Force and myosin content variation in isolated intact single muscle fibres from Rana temporaria

We studied the relation between force normalized by dry mass per unit length and the myosin fraction of muscle dry mass. The two tibialis anterior muscles were dissected from 12 frogs (Rana temporaria). Then, from one muscle, two single fast-twitch fibres were isolated. Each fibre was mounted isometrically in Ringer's solution, and electrically stimulated using a standardized protocol. Peak force production, normalized by the fibre's dry mass per unit length, varied by a factor of 1.4. Little variation in normalized force was measured between fibres from the same animal, whereas between animals a significant difference was found (P<0.05). The contralateral muscle was used to determine the myosin fraction of the dry mass. The relationship between the fraction myosin of the dry mass and force normalized by dry mass per unit length showed a high correlation (r = 0.81; n = 12). From this we conclude that variation in normalized tetanic force is determined greatly (65%) by variations in myosin content.     

Optical trap stiffness in the presence and absence of spherical aberrations

Optical traps are commonly constructed with high-numerical-aperture objectives. Oil-immersion objectives suffer from spherical aberrations when used for imaging in aqueous solutions. The effect of spherical aberrations on trapping strength has been modeled by approximation, and only a few experimental results are available in the case of micrometer-sized particles. We present an experimental study of the dependence of lateral and axial optical-trap stiffness on focusing depth for polystyrene and silica beads of 2 microm diameter by using oil- and water-immersion objectives. We demonstrate a strong depth dependence of trap stiffness with the oil-immersion objective, whereas no depth dependence was observed with the water-immersion objective.     

Rate of phosphate release after photoliberation of adenosine 5'-triphosphate in slow and fast skeletal muscle fibers

Inorganic phosphate (Pi) release was determined by means of a fluorescent Pi-probe in single permeabilized rabbit soleus and psoas muscle fibers. Measurements of Pi release followed photoliberation of approximately 1.5 mM ATP by flash photolysis of NPE-caged ATP in the absence and presence of Ca2+ at 15 degrees C. In the absence of Ca2+, Pi release occurred with a slow rate of 11 +/- 3 microM . s-1 (n = 3) in soleus fibers and 23 +/- 1 microM . s-1 (n = 10) in psoas fibers. At saturating Ca2+ concentrations (pCa 4.5), photoliberation of ATP was followed by rapid force development. The initial rate of Pi release was 0.57 +/- 0.05 mM . s-1 in soleus (n = 13) and 4.7 +/- 0.2 mM . s-1 in psoas (n = 23), corresponding to a rate of Pi release per myosin head of 3.8 s-1 in soleus and 31.5 s-1 in psoas. Pi release declined at a rate of 0.48 s-1 in soleus and of 5.2 s-1 in psoas. Pi release in soleus was slightly faster in the presence of an ATP regenerating system but slower when 0.5 mM ADP was added. The reduction in the rate of Pi release results from an initial redistribution of cross-bridges over different states and a subsequent ADP-sensitive slowing of cross-bridge detachment.     

Influence of phosphate and pH on myofibrillar ATPase activity and force in skinned cardiac trabeculae from rat

