Force Generation In Skeletal Muscle Research Articles

Sex differences in isolated skeletal muscle force production: a spectrum of responses

Whether sex differences in skeletal muscle microvascular responses differ in response to muscle contraction is currently unknown. To determine vascular responses to muscle contraction in vivo we use intravital microscopy using thin skeletal muscle models of mixed fibre types (i.e. hamster retractor muscle). However, to effectively quantify this relationship in vivo, the initial muscle contraction stimulus must be constant across the sexes and thus, we must first determine whether there are sex differences in skeletal muscle force generation. We hypothesized that sexual dimorphisms in force production would not be apparent in muscles of any fibre type composition. To test this hypothesis, we used skeletal muscles with unique fibre type characteristics and oxidative capacities. Mouse soleus (SOL: primarily type 1 >75%), extensor digitorum longus (EDL: primarily type IIB ~60%), and costal diaphragm (DIA: primarily type IIA ~55%) muscles were isolated from adult male and female CD-1 mice. The retractor muscle (RET: primarily type IIX ~58%) was also isolated from adult male and female Syrian hamsters. Muscles were electrically stimulated in vitro and force production was quantified at optimal length during a force-frequency test (1-120Hz) and twitch (1Hz) contractions. In mice, females generated greater force during the force-frequency test in SOL (10-120Hz) and EDL (30-120Hz) and generated greater twitch forces compared to males (SOL: female 34.7 ± 2.4 mN/mm 2 , n=21 vs male 27.8 ± 2.0 mN/mm 2 , n=17; EDL: female 41.2 ± 2.2 mN/mm 2 , n=35 vs male 32.5 ± 2.1 mN/mm 2 , n=29). Females generated greater force during the force frequency test in DIA at 30Hz only and no significant sex differences were observed in DIA twitch force (female 0.24 ± 0.02 g/mg, n=19 vs male 0.20 ± 0.02 g/mg, n=21). In hamster RET no significant sex differences in force were observed during the force frequency test or during the twitch contraction (female 0.13 ± 0.01 g/mg, n=19 vs male 0.14 ± 0.02 g/mg, n=20). Collectively, these findings demonstrate that sex differences in force production were observed in muscles primarily comprised of slow oxidative (type 1) and fast glycolytic (type IIB) fibres but not in muscles primarily comprised of intermediate fibres (type IIA and type IIX). These data show that on a spectrum of muscle fibres from slow oxidative to fast glycolytic, sex differences are most apparent on the extremes of the spectrum, whereas sexual dimorphisms in the middle of the spectrum are not apparent. This work is supported by NSERC. NSERC This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Characterization of Motor Unit Firing and Twitch Properties for Decoding Musculoskeletal Force in the Human Ankle Joint In Vivo.

Understanding how motor units (MUs) contribute to skeletal mechanical force is crucial for unraveling the underlying mechanism of human movement. Alterations in MU firing, contractile and force-generating properties emerge in response to physical training, ageing or injury. However, how changes in MU firing and twitch properties dictate skeletal muscle force generation in healthy and impaired individuals remains an open question. In this work, we present a MU-specific approach to identify firing and twitch properties of MU samples and employ them to decode musculoskeletal function in vivo. First, MU firing events were decomposed offline from high-density electromyography (EMG) of six lower leg muscles involved in ankle plantar-dorsi flexion.We characterized their twitch responses based on the statistical distributions of their firing properties and employed them to compute MU-specific activation dynamics. Subsequently, we decoded ankle joint moments by linking our framework to a subject-specific musculoskeletal model. We validated our approach at different ankle positions and levels of activation and compared it with traditional EMG-driven models. Our proposed MU-specific formulation achieves higher generalization across conditions than the EMG-driven models, with significantly lower coefficients of variation in torque predictions. Furthermore, our approach shows distinct neural strategies across a large repertoire of contractile conditions in different muscles. Our proposed approach may open new avenues for characterizing the relationship between MU firing and twitch properties and their influence on force capacity. This can facilitate the development of targeted rehabilitation strategies tailored to individuals with specific neuromuscular conditions.

