha-actinin resulted in a dose-dependent decrease of oscillation amplitude, while the period was not affected. Tension oscillations could not be detected in single myofibrils but were frequently detectable in myofibril doublets, where the oscillation magnitude (approximately 1 microgram) was above the noise floor. Addition of 10 mumol/L ryanodine to the activating solution did not alter oscillation characteristics, as expected, since the oscillations are unrelated to SR calcium release. On the basis of our results, we consider a mechanism for the oscillations in which a length dependence of myofibrillar Ca2+ sensitivity and a dynamic Z-line structure are essential.

Actinin↗
PubMed1993

I-band periodicity in rabbit psoas muscle fibers measured using digital image-analysis techniques.

The fine periodicity of I-band was measured in single fibers of glycerinated rabbit psoas muscle, in relaxed, activated and rigor states at short (2.5 microns) and long (3.2 microns) sarcomere lengths. Measurements were carried out using fast Fourier transforms obtained from I-band regions of digitized electron micrographic images. The mean periodicity, averaged among all conditions, was 42.5 +/- 1.5 nm. Some differences were found among the different conditions, but they were too small to be conclusive. The observed periodicity value was considerably higher than the generally assumed value of 38.5 nm, and is surprisingly close to the myosin-repeat spacing.

Actins↗
PubMed1992

Effect of small release on force during sarcomere-isometric tetani in frog muscle fibers.

We investigated the effect of small shortening imposed on frog muscle fibers during sarcomere-isometric tetani. Sarcomere length was initially kept constant, then slightly shortened (1%-5% of initial length) and clamped again for the remainder of the tetanus. Force level after the shortening was higher than the force level preceding the release. The size of the increase was larger than that predicted by the descending limb of the linear force-length relation. The difference between measured and predicted force levels increased with sarcomere length. At a sarcomere length of 3.2 microns, the force level after the shortening was higher by 50% than the force level expected from the linear descending limb. Dispersion of sarcomere-length within the sampled region was measured by two independent methods: striation imaging and analysis of the intensity profile of the first diffraction order. Sarcomere-length inhomogeneity in the sampled region was too small (standard deviation from the average sarcomere-length was +/- 0.03 microns) to account for the size of the increase in force. We studied the dependence of increase in tetanic force level after small sarcomere-length release on the size, velocity and timing of the release, as well as on initial sarcomere-length. Release size was the major determinant of the amount of increase in force. Release of 20 nm per half sarcomere was sufficient to produce an almost full force increase. Larger releases increased the force only moderately. Over the range studied, release velocity and timing had little or no effect.

Animals↗
PubMed1991

Nature and origin of gap filaments in striated muscle.

Immunoelectron microscopy was used to study the nature and origin of 'gap' filaments in frog semitendinosus muscle. Gap filaments are fine longitudinal filaments observable only in sarcomeres stretched beyond thick/thin filament overlap: they occupy the gap between the tips of thick and thin filaments. To test whether the gap filaments are part of the titin-filament system, we employed monoclonal antibodies to titin (T-11, Sigma) and observed the location of the epitope at a series of sarcomere lengths. At resting sarcomere length, the epitope was positioned in the I-band approximately 50 nm beyond the apparent ends of the thick filament. The location did not change perceptibly with increasing sarcomere length up to 3.6 microns. Above 3.6 microns, the span between the epitope and the end of the A-band abruptly increased, and above 4 microns, the antibodies could be seen to decorate the gap filaments. Between 5 and 6 microns, the epitope remained approximately in the middle of the gap. Even with this high degree of stretch, the label remained more or less aligned across the myofibril. The abrupt increase of span beyond 3.6 microns implies that the A-band domain of titin is pulled free of its anchor points along the thick filament, and moves toward the gap. Although this domain is functionally inextensible at physiological sarcomere length, the epitope movement in extremely stretched muscle shows that it is intrinsically elastic. Thus, the evidence confirms that gap filaments are clearly part of the titin-filament system. They are derived not only from the I-band domain of titin, but also from its A-band domain.

Animals↗
PubMed1990

Immunoelectron microscopic observations on tropomyosin localization in striated muscle.