1. The effects of inorganic phosphate (Pi) and pH on maximal calcium-activated isometric force and MgATPase activity were studied in chemically skinned cardiac trabeculae from rat. ATP hydrolysis was coupled enzymatically to the breakdown of NADH, and its concentration was determined photometrically. Measurements were performed at 2.1 microns sarcomere length and 20 degrees C. ATPase activity and force were also determined when square-wave-shaped length changes were applied, with a frequency of 23 Hz and an amplitude of 2.5%. 2. At pH 7.0 without added Pi, the average isometric force (+/- S.E.M.) was 51 +/- 3 kN m-2 (n = 23). The average isometric ATPase activity was 0.43 +/- 0.02 mM s-1 (n = 23). During the changes in length ATPase activity increased to 152 +/- 3% of the isometric value, while the average force level decreased to 48 +/- 2%. 3. Isometric force gradually decreased to 31 +/- 2% of the control value when the Pi concentration was increased to 30 mM. Isometric ATPase activity, however, remained constant for Pi concentrations up to 5 mM and decreased to 87 +/- 3% at 30 mM Pi. When Pi accumulation inside the preparation due to ATP hydrolysis was taken into account, a linear relationship was found between isometric force and log [Pi]. The decrease in relative force was found to be 44 +/- 4% per decade. 4. During the length changes, ATPase activity and average force showed, apart from the increase in ATPase activity and decrease in average force, the same dependence on Pi as the isometric values. Stiffness, estimated from the amplitude of the force responses during the length changes, decreased in proportion to isometric force when the Pi concentration was increased. The changes in the shape of the force responses due to the repetitive changes in length as a function of the Pi concentration were relatively small. These results suggest that the effect of Pi on the transitions which influence ATP turnover is rather insensitive to changes in cross-bridge strain. 5. Isometric force, normalized to the control value at pH 7.0, increased gradually from 54 +/- 1% at pH 6.2 to 143 +/- 10% at pH 7.5. ATPase activity remained practically constant for pH values from 6.8 to 7.2 but decreased to 80 +/- 1% at pH 6.2 and to 83 +/- 5% at pH 7.5. ATPase activity during the length changes was reduced more than the isometric ATPase activity when pH was lowered.(ABSTRACT TRUNCATED AT 400 WORDS)     

Calcium modulates the influence of length changes on the myofibrillar adenosine triphosphatase activity in rat skinned cardiac trabeculae

The relationship between adenosine triphosphate (ATP) turnover and muscle performance was investigated in skinned cardiac trabeculae of the rat at different [Ca2+] and two different sarcomere lengths (1.8 microns and 2.2 microns) at 20 degrees C. ATP turnover was measured photometrically by enzymatic coupling of the regeneration of ATP to the oxidation of reduced nicotinamide adenine dinucleotide. The trabeculae were studied under isometric conditions and when the length was altered repetitively at a frequency of 23 Hz, with a square wave, by 5% of the initial length. The isometric ATPase activity amounted to 0.48 mM/s. Isometric ATP turnover and force were proportional at different [Ca2+]. During length changes at maximal activation (pCa 4.27) and 2.2 microns sarcomere length, ATPase activity increased to up to 162% whereas at low [Ca2+], ATPase activity decreased with respect to the isometric value at that pCa. At pCa 5.5, ATPase activity was reduced to 33%. These results indicate that during the length changes the apparent cross-bridge detachment rate is increased and the apparent attachment rate is decreased. The findings suggest that the Fenn effect, i.e. the increase in energy turnover above the isometric value during shortening, is present in cardiac trabeculae at high levels of activation, but is absent or reversed at lower levels of activity.     

Magnetism of single-crystalline Fe nanostructures

The quantitative investigation of magnetic nanostructures by means of ferromagnetic resonance is demonstrated for single-crystalline iron nanostructures. It is shown that the single-crystalline nature leads to effects not being present in polycrystalline ones and helps to quantitatively interpret the results. First a method is presented that enables one to fabricate epitaxial Fe nanowires starting from a thin film of Fe grown under ultrahigh vacuum conditions on GaAs (110). The system allows, due to the combination of cubic and twofold magnetic anisotropy, to prepare wires whose easy axis in remanence is oriented perpendicular to the wires axis. This unique feature is only achievable in epitaxial systems. Furthermore, nearly perfect Fe nanocubes with 13.6 nm edge length prepared by wet-chemical methods are studied. While the shell of the particles is composed of either Fe3O4 or gamma-Fe2O3, the core consists of metallic Fe. Oxygen and hydrogen plasma are used to remove the ligand system and the oxide shell. The single-crystalline nature of the cubes enables one to quantitatively determine the magnetic properties of the individual particle by means of ferromagnetic resonance measurements on an ensemble together with a model based on the Landau-Lifshitz equation. The measurements reveal a magneto-crystalline anisotropy of K4 = 4.8. 10(4) J/m3 being equal to bulk value and a saturation magnetization which is reduced to M(5K) = (1.2 +/- 0.12). 10(6) A/m (70% of bulk value). The effective damping parameter alpha = 0.03 is increased by one order of magnitude with respect to bulk Fe, showing that magnetic damping in nanostructures differs from the bulk.     

