Effect of inorganic phosphate on the force and number of myosin cross-bridges during the isometric contraction of permeabilized muscle fibers from rabbit psoas

The relation between the chemical and mechanical steps of the myosin-actin ATPase reaction that leads to generation of isometric force in fast skeletal muscle was investigated in demembranated fibers of rabbit psoas muscle by determining the effect of the concentration of inorganic phosphate (Pi) on the stiffness of the half-sarcomere (hs) during transient and steady-state conditions of the isometric contraction (temperature 12°C, sarcomere length 2.5 μm). Changes in the hs strain were measured by imposing length steps or small 4 kHz oscillations on the fibers in control solution (without added Pi) and in solution with 3-20 mM added Pi. At the plateau of the isometric contraction in control solution, the hs stiffness is 22.8 ± 1.1 kPa nm⁻¹. Taking the filament compliance into account, the total stiffness of the array of myosin cross-bridges in the hs (e) is 40.7 ± 3.7 kPa nm⁻¹. An increase in [Pi] decreases the stiffness of the cross-bridge array in proportion to the isometric force, indicating that the force of the cross-bridge remains constant independently of [Pi]. The rate constant of isometric force development after a period of unloaded shortening (r_F) is 23.5 ± 1.0 s⁻¹ in control solution and increases monotonically with [Pi], attaining a maximum value of 48.6 ± 0.9 s⁻¹ at 20 mM [Pi], in agreement with the idea that Pi release is a relatively fast step after force generation by the myosin cross-bridge. During isometric force development at any [Pi], e and thus the number of attached cross-bridges increase in proportion to the force, indicating that, independently of the speed of the process that leads to myosin attachment to actin, there is no significant (>1 ms) delay between generation of stiffness and generation of force by the cross-bridges.     

Effects of contraction duration on low-frequency fatigue in voluntary and electrically induced exercise of quadriceps muscle in humans

The aims of this study were to investigate if low-frequency fatigue (LFF) dependent on the duration of repeated muscle contractions and to compare LFF in voluntary and electrically induced exercise. Male subjects performed three 9-min periods of repeated isometric knee extensions at 40% maximal voluntary contraction with contraction plus relaxation periods of 30 plus 60 s, 15 plus 30 s and 5 plus 10 s in protocols 1, 2 and 3, respectively. The same exercise protocols were repeated using feedback-controlled electrical stimulation at 40% maximal tetanic torque. Before and 15 min after each exercise period, knee extension torque at 1, 7, 10, 15, 20, 50 and 100 Hz was assessed. During voluntary exercise, electromyogram root mean square (EMG_rms) of the vastus lateralis muscle was evaluated. The 20-Hz torque:100-Hz torque (20:100 Hz torque) ratio was reduced more after electrically induced than after voluntary exercise (P < 0.05). During electrically induced exercise, the decrease in 20:100 Hz torque ratio was gradually (P < 0.05) reduced as the individual contractions shortened. During voluntary exercise, the decrease in 20:100 Hz torque ratio and the increase in EMG_rms were greater in protocol 1 (P < 0.01) than in protocols 2 and 3, which did not differ from each other. In conclusion, our results showed that LFF is dependent on the duration of individual muscle contractions during repetitive isometric exercise and that the electrically induced exercise produced a more pronounced LFF compared to voluntary exercise of submaximal intensity. It is suggested that compensatory recruitment of faster-contracting motor units is an additional factor affecting the severity of LFF during voluntary exercise. Accepted: 5 November 1997     

The Mechanical Properties of Drosophila Jump Muscle Expressing Wild-Type and Embryonic Myosin Isoforms