Tropomyosin localization in striated muscle was studied by means of immunoelectron microscopy. Polyclonal and monoclonal antibodies to tropomyosin were allowed to diffuse into mechanically skinned single fibres dissected from frog semitendinosus muscle. Antibodies produced transverse I-band stripes with the expected periodicity of 38 nm. However, some differences were revealed among the various antibodies. While polyclonal antibodies generally showed 23 stripes, monoclonal antibodies showed an extra 24th stripe immediately adjacent to the Z-line, implying some structural/functional uniqueness of this terminal tropomyosin. Furthermore, the stripes did not always lie parallel to the Z-line. When the Z-line was straight or slightly skewed, the stripes generally were parallel to it. However, when Z-line skew was more severe, the stripes remained perpendicular to the fibre axis, indifferent to the Z-line skew. This may implay that the coupling of tropomyosin to the thin filament is not tight. Finally, the monoclonal antibodies themselves exerted an anomalous effect on the Z-line, apparently extracting or shifting some of its mass.

Animals↗
PubMed1990

A-band shortening in single fibers of frog skeletal muscle.

The question of whether A-bands shorten during contraction was investigated using two methods: high-resolution polarization microscopy and electron microscopy. During shortening from extended sarcomere lengths in the passive state, sarcomere-length changes were essentially accounted for by I-band shortening. During active shortening under otherwise identical conditions, the sarcomere length change was taken up approximately equally by A- and I-bands. Several potential artifacts that could give rise to apparent A-band shortening were considered and judged unlikely. Results obtained with polarization microscopy were similar to those obtained with electron microscopy. Thus, modest but significant thick filament shortening appears to occur during active sarcomere shortening under physiological conditions.

Animals↗
PubMed1990

The descending limb of the force-sarcomere length relation of the frog revisited.

1. We studied the descending limb of the force-sarcomere length relation in single frog muscle fibres using sarcomere isometric contractions. 2. Sarcomere length was measured simultaneously with two independent methods: a laser diffraction method and a segment length method that detects the distance between two markers attached to the surface of the fibre, about 800 microns apart. Both methods were used to keep sarcomeres at constant length during contraction. 3. Fibres were selected for low resting tension since it was known from previous experiments that for such fibres the force developed by fixed-end tetani is much higher than that predicted by the degree of filament overlap. 4. With fixed-end tetani, force decline with increase of sarcomere length was small between 2.0 and 3.0 microns. At a sarcomere length of 3.0 microns, force was about 90% of maximal. 5. With sarcomere isometric tetani, force was considerably lower than with fixed-end tetani. Force was maximal at about 2.1 microns and decreased to zero at about 3.6 microns. At intermediate lengths the descending limb was within 80 nm of the values predicted from filament overlap. 6. We investigated why force of fixed-end contractions was much higher than that generated by sarcomere isometric contractions. 7. During the force plateau of fixed-end tetani at sarcomere lengths longer than about 2.0-2.2 microns, sarcomeres in the fibres mid-region were not isometric, but instead stretched slowly. By measuring the force-velocity relation it was shown that this slow stretch elevates active force well beyond sarcomere isometric force. 8. Stretch of the central region was also observed during the tetanic force rise. This was shown to result in an increase of passive force that grew larger at longer sarcomere lengths. At about 3.6 microns the increase of passive force was similar to the total force generated by fixed-end contractions at this length. 9. Laser diffraction and segment length methods gave the same results, diminishing the chance that any systematic artifact underlies our findings. 10. While earlier experiments from this laboratory carried out on fibres held at constant length during contraction did not reveal a linear descending limb, the present results support the linear descending limb as a characteristic feature of isometrically contracting sarcomeres.

Animals↗
PubMed1990

Sarcomere dynamics during isotonic velocity transients in single frog muscle fibers.

If the load on a tetanized fiber is abruptly changed to a new steady value, the ensuing fiber length change shows the well-known "isotonic velocity transient," in which the velocity oscillates before settling at some steady value. We studied sarcomere dynamics during these transients using two methods: optical diffraction and a segment-length method. Our principal aim was to determine whether these transients might be a reflection of the fact that sarcomere shortening is often found to be stepwise. We found that pauses in sarcomere shortening occurred during the low-velocity phases of the transient and that steps of sarcomere shortening occurred during the high-velocity phases. Thus the isotonic transient appears to arise from the steps. In addition to the isotonic transient, we studied the well-known isometric transient, in which fiber length is abruptly changed, and ensuing tension response is measured. Again, we found that the transient may be a reflection of the stepwise shortening pattern.

Animals↗
PubMed1989

Sarcomere length dependence of the force-velocity relation in single frog muscle fibers.