Force production in mechanically isolated cardiac myocytes from human ventricular muscle tissue

OBJECTIVE: The expression of contractile isoforms changes during various pathological conditions but little is known about the consequences of these changes for the mechanical properties in human ventricular muscle. We investigated the feasibility of simultaneous determination of protein composition and isometric force development in single cardiac myocytes from human ventricular muscle tissue obtained from small biopsies taken during open heart surgery. METHODS: Small biopsies of about 3 mg wet weight were taken during open heart surgery from patients with aortic valve stenosis. These biopsies were divided in two parts. One part (approximately 2 mg) was used for mechanical isolation of single myocytes and subsequent force measurement while the remaining part was used, in aliquots of 1 microgram dry weight, for protein analysis by polyacrylamide gel electrophoresis. The myocytes were attached with silicon glue to a sensitive force transducer and a piezoelectric motor, mounted on an inverted microscope and permeabilized by means of Triton X-100. Force development was studied at various free calcium concentrations. RESULTS: From all biopsies, myocytes could be obtained and the composition of contractile proteins could be determined. The average isometric force (+/- s.e.m.) at saturating calcium concentration obtained on 20 myocytes from 5 patients amounted to 51 +/- 8 kN/m2. Force was half maximal at a calcium concentration of 2.47 +/- 0.10 microM. CONCLUSION: These measurements indicate that it is possible to study the correlation between mechanical properties and protein composition in small biopsies from human ventricular muscle.     

Calcium handling by the sarcoplasmic reticulum during oscillatory contractions of skinned skeletal muscle fibres

Isometric ATP consumption and force were investigated in mechanically skinned fibres from iliofibularis muscle of Xenopus laevis. Measurements were performed at different [Ca2+], in the presence and absence of caffeine (5 nM). In weakly Ca2+-buffered solutions without caffeine, spontaneous oscillations in force and ATPase activity occurred. The repetition frequency was [Ca2+]-and temperature-dependent. The Ca2+ threshold (+/- SEM) for the oscillations corresponded to a pCa of 6.5 +/- 0.1. The maximum ATP consumption associated with calcium uptake by the sarcoplasmic reticulum (SR) reached during the oscillations was similar to the activity under steady-state conditions at saturating calcium concentrations in the presence of caffeine. Maximum activity was reached when the force relaxation was almost complete. The calculated amount of Ca2+ taken up by the SR during a complete cycle corresponded to 5.4 +/ 0.4 mmol per litre cell volume. In strongly Ca2+-buffered solutions, caffeine enhanced the calcium sensitivity of the contractile apparatus and, at low calcium concentrations, SR Ca uptake. These results suggest that when the SR is heavily loaded by net Ca uptake, there is a massive calcium-induced calcium release. Subsequent net Ca uptake by the SR then gives rise to the periodic nature of the calcium transient.     

Visualization of spin dynamics in single nanosized magnetic elements

The design of future spintronic devices requires a quantitative understanding of the microscopic linear and nonlinear spin relaxation processes governing the magnetization reversal in nanometer-scale ferromagnetic systems. Ferromagnetic resonance is the method of choice for a quantitative analysis of relaxation rates, magnetic anisotropy and susceptibility in a single experiment. The approach offers the possibility of coherent control and manipulation of nanoscaled structures by microwave irradiation. Here, we analyze the different excitation modes in a single nanometer-sized ferromagnetic stripe. Measurements are performed using a microresonator set-up which offers a sensitivity to quantitatively analyze the dynamic and static magnetic properties of single nanomagnets with volumes of (100 nm)(3). Uniform as well as non-uniform volume modes of the spin wave excitation spectrum are identified and found to be in excellent agreement with the results of micromagnetic simulations which allow the visualization of the spatial distribution of these modes in the nanostructures.