Transgenic Drosophila are highly useful for structure-function studies of muscle proteins. However, our ability to mechanically analyze transgenically expressed mutant proteins in Drosophila muscles has been limited to the skinned indirect flight muscle preparation. We have developed a new muscle preparation using the Drosophila tergal depressor of the trochanter (TDT or jump) muscle that increases our experimental repertoire to include maximum shortening velocity (V_slack), force-velocity curves and steady-state power generation; experiments not possible using indirect flight muscle fibers. When transgenically expressing its wild-type myosin isoform (Tr-WT) the TDT is equivalent to a very fast vertebrate muscle. TDT has a V_slack equal to 6.1 ± 0.3 ML/s at 15°C, a steep tension-pCa curve, isometric tension of 37 ± 3 mN/mm², and maximum power production at 26% of isometric tension. Transgenically expressing an embryonic myosin isoform in the TDT muscle increased isometric tension 1.4-fold, but decreased V_slack 50% resulting in no significant difference in maximum power production compared to Tr-WT. Drosophila expressing embryonic myosin jumped <50% as far as Tr-WT that, along with comparisons to frog jump muscle studies, suggests fast muscle shortening velocity is relatively more important than high tension generation for Drosophila jumping.     

Magnitude of sarcomere extension correlates with initial sarcomere length during lengthening of activated single fibers from soleus muscle of rats

A laser-diffraction technique was developed that rapidly reports the lengths of sarcomeres (L_s) in serially connected sectors of permeabilized single fibers. The apparatus translates a laser beam along the entire length of a fiber segment within 2 ms, with brief stops at each of 20 contiguous sectors. We tested the hypothesis that during lengthening contractions, when maximally activated fibers are stretched, sectors that contain the longer sarcomeres undergo greater increases in L_s than those containing shorter sarcomeres. Fibers (n = 16) were obtained from the soleus muscles of adult male rats and the middle portions (length = 1.05 ± 0.11 mm; mean ± SD) were investigated. Single stretches of strain 27% and a strain rate of 54% s⁻¹ were initiated at maximum isometric stress and resulted in a 19 ± 9% loss in isometric stress. The data on L_s revealed that 1), the stretch was not distributed uniformly among the sectors, and 2), during the stretch, sectors at long L_s before the stretch elongated more than those at short lengths. The findings support the hypothesis that during stretches of maximally activated skeletal muscles, sarcomeres at longer lengths are more susceptible to damage by excessive strain.     

The isometric force that induces maximal surface muscle deoxygenation

To determine the external force that induces maximal deoxygenation of brachioradialis muscle 32 trained male subjects maintained isometric contractions using the elbow flexor muscles up to the limit time (isotonic part of the isometric contraction, IIC) and beyond that time for 120 s (anisotonic part of the isometric contraction). During IIC each subject maintained relative forces of either 25% and 70% maximal voluntary contraction (MVC), 50% and 100% MVC, or 40% and 60% MVC. Muscle oxygenation was assessed using a near infrared spectroscope, and expressed as a percentage of the reference value (ΔO_2rest) which was the difference between the minimal oxygenation obtained after 6 min of ischaemia at rest and the maximal reoxygenation following the release of the tourniquet. During IIC at 25% MVC, muscle oxygenation decreased to 17 (SEM 3)% ΔO_2rest, then it levelled off [25 (SEM 1)% ΔO_2rest]. After the point at which target force could not be maintained, reoxygenation was very weak. During IIC at 40%, 50%, 60%, and 70% MVC, the lowest muscle oxygenation values were obtained after 15–20 s of contraction and corresponded to −18 (SEM 6), −59 (SEM 12) −31 (SEM 6), and −29 (SEM 6)% ΔO_2rest, respectively. For the contraction at 100% MVC, the lowest oxygenation [−19 (SEM 9)% ΔO_2rest] was obtained while force was decreasing (69% MVC). During the anisotonic part of the isometric contractions, the greatest reoxygenation rate was obtained after 50% MVC IIC (P < 0.001). Our results showed that during isometric elbow flexions between 25% and 100% MVC, there was no linear relationship between external force and muscle oxygenation, and that the maximal deoxygenation of the brachioradialis muscle was obtained at 50% MVC. Accepted: 16 February 1998     

Cardiovascular responses to sustained maximal isometric contractions of the finger flexors