An Electromyography-Based Constitutive Law for Force Generation in Skeletal Muscle-Part I: Model Development.

This paper proposes a new method for estimating skeletal muscle forces using a model derived from dimensional analysis. It incorporates electromyography signals and muscle force-length, force-velocity, and force-frequency relationships as inputs. The purpose of this model is to provide more accurate estimates of individualized muscle forces to better predict surrounding musculoskeletal tissue and joint contact loading. The derivation begins with dimensional analysis and a selection of critical parameters that define muscle force generation. The resulting constitutive equation gives way to a unique application of inverse-dynamics, one which avoids the issue of indeterminacy when reaction moments and ligament loading are minimized in a joint. The ankle joint is used as an example for developing the equations that culminate into a system of linear equations. A muscle force model capable of being calibrated and then used to predict joint contact and surrounding tissue loading is critical in advancing biomechanics research areas like injury prevention, performance optimization, and tissue engineering, among others. This model's foundation in dimensional analysis, along with its inclusion of electromyography signals, gives promise that it will be physiologically relevant and suitable for application-based studies. A following paper, Part II, will evaluate this premise in an experimental setting.

Activation Of Specific Estrogen Receptor Isotype Mediates Skeletal Muscle Force Potentiation In Ovariectomized Mice

Estrogens affect force generation of skeletal muscle. Post-tetanic potentiation (PTP) of force is low in ovariectomize mice, and estradiol supplementation rescues potentiation both in vivo and in vitro. However, it is not known whether this estrogenic influence is through estrogen receptors, and if so, which isotype. PURPOSE: We hypothesized activation of estrogen receptor beta (ERβ) or G protein-coupled estrogen receptor (GPER), but not estrogen receptor alpha (ERα) would enhance in vivo muscle PTP in ovariectomized mice. METHODS: Adult female C57BL/6J mice (n = 4-12 per treatment) had a nerve cuff surgically implanted on the left common peroneal nerve. Six weeks later mice were ovariectomized (OVX). Four weeks later, PTP of the anterior crural muscles was measured immediately before and 1 h after treatment with either vehicle (OVX+Veh), 17β-estradiol (OVX+E2), ERα agonist PPT (OVX+PPT; 1 μM PPT), ERβ agonist DPN (OVX+DPN; 50 nM DPN) or GPER agonist G1 (OVX+G1, 2.4 nM G1) via tail vein injection. PTP was calculated as the percent increase in twitch torque from baseline to the highest torque of the post-tetanic twitches. One-way ANOVAs with Holm-Sidak post hoc tests were used for data analysis. Results are reported as mean ± SE. RESULTS: Peak torques of unpotentiated, baseline twitches were not different among the five groups: 0.47 ± 0.03, 0.59 ± 0.04, 0.62 ± 0.12, 0.52 ± 0.11 and 0.54 ± 0.03 mN*m, respectively (p=0.25). Potentiated twitches generated 7-108% more torque than unpotentiated twitches. PTP of OVX+E2 and OVX+G1 was significantly greater than that of OVX+Veh mice (59 ± 5% and 61 ± 7% vs 35 ± 5% increase, respectively; p=0.01). PTP of OVX+PPT and OVX+DPN were not significantly different from OVX+Veh mice (41 ± 4% and 49 ± 7% vs 35 ± 5% increase, respectively; p=0.37). CONCLUSION: Acute treatment with ERα or ERβ agonist does not increase in vivo PTP of the anterior crural muscles in OVX mice to the extent that E2 or GPER agonist does. This result partially supports our hypothesis that estrogenic modulation of skeletal muscle force potentiation is not through ERα, and is through GPER. Supported by NIH grants R01-AG031743 and T32-AR050938, and Metropolitan State University Professional Development Grant.

Stress-Dependent Activation of Myosin Motors Controls the Cooperativity and Dynamics of Force Generation in Skeletal Muscle

Stress-Dependent Activation of Myosin Motors Controls the Cooperativity and Dynamics of Force Generation in Skeletal Muscle

Microstructural analysis of skeletal muscle force generation during aging.