The force-velocity relation of single frog fibers was measured at sarcomere lengths of 2.15, 2.65, and 3.15 microns. Sarcomere length was obtained on-line with a system that measures the distance between two markers attached to the surface of the fiber, approximately 800 microns apart. Maximal shortening velocity, determined by extrapolating the Hill equation, was similar at the three sarcomere lengths: 6.5, 6.0, and 5.7 microns/s at sarcomere lengths of 2.15, 2.65, and 3.15 microns, respectively. For loads not close to zero the shortening velocity decreased with increasing sarcomere length. This was the case when force was expressed as a percentage of the maximal force at optimal fiber length or as a percentage of the sarcomere-isometric force at the respective sarcomere lengths. The force-velocity relation was discontinuous around zero velocity: load clamps above the level that kept sarcomeres isometric resulted in stretch that was much slower than when the load was decreased below isometric by a similar amount. We fitted the force-velocity relation for slow shortening (less than 600 nm/s) and for slow stretch (less than 200 nm/s) with linear regression lines. At a sarcomere length of 2.15 microns the slopes of these lines was 8.6 times higher for shortening than for stretch. At 2.65 and 3.15 microns the values were 21.8 and 14.1, respectively. At a sarcomere length of 2.15 microm, the velocity of stretch abruptly increased at loads that were 160-170% of the sarcomere isometric load, i.e., the muscle yielded. However, at a sarcomere length of 2.65 and 3.15 microm yield was absent at such loads. Even the highest loads tested (260%) resulted in only slow stretch. It is concluded that properties of the force generators change with sarcomere length. This is not anticipated by the cross-bridge model of muscle contraction.

Animals↗
PubMed1989

Effect of active pre-shortening on isometric and isotonic performance of single frog muscle fibres.

1. We studied the effects of shortening history on isometric force and isotonic velocity in single intact frog fibres. Fibres were isometrically tetanized. When force reached a plateau, shortening was imposed, after which the fibre was held isometric again. Isometric force after shortening could then be compared with controls in which no shortening had taken place. 2. Sarcomere length was measured simultaneously with two independent methods: a laser-diffraction method and a segment-length method that detects the distance between two markers attached to the surface of the fibre, about 800 microns apart. 3. The fibre was mounted between two servomotors. One was used to impose the load clamp while the other cancelled the translation that occurred during this load clamp. Thus, translation of the segment under investigation could be minimized. 4. Initial experiments were performed at the fibre level. We found that active preshortening reduced isometric force considerably, thereby confirming earlier work of others. Force reductions as large as 70% were observed. 5. Under conditions in which there were large effects of shortening at the fibre level, we measured sarcomere length changes in the central region of the fibre. These sarcomeres shortened much less than the fibre's average. In fact, when the load was high, these sarcomeres lengthened while the fibre as a whole shortened. Thus, while the fibre-length signal implied that sarcomeres might have shortened to some intermediate length, in reality some sarcomeres were much longer, others much shorter. 6. Experiments performed at the sarcomere level revealed that isometric force was unaffected by previous sarcomere shortening provided the shortening occurred against either a low load or over a short distance. However, if the work done during shortening was high, force after previous shortening was less than if sarcomeres had remained at the final length throughout contraction. The correlation between the force deficit and the work done during shortening was statistically significant (P = 0.0001). 7. Interrupting the tetanus for 0.5-3.0 s did not reverse the effects of shortening on isometric force; at least 5-10 min of rest were required before force recovered completely. 8. Sarcomeres accelerated during the period of shortening under constant load, indicating that the sarcomeres became progressively stronger. However, the acceleration was less than that predicted from the force-velocity relation applicable at each of the sarcomere lengths transversed during shortening. 9. Velocity of shortening appeared to be much more sensitive to previous shortening than isometric force. 10. Results obtained with the diffraction method were the same as those obtained with the segment method.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

I-bands of striated muscle contain lateral struts.

In electron micrographs of striated muscle, the I-band often shows a distinct cross-striation. The periodicity of this striation is near 40 nm and has been attributed to troponin, which is localized along the thin filament. However, the cross-striation is often so prominent as to be suggestive of physical structures running transversely across the I-band. We examined I-band ultrastructure using three independent methods: thin sections of chemically fixed specimens; freeze-fracture; and freeze-substitution. With all three methods we found transverse structures distributed throughout the I-band, many of which bridged the gap between neighboring filaments. Such structures were observed in each of the several species studied. In fish muscle in particular, which has a highly regular lattice, it was obvious that these structures gave rise to the observed periodicity.