This study investigated cardiovascular responses to 2 min sustained submaximal (20% MVC) and maximal (100% MVC) voluntary isometric contractions of the finger flexors in healthy young women. Cardiovascular variables investigated were: heart rate (f _c), mean arterial pressure ( _a), and stroke volume (SV). Doppler echocardiography was used to estimate SV from measures of aortic diameter (AD) and time-velocity integrals. Preliminary studies indicated that AD did not change significantly after 2 min sustained 100% MVC. Therefore, pre-exercise AD values were used to calculate SV before, during and after exercise. During the 2-min 100% MVC period, f _c and _aincreased significantly during the first 30 s of contraction. f _c then remained constant during the remainder of the 2-min contraction period, while _acontinued to rise. SV did not change significantly during the 100% MVC task but increased significantly during recovery from sustained 100% MVC. The data suggest that the magnitude of cardiovascular responses to isometric exercise is dependent on the specific task performed, and that there is a different pattern of response for f _c, _a, and SV during 20% and 100% MVC tasks. Unlike f _c and _a, SV did not change significantly during isometric exercise, but increased significantly after sustained 100% MVC.     

Initial reaction kinetics of succinate dehydrogenase in mouse liver studied with a real-time image analyser system

Summary The initial reaction kinetics of succinate dehydrogenase in situ were investigated in sections of mouse unfixed liver using an ARGUS-100 image analyser system. The sections were incubated on substrate-containing agarose gel films. Images of a section, illuminated with monochromatic light (584 nm), were captured with the image analyser in real time at intervals of 10 s during the incubation. The absorbances of selected hepatocytes in the successive images were determined as a function of time. In every cell, the absorbance increased non-linearly after the first minute of incubation. The initial velocity of the dehydrogenase was calculated from the linear activities during the first 20 s of incubation. Hanes plots of the initial velocities and succinate concentration yielded the following mean kinetic constants. For periportal hepatocytes, the apparentK _m=1.2±0.8 mM andV _max=29±2 mol hydrogen equivalents formed/cm³ hepatocyte cytoplasm per min. For pericentral hepatocytes,K _m=1.4±1.0 mM andV _max=21±2 mol hydrogen equivalents/cm³ per min. TheK _m values are very similar to those determined previously from biochemical assays. These results, and the observed dependence of the initial velocity on the enzyme concentration, suggest that the technique reported here is valid for the histochemical assay of succinate dehydrogenase.     

The effect of exercise induced hyperthermia on muscle fibre conduction velocity during sustained isometric contraction

This study investigated the effect of dynamic exercise in a hot environment on muscle fibre conduction velocity (MFCV) of the knee extensors during a sustained isometric contraction. Seven trained male cyclists (mean [±SD], age, and were 35 ± 9.9 and 57.4 ± 6.6 ml kg⁻¹ min⁻¹) cycled for 50 min at 60% of peak power output in either: (1) 40 °C (HOT); or (2) 19 °C (NEUTRO); and (3) remained passive in 40 °C (PASS). Post-intervention a 100 s maximal sustained isometric contraction (SMC) of the knee extensors was performed. Rectal temperature increased (p < 0.01) for both HOT and NEUTRO with PASS unchanged and with HOT rising higher (p < 0.01) than NEUTRO (38.6 ± 0.4 vs. 37.6 ± 0.4 °C). Muscle temperature increased (p < 0.01) for all three conditions with HOT rising the highest (p < 0.01) (40.3 ± 0.5 vs. 38.3 ± 0.3 and 37.6 ± 1.3 °C for NEUTRO and PASS, respectively). Lactate showed higher accumulation (p < 0.01) for HOT than NEUTRO (6.9 ± 2.3 vs. 4.2 ± 2.1 mmol l⁻¹). During SMC the torque, electromyography root mean squared (RMS) and MFCV all significantly (p < 0.01) declined. Only in HOT did MFCV decline significantly (p < 0.01) less than torque and RMS (9.9 ± 6.2% vs. 37.5 ± 17.8% and 37.6 ± 21.4%, respectively). In conclusion, during exercise induced hyperthermia, reduced motor unit recruitment as opposed to slower conducting properties of the muscle fibre appears to be responsible for the greater reduction in torque output.     