Human aging results in a progressive decline in the active force generation capability of skeletal muscle. While many factors related to the changes of morphological and structural properties in muscle fibers and the extracellular matrix (ECM) have been considered as possible reasons for causing age-related force reduction, it is still not fully understood why the decrease in force generation under eccentric contraction (lengthening) is much less than that under concentric contraction (shortening). Biomechanically, it was observed that connective tissues (endomysium) stiffen as ages, and the volume ratio of connective tissues exhibits an age-related increase. However, limited skeletal muscle models take into account the microstructural characteristics as well as the volume fraction of tissue material. This study aims to provide a numerical investigation in which the muscle fibers and the ECM are explicitly represented to allow quantitative assessment of the age-related force reduction mechanism. To this end, a fiber-level honeycomb-like microstructure is constructed and modeled by a pixel-based Reproducing Kernel Particle Method (RKPM), which allows modeling of smooth transition in biomaterial properties across material interfaces. The numerical investigation reveals that the increased stiffness of the passive materials of muscle tissue reduces the force generation capability under concentric contraction while maintains the force generation capability under eccentric contraction. The proposed RKPM microscopic model provides effective means for the cellular-scale numerical investigation of skeletal muscle physiology. NOVELTY STATEMENT: A cellular-scale honeycomb-like microstructural muscle model constructed from a histological cross-sectional image of muscle is employed to study the causal relations between age-associated microstructural changes and age-related force loss using Reproducing Kernel Particle Method (RKPM). The employed RKPM offers an effective means for modeling biological materials based on pixel points in the medical images and allow modeling of smooth transition in the material properties across interfaces. The proposed microstructure-informed muscle model enables quantitative evaluation on how cellular-scale compositions contribute to muscle functionality and explain differences in age-related force changes during concentric, isometric and eccentric contractions.

Assessing the role of Ca[formula omitted] in skeletal muscle fatigue using a multi-scale continuum model

The Calcium ion Ca2+ plays a critical role as an initiator and preserving agent of the cross-bridge cycle in the force generation of skeletal muscle. A new multi-scale chemo-mechanical model is presented in order to analyze the role of Ca2+ in muscle fatigue and to predict fatigue behavior. To this end, a cross-bridge kinematic model was incorporated in a continuum based mechanical model, considering a thermodynamic compatible framework. The contractile velocity and the generated active force were directly related to the force-bearing states that were considered for the cross-bridge cycle. In order to determine the values of the model parameters, the output results of an isometric simulation were initially fitted with experimental data obtained for rabbit Extensor Digitorum Longus muscle. Furthermore, a simulated force-velocity curve under concentric contractions was compared with reported experimental results. Finally, by varying the Ca2+ concentration level and its kinetics in the tissue, the model was able to predict the evolution of the active force of an experimental fatigue protocol. The good agreement observed between the simulated results and the experimental outcomes proves the ability of the model to reproduce the fatigue behavior and its applicability for more detailed multidisciplinary investigations related to chemical conditions in muscle performance.

A novel three-filament model of force generation in eccentric contraction of skeletal muscles.

We propose and examine a three filament model of skeletal muscle force generation, thereby extending classical cross-bridge models by involving titin-actin interaction upon active force production. In regions with optimal actin-myosin overlap, the model does not alter energy and force predictions of cross-bridge models for isometric contractions. However, in contrast to cross-bridge models, the three filament model accurately predicts history-dependent force generation in half sarcomeres for eccentric and concentric contractions, and predicts the activation-dependent forces for stretches beyond actin-myosin filament overlap.

MG29/SYPL2 Contributes to Dysregulation of Lipid Composition and Store Operated Ca2+ Entry in Aging Skeletal Muscle