Animals↗
PubMed1988

Thick filaments of striated muscle are laterally interconnected.

Earlier reports from this and other laboratories indicated that thick filaments may be interconnected along their length by rung-like structures. This study was carried out to test whether these interconnections are genuine structures; whether they appear in different muscle types; and whether they arise from myosin cross-bridges. We studied insect flight muscles because of their well-known ultrastructure, and frog heart and rabbit psoas muscles secondarily. Ultrastructure was examined with freeze-fracture; with conventionally prepared thin sections; and with negative stain. All three methods showed rung-like interconnections between thick filaments. The interconnections spanned the length of the cross-bridge zone, i.e., along all but the central bare zone of the thick filament. They were observed consistently in relaxed, activated, and rigor states. We considered potential artifacts that might cause apparent interconnections where none existed in vivo, but were unable to identify a source of artifact common to all methods. Several features of the interconnections imply that for the most part they may be composed of S-1 heads from adjacent thick filaments binding to one another at their tips.

Animals↗
PubMed1988

Effect of tension on the rigor cross-bridge angle.

The effect of resting tension and external force on the rigor crossbridge angle was investigated in insect flight muscle (Honeybee, Apis mellifera). In the presence of resting tension, bridges were either perpendicular to the filament axis or tilted towards the M-line. In shortened, slack muscle, bridges remained perpendicular or were tilted towards the Z-line. Thus, the rigor bridge angle appears to depend on the state of the thick filament. With the thick filament under stress, the angle may be quite different than when it is slack. On the other hand, external force applied directly to the rigor bridges did not change their configuration. Bridge angle remained unchanged, irrespective of the amount of tension applied to the bridge. This implies that a very tight bond exists between the rigor bridge and thin filament, allowing essentially no rotation even with a large applied force.

Animals↗
PubMed1988

Pauses, steps, and the mechanism of contraction.

Despite widespread controversy still surrounding the phenomenon, stepwise shortening has now been confirmed by five independent methods in this laboratory, and by several other methods in different laboratories. In this paper we offer preliminary evidence obtained with the most recent method--measurement of 'isotonic muscle length transients'. We find that the muscle length inflections observed after quick release to an isotonic load correspond to pauses and steps at the sarcomere level. Thus, pauses and steps are reflected not only in sarcomere length and segment length signals, but in the muscle length signal as well. We review several of the more illuminating features of stepwise shortening, as well as new ultrastructural observations which, taken together, point to an hypothesis for the generation of steps. The steps may be generated by shortening of one or another of the sarcomere's filaments: connecting filaments in the unactivated myofibril and thick filaments in the activated myofibril. Supporting evidence is considered.

Animals↗
PubMed1987

Stepwise shortening of muscle fibre segments.

Shortening dynamics were measured in single fibres of frog skeletal muscle using a system that could track the spacing between hairs mounted on the fibre surface. Segment length changes were predominantly stepwise. The objective of the study was to identify potential artifacts and check their relevance. Several possible causes of artifactual steps were evaluated quantitatively and ruled out. In addition, the surface marker method and an independent length-detection method based on light diffraction were used simultaneously. The concurrence of results confirmed that it is highly unlikely that stepwise shortening could arise out of instrument artifact. Possible mechanisms underlying the phenomenon are considered.

Animals↗
PubMed1986

Bridgelike interconnections between thick filaments in stretched skeletal muscle fibers observed by the freeze-fracture method.

The ultrastructure of frog semitendinosus muscle was explored using the freeze-fracture, deep-etch, rotary-shadowing technique. Mechanically skinned fibers were stretched to decrease or eliminate the overlap of thick and thin filaments before rapid freezing with liquid propane. In relaxed, contracting, and rigor fibers, a significant number of bridgelike interconnections, distinct from those observed in the M-region, were observed between adjacent thick filaments in the non-overlap region. Their half-length and diameter corresponded approximately to the known dimensions of the cross-bridge (or myosin S-1). The interconnection may thus be formed by the binding of two apposed cross-bridges projecting from adjacent thick filaments. Fixation with 0.5% glutaraldehyde for 5-10 min before freezing effectively preserved these structures. The results indicate that the interconnections are genuine structures that appear commonly in stretched muscle fibers. They may play a role in stabilizing the thick filament lattice, and possibly in the contractile process.

Actin Cytoskeleton↗