Energy balance studies in frog skeletal muscles shortening at one-half maximal velocity

High-energy phosphate metabolism and energy liberated as heat and work were measured in 3-s tetani of frog sartorius muscle at 0 degree C. Two contraction periods were studied: (a) a 0.35-s period of shortening near half-maximum velocity beginning after 2 s of isometric stimulation, and (b) a 0.65-s isometric period immediately following the shortening. There were no significant changes in levels of ATP, ADP, or AMP in the two contraction periods. The observed changes in inorganic phosphate and creatine levels indicated that the only significant reaction occurring was phosphocreatine splitting. The mean rate of high-energy phosphate splitting during the shortening, 1.60 +/- 0.23 mumol X g-1 X s-1 (n = 24), was about fivefold higher than that in the 1-s period in the isometric tetanus, 0.32 +/- 0.11 mumol X g-1 X s- 1 (n = 17), observed in our previous study. The mean rate in the post- shortening period, 0.46 +/- 0.13 mumol X g-1 X s-1 (n = 17), was not significantly different from that in the 1-s period in the isometric tetanus. A large amount of heat plus work was produced during the shortening period, and this could be accounted for by simultaneous chemical changes. In the post-shortening period, the observed enthalpy was also accounted for by simultaneous chemical reactions. Thus, the present result is in sharp contrast to that obtained from a similar study performed at a shortening at Vmax, where an enthalpy excess was produced during shortening and an enthalpy deficit was produced during the period following the shortening.     

Sodium citrate ingestion and muscle performance in acute hypobaric hypoxia

Eight subjects were studied on four occasions following ingestion of a 300-ml solution containing either sodium citrate (C, 0.4g · kg^–1 body mass) or placebo (P, sodium chloride 0.045 g · kg^–1 body mass), at local barometric pressure (N, P _B approximately 740 mmHg, 98.7 kPa) or hypobaric hypoxia (HH, P _B = 463 mmHg, 61.7 kPa). At 2 h after ingestion of the solution, the subjects performed prolonged isometric knee-extension at 35% of the maximal voluntary contraction (MVC) measured either in N or HH. Results showed that ingestion of C led to an improvement in muscle endurance (P < 0.01). However, this increase in endurance time for knee extensor muscles was only significant in N ( +22%, P < 0.05, compared to + 15%, NS, at N and HH, respectively). Following ingestion of sodium citrate, pre-exercise bicarbonate concentrations and pH levels were significantly higher than those measured after P ingestion. A significant treatment effect was observed for blood lactate concentrations with values higher for C than for P after 4, 6 and 10 min of recovery (P < 0.05). Electromyographic signals (EMG) were obtained from the vastus lateralis muscle during the prolonged isometric contraction at 35% MVC. The mean power frequency (MPF) significantly decreased in time under both N-P and N-C conditions. In HH, no significant decrease in MPF was observed with time. The results suggest that C ingestion was an ergogenic aid enhancing endurance during a sustained isometric contraction. In addition, it is suggested that fatigue during prolonged isometric contraction in HH was not directly related to factors determining the EMG signs of fatigue.     

Cardiodepressive effect elicited by the essential oil of Alpinia speciosa is related to L-type Ca current blockade

This study was undertaken to elucidate the effect of the essential oil from Alpinia speciosa (EOAs) on cardiac contractility and the underlying mechanisms. The essential oil was obtained from Alpinia speciosa leaves and flowers and the oil was analyzed by GC-MS method. Chemical analysis revealed the presence of at least 18 components. Terpinen-4-ol and 1,8-cineole corresponded to 38% and 18% of the crude oil, respectively. The experiments were conducted on spontaneously-beating right atria and on electrically stimulated left atria isolated from adult rats. The effect of EOAs on the isometric contractions and cardiac frequency in vitro was examined. EOAs decreased rat left atrial force of contraction with an EC₅₀ of 292.2 ± 75.7 μg/ml. Nifedipine, a well known L-type Ca²⁺ blocker, inhibited in a concentration-dependent manner left atrial force of contraction with an EC₅₀ of 12.1 ± 3.5 μg/ml. Sinus rhythm was diminished by EOAs with an EC₅₀ of 595.4 ± 56.2 μg/ml. Whole-cell L-type Ca²⁺ currents were recorded by using the patch-clamp technique. EOAs at 25 μg/ml decreased I_Ca,L by 32.6 ± 9.2% and at 250 μg/ml it decreased by 89.3 ± 7.4%. Thus, inhibition of L-type Ca²⁺ channels is involved in the cardiodepressive effect elicited by the essential oil of Alpinia speciosa in rat heart.     