We proposed over one decade ago that age-related muscle weakness and not the loss of muscle size is the principal contributor to disability. In searching for cellular and molecular explanations for this weakness that exceeds the loss of muscle mass, we found that elements of contractile force generation in skeletal muscle are dependent on store-operated Ca2+ entry (SOCE) and that this capacity is lost in aging skeletal muscle. This altered SOCE is directly associated with reduced content of mitsugumin 29 (MG29), a synaptophysin family protein (SYPL2) containing a MARVEL domain involved in cholesterol binding and formation of oligomers of these proteins. MG29 is essential for the formation of the transverse-tubule system and efficient SOCE, creating a link between reduced MG29 expression in aged muscle and the development of muscle dysfunction in sarcopenia as our previous data demonstrate that aspects of aged skeletal muscle are present in young mg29-/- muscle. Here we test how the loss of MG29 affects the phospholipid composition of skeletal muscle fibers and if producing similar changes in muscle fibers can influence SOCE activity. Lipidomic analysis of young mg29-/- skeletal muscle showed decreased cholesterol content and altered levels of various phospholipid species compared to young control muscle. To simulate the reduced cholesterol content observed in mg29-/- muscle we partially extracted it from mature C2C12 myotubes with methyl-β-cyclodextrin (MCD) and then measured SOCE in these myotubes. Reduced SOCE was observed following MCD extraction, similar to the limited SOCE we previously observed in young mg29-/- and wild-type aged muscle fibers. Further studies will determine if mutation of conserved residues in the MARVEL domain of MG29 modulate SOCE function in, and the lipid composition of, skeletal muscle cells.

Investigation of Bisphenol A administration on skeletal muscle force generation in mice

Bisphenol A (BPA) is a chemical used in the manufacturing of numerous products including plastics and food‐contact materials and is classified as an endocrine disruptor because it can mimic actions of estrogen in the body. The purpose of this study was to examine BPA effects on muscle force generation since estrogen may influence muscle contractile properties. Eight‐week old male CD‐1 mice were housed in polypropylene cages and provided a soybean oil‐free diet (AIN‐93G) (control; C) for two weeks. Thereafter, animals were randomly assigned to C (n=5), C+BPA (BPA; 50 mg BPA/kg diet; n=4), or C+ethinyl estradiol (EE; 0.1 ppb; n=5) diets for one month. Soleus muscles were harvested for measurement of twitch and tetanic force generation (N/cm2) in vitro. Peak twitch tension of EE (3.5±0.1) was greater (p<0.05) than BPA (2.6±0.1) and C (2.8±0.2). Peak tetanic tension of EE (24.8±1.2) was greater (p<0.05) than BPA (19.2±0.7) but was similar (p>;0.05) to C (22.31±0.3). Soleus force generation between BPA and C was similar (p>;0.05). Post‐experiment animal body mass and soleus muscle mass did not differ between groups (p>;0.05). Preliminary findings revealed augmented force generation of the soleus following administration of EE, but not BPA. Therefore, BPA administration does not appear to influence twitch or maximal force generation of mouse soleus muscle.Supported by the Hill Collaboration on Environmental Medicine (KCD)

Systems biology of skeletal muscle: fiber type as an organizing principle

Skeletal muscle force generation and contraction are fundamental to countless aspects of human life. The complexity of skeletal muscle physiology is simplified by fiber type classification where differences are observed from neuromuscular transmission to release of intracellular Ca(2+) from the sarcoplasmic reticulum and the resulting recruitment and cycling of cross-bridges. This review uses fiber type classification as an organizing and simplifying principle to explore the complex interactions between the major proteins involved in muscle force generation and contraction.

Regulation of Force Dynamics in Fast Twitch Muscle

Several regulatory steps controlling skeletal muscle force generation have been experimentally demonstrated, and mathematical models have been developed to understand these mechanisms. Here, we investigated the minimal set of mechanisms required to describe both the range in peak force from twitch to tetanus and the characteristics of contraction and relaxation. Hereto, we measured calcium fluorescence using Rhod‐2 and force kinetics in murine EDL muscles at different stimulation frequencies, ranging from twitch to tetanus. Next, we tested available models in literature with a validated spatiotemporal calcium model as input. Monte Carlo simulations, covering a large parameter space, have been performed to test if the evaluated force models could describe the force dynamics.These simulations showed that calcium activation of the cross bridges sufficed to describe the increment in force from twitch to tetanus. In addition, numerical analysis indicated that extending the models with cooperative activation through strongly attached cross‐bridges was required to reproduce the dynamics of contraction and especially of relaxation. Therefore, we hypothesize that the minimal set of mechanisms to describe force dynamics in fast twitch muscle includes both activation by calcium bound to troponin and cooperative activation through strongly attached cross‐bridges.