Effects of aerobic exercise on the torque-velocity relationship in cycling

The kinetics of the torque-velocity (T-ω) relationship after aerobic exercise was studied to assess the effect of fatigue on the contractile properties of muscle. A group of 13 subjects exercised until fatigued on a cycle ergometer, at an intensity which corresponded to 60% of their maximal aerobic power for 50 min (MAP60%); ten subjects exercised until fatigued at 80% of their maximal aerobic power for 15 min (MAP80%). Of the subjects 7 exercised at both intensities with at least a 1-week interval between sessions. Pedalling rate was set at 60 rpm. The T-ω relationship was determined from the velocity data collected during all-out sprints against a 19 N · m braking torque on the same ergometer, according to a method proposed previously. Maximal theoretical velocity (ω₀) and maximal theoretical torque (T ₀) were estimated by extrapolation of the linear T-ω relationship. Maximal power (P _max) was calculated from the values of T ₀ and ω₀ (P _max = 0.25 ω₀ T ₀). The T-ω relationships were determined before, immediately after and 5 and 10 min after the aerobic exercise. The kinetics of ω₀, T ₀ and P _max was assumed to express the effects of fatigue on the muscle contractile properties (maximal shortening velocity, maximal muscle strength and maximal power). Immediately after exercise at MAP60% a 7.8% decrease in T ₀ and 8.8% decrease in P _max was seen while the decrease in ω₀ was nonsignificant, which suggested that P _max decreased in the main because of a loss in maximal muscle strength. In contrast, MAP80% induced a 8.1% decrease in ω₀ and 12.8% decrease in P _max while the decrease in T ₀ was nonsignificant, which suggested that the main cause of the decrease in P _max was probably a slowing of maximal shortening velocity. The short recovery time of the T-ω relationship suggests that the causes of the decrease of torque and velocity are processes which recover rapidly. Accepted: 25 November 1996     

Temperature adaptation and the contractile properties of live muscle fibres from teleost fish

Summary The contractile properties of swimming muscles have been investigated in marine teleosts from Antarctic (Trematomus lepidorhinus, Pseudochaenichthys georgianus), temperate (Pollachius virens, Limanda limanda, Agonis cataphractus, Callionymus lyra), and tropical (Abudefduf abdominalis, Thalassoma duperreyi) latitudes. Small bundles of fast twitch fibres were isolated from anterior myotomes and/or the pectoral fin adductor profundis muscle (m. add. p). Live fibre preparations were viable for several days at in vivo temperatures, but became progressively inexcitable at higher or lower temperatures. The stimulation frequency required to produce fused isometric tetani increased from 50 Hz in Antarctic species at 0°C to around 400 Hz in tropical species at 25°C. Maximum isometric tension (P_o) was produced at the normal body temperature (NBT) of each species (Antarctic, 0–2°C; North Sea and Atlantic, 8–10°C; Indo-West Pacific, 23–25°C). P₀ values at physiological temperatures (200–300 kN·m^–2) were similar for Antarctic, temperate, and tropical species. A temperature induced tension hysteresis was observed in muscle fibres from some species. Exposure to <0°C in Antarctic and <2°C in temperate fish resulted in the temporary depression of tension over the whole experimental range, an effect reversed by incubation at higher temperatures. At normal body temperatures the half-times for activation and relaxation of twitch and tetanic tension increased in the order Antarctic>temperate>tropical species. Relaxation was generally much slower at temperatures <10°C in fibres from tropical than temperate fish. Q₁₀ values for these parameters at NBTs were 1.3 2.1 for tropical species, 1.7–2.6 for temperate species, and 1.6–3.5 for Antarctic species. The forcevelocity (P-V) relationship was studied in selected species using iso-velocity releases and the data below 0.8 P₀ iteratively fitted to Hill's equation. The P-V relation at NBT was found to be significantly less curved in Antarctic than temperate species. The unloaded contraction velocity (V_max) of fibres was positively correlated with NBT increasing from about 1 muscle fibre length·s^–;1 in an Antarctic fish (Trematomus lepidorhinus) at 1°C to around 16 muscle fibre lengths·s^–1 in a tropical species (Thalassoma duperreyi) at 24°C. It is concluded that although muscle contraction in Antarctic fish shows adaptations for low temperature function, the degree of compensation achieved in shortening speed and twitch kinetics is relatively modest.Abbreviations ET environmental temperature - m. add. p major adductor profundis - m. add. s. major adductor superficialis - NBT normal body temperature - P ₀ maximum isometric tension - P-V force velocity - SR sarcoplasmic reticulum - T 1/2 a half activation time - T 1/2 r half relaxation time - V _max unloaded contraction     

Expression of Green Fluorescent Protein Impairs the Force-Generating Ability of Isolated Rat Ventricular Cardiomyocytes

Green fluorescent protein (GFP) is widely used as a biologically inert expression marker for studying the effects of transgene expression in heart tissue, but its influence on the contractile function of cardiomyocytes has not yet been fully evaluated. We measured the contractile function of isolated rat ventricular myocytes before and after infection with a recombinant adenovirus expressing GFP (Adv-GFP). Myocytes infected with a non-transgene-containing adenovirus (Adv-Null) or uninfected myocytes (UI) served as controls. Using a carbon-fiber-based force-length measurement system for single cardiomyocytes, we evaluated the contractile function over a wide range of loading conditions including the shortening fraction (%FS) and maximal shortening velocity (Vmax) under the unloaded condition, and isometric force. At 24 hours after infection, nearly 80% of the Adv-GFP-infected myocytes expressed GFP. We found that the %FS and Vmax did not differ among the three groups, however, the isometric force showed a mild, but significant, decrease only in Adv-GFP myocytes (Adv-GFP: 29.1 ± 4.0 mN/mm²; Adv-Null: 42.8 ± 6.2 mN/mm²; UI: 47.1 ± 4.8 mN/mm²; p = 0.03). An evaluation of the contractile function of isolated cardiomyocytes under high load conditions revealed impaired isometric contractility by GFP expression. Adv-GFP expression may not be an ideal control for specific gene expression experiments in myocardial tissue.     

Biological evidence for the direct stimulating effect of PGF2α on the release of pituitary luteolytic agent(s) of pseudopregnant rats

Stereotaxic implantation of PGF_2α in the thick portion of the posterior part of the anterior pituitary(AP) and in the vicinity of the median eminence(ME), during various interval of the active phase of pseudopregnancy were studied. AP implantation of the drug on L₀, L₄, L₇, and L₉ showed significant shortened normal duration of pseudopregnancy of animals without PGF_2α implantation from 13.4±0.4 days to 9.3±1.9, 11.6±0.8, 11.0±0.5 and 10.7±0.7 days respectively. The same implantation also capable of preventing the postponement of leucocytic vaginal smear of hysterectomized pseudopregnant animals. The overall means duration of pseudopregnancy of L₀-L₉ hysterectomized animals who had no PGF_2α implant were 18.7±2.3 days while overall means of PGF_2α implanted animals were only 12.9±0.9 days. ME implant of the drug failed to show clear cut statistically shortening of the duration of pseudopregnancy. The overall means of the duration of pseudopregnancy of these animals were 11.8±0.5 days. ME implant also failed to prevent postponement of the duration of leucocytic vaginal smear of hysterectomized pseudopregnant animals in all groups observed. These evidences favor the possibility of direct stimulating effect of PGF_2α on the release of pituitary luteolytic agent in rats, possibly LH in nature.     

The role of central command in ventilatory control during static exercise

The role of central command in the respiratory response to 15 min of rhythmic-static (isometric) exercise was studied in humans. Voluntary exercise (VE) was compared with electrically induced exercise (EE) at three different work intensities, i.e. 5%, 15% and 25% of maximal voluntary contraction. A group of 12 volunteers participated in the study and each of them performed six sessions. A session consisted of at least 5 min rest, 15 min rhythmic-static single leg exercise (4 s contraction/12 s relaxation) and at least 5 min recovery. Force, minute ventilation (V _E) and oxygen uptake (VO₂) were measured. In EE, both V _E and VO₂ increased continuously during the entire exercise period after an initial rapid increase at all three work intensities. Correlation between VE and VO₂ was highly significant during EE. During all three work intensities of VE, VE and VO₂ achieved a steady-state after the initial increase. During VE, VE did not correlate as closely with VO₂ as during EE. All these findings indicate that central command was not imperative for an adequate ventilatory response to exercise within all three work intensities investigated. Without the influence of central command, correlation between VE and VO₂ was even better than during VE.     

Force-velocity shifts with repetitive isometric and isotonic contractions of canine gastrocnemius in situ.

The force-velocity (F-V) relationships of canine gastrocnemius-plantaris muscles at optimal muscle length in situ were studied before and after 10 min of repetitive isometric or isotonic tetanic contractions induced by electrical stimulation of the sciatic nerve (200-ms trains, 50 impulses/s, 1 contraction/s). F-V relationships and maximal velocity of shortening (Vmax) were determined by curve fitting with the Hill equation. Mean Vmax before fatigue was 3.8 +/- 0.2 (SE) average fiber lengths/s; mean maximal isometric tension (Po) was 508 +/- 15 g/g. With a significant decrease of force development during isometric contractions (-27 +/- 4%, P < 0.01, n = 5), Vmax was unchanged. However, with repetitive isotonic contractions at a low load (P/Po = 0.25, n = 5), a significant decrease in Vmax was observed (-21 +/- 2%, P < 0.01), whereas Po was unchanged. Isotonic contractions at an intermediate load (P/Po = 0.5, n = 4) resulted in significant decreases in both Vmax (-26 +/- 6%, P < 0.05) and Po (-12 +/- 2%, P < 0.01). These results show that repeated contractions of canine skeletal muscle produce specific changes in the F-V relationship that are dependent on the type of contractions being performed and indicate that decreases in other contractile properties, such as velocity development and shortening, can occur independently of changes in isometric tension.     

Hydraulic Conductivity (Lp) and Its Activation Energy (Ea), Cryoprotectant Agent Permeability (Ps) and ItsEa, and Reflection Coefficients (?) for Golden Hamster Individual Pancreatic Islet Cell Membranes

Long-term cryopreservation of islets of Langerhans would be advantageous to a clinical islet transplantation program. Fundamental cryobiology utilizes knowledge of basic biophysical characteristics to increase the understanding of the preservation process and possibly increase survival rate. In this study several of these previously unreported characteristics have been determined for individual islet cells isolated from Golden hamster islets. Using an electronic particle counting device and a temperature control apparatus, dynamic volumetric response of individual islet cells to anisosmotic challenges of 1.5 M dimethyl sulfoxide (DMSO) and 1.5 M ethylene glycol (EG) were recorded at four temperatures (8, 22, 28, and 37°C). The resulting curves were fitted using Kedem and Katchalsky equations which describe water flux and cryoprotectant agent (CPA) flux based on hydraulic conductivity (L_p), CPA permeability (P_s), and reflection coefficient (?) for the membrane. For Golden hamster islet cells,L_p,P_s, and ? for DMSO at 22°C were found to be 0.23 ± 0.06 μm/min/atm, 0.79 ± 0.32 × 10⁻³cm/min, and 0.55 ± 0.37 (n= 11) (mean ± SD), respectively. For EG at 22°C,L_pequaled 0.23 ± 0.06 μm/min/atm,P_sequaled 0.63 ± 0.20 × 10⁻³cm/min, and ? was 0.75 ± 0.17 (n= 9). Arrhenius plots (lnL_por lnP_sversus 1/temperature (K)) were created by adding the data from the other three temperatures and the resulting linear regression yielded correlation coefficients (r) of 0.99 for all four plots (L_pandP_sfor both CPAs). Activation energies (E_a) ofL_pandP_swere calculated from the slopes of the regressions. The values for DMSO were found to be 12.43 and 18.34 kcal/mol forL_pandP_s(four temperatures, totaln= 52), respectively. For EG,E_aofL_pwas 11.69 kcal/mol andE_aofP_swas 20.35 kcal/mol (four temperatures, totaln= 58).     

LHRH-receptor-regulated Ca2+-ATPase activity in murine pituitary gland

Addition of luteinizing hormone releasing hormone (LHRH) in vitro (10^–5–5×10^–9 M) to murine pituitary membranes resulted in a dose-related decrease in Ca²⁺-ATPase activity within 15 min. Inhibitory effects of LHRH (10^–7 M) occurred after 90 sec, and appeared maximal by 120 sec. Eadie-Hofstee analysis at 10^–7 M LHRH, at varying [Ca²⁺]_free, resulted in aK _m=0.89±0.06 M and aV _max=18.8±0.71 nmol/mg per 2 min, compared to aK _m=0.69±0.06 M and aV _max=32.8±1.21 nmol/mg per 2 min for controls. Pre-incubation for 5 min with LHRH antagonist (10^–8 M) significantly attenuated (50%) the inhibitory effects of 10^–7 M LHRH on pituitary Ca²⁺ ATPase activity with aK _m=0.97±0.24 M and aV _max=28.1±2.8 nmol/mg per 2 min. The addition of LHRH (10^–7 M) to pituitary homogenates significantly increased luteinizing hormone (LH) release already at 10 and up to 40 sec compared to basal LH release. Systemic administration of 50 ng LHRH (i.p.), significantly (P<0.05) reduced pituitary Ca²⁺-ATPase after 30, 60 and 90 min, with a return to control levels by 120 min. Pituitary LH content was reduced slightly at 15 min, but was increased significantly at 90 and 120 min post-treatment. Plasma LH levels were elevated by 5 min, reached a peak by 15 min and returned to control within 60 min. The present findings indicate that LHRH receptor activation may influence cytosolic Ca²⁺ transport through effects on membrane Ca²⁺-ATPase activity. These actions may regulate LHRH-induced synthesis, storage and release of LH from pituitary gonadotropes.     

Release of fatty acid-binding protein and long chain fatty acids from isolated rat heart after ischemia and subsequent calcium paradox

To obtain insight into the relation between the release of heart-type fatty acid-binding protein (H-FABP_c) and of long-chain fatty acids (FA) from injured cardiac tissue, rat hearts were Langendorff perfused according to the following scheme: 30 min normoxia, 60 min ischemia, 30 min reperfusion, 10 min Ca²⁺ free perfusion and finally 10 min Ca²⁺ repletion. During this protocol right ventricular (Q _rv ) and interstitial effluent samples (Q _i ) were collected at regular intervals. During reperfusion a total of 0.8±0.1 nmol H-FABP_c but no FA were detected in the effluents. However, during Ca²⁺ readmission, 45±4 nmol H-FABP_c (80–90% of total tissue content) was released with an initial (first 3 min) simultaneous release of FA (FA/H-FABP_c ratio 0.90±0.07 mol/mol). Thereafter, FA release continued at 10–15 nmol per min mainly inQ _rv while the rate of H-FABP_c release decreased. During Ca²⁺ repletion, tissue FA content raised rapidly from 168±20 to 1918±107 nmol/g dry weight. These findings suggest that after severe cardiac damage initially FA is released bound to H-FABP_c, whereas further FA release occurs in a non-protein bound manner.