Effect of xanthine oxidase-generated extracellular superoxide on skeletal muscle force generation

Skeletal muscle contractions increase superoxide anion in skeletal muscle extracellular space. We tested the hypotheses that 1) after an isometric contraction protocol, xanthine oxidase (XO) activity is a source of superoxide anion in the extracellular space of skeletal muscle and 2) the increase in XO-derived extracellular superoxide anion during contractions affects skeletal muscle contractile function. Superoxide anion was monitored in the extracellular space of mouse gastrocnemius muscles by following the reduction of cytochrome c in muscle microdialysates. A 15-min protocol of nondamaging isometric contractions increased the reduction of cytochrome c in microdialysates, indicating an increase in superoxide anion. Mice treated with the XO inhibitor oxypurinol showed a smaller increase in superoxide anions in muscle microdialysates following contractions than in microdialysates from muscles of vehicle-treated mice. Intact extensor digitorum longus (EDL) and soleus muscles from mice were also incubated in vitro with oxypurinol or polyethylene glycol-tagged Cu,Zn-SOD. Oxypurinol decreased the maximum tetanic force produced by EDL and soleus muscles, and polyethylene glycol-tagged Cu,Zn-SOD decreased the maximum force production by the EDL muscles. Neither agent influenced the rate of decline in force production when EDL or soleus muscles were repeatedly electrically stimulated using a 5-min fatiguing protocol (stimulation at 40 Hz for 0.1 s every 5 s). Thus these studies indicate that XO activity contributes to the increased superoxide anion detected within the extracellular space of skeletal muscles during nondamaging contractile activity and that XO-derived superoxide anion or derivatives of this radical have a positive effect on muscle force generation during isometric contractions of mouse skeletal muscles.

Mice Lacking Homer 1 Exhibit a Skeletal Myopathy Characterized by Abnormal Transient Receptor Potential Channel Activity

Transient receptor potential (TRP) channels are nonselective cation channels, several of which are expressed in striated muscle. Because the scaffolding protein Homer 1 has been implicated in TRP channel regulation, we hypothesized that Homer proteins play a significant role in skeletal muscle function. Mice lacking Homer 1 exhibited a myopathy characterized by decreased muscle fiber cross-sectional area and decreased skeletal muscle force generation. Homer 1 knockout myotubes displayed increased basal current density and spontaneous cation influx. This spontaneous cation influx in Homer 1 knockout myotubes was blocked by reexpression of Homer 1b, but not Homer 1a, and by gene silencing of TRPC1. Moreover, diminished Homer 1 expression in mouse models of Duchenne's muscular dystrophy suggests that loss of Homer 1 scaffolding of TRP channels may contribute to the increased stretch-activated channel activity observed in mdx myofibers. These findings provide direct evidence that Homer 1 functions as an important scaffold for TRP channels and regulates mechanotransduction in skeletal muscle.

Response of Equatorial X-Ray Reflections and Stiffness to Altered Sarcomere Length and Myofilament Lattice Spacing in Relaxed Skinned Cardiac Muscle

Low angle x-ray diffraction measurements of myofilament lattice spacing ( D 1,0) and equatorial reflection intensity ratio ( I 1,1/ I 1,0) were made in relaxed skinned cardiac trabeculae from rats. We tested the hypothesis that the degree of weak cross-bridge (Xbr) binding, which has been shown to be obligatory for force generation in skeletal muscle, is modulated by changes in lattice spacing in skinned cardiac muscle. Altered weak Xbr binding was detected both by changes in I 1,1/ I 1,0 and by measurements of chord stiffness (chord K). Both measurements showed that, similar to skeletal muscle, the probability of weak Xbr binding at 170-mM ionic strength was significantly enhanced by lowering temperature to 5°C. The effects of lattice spacing on weak Xbr binding were therefore determined under these conditions. Changes in D 1,0, I 1,1/ I 1,0, and chord K by osmotic compression with dextran T500 were determined at sarcomere lengths (SL) of 2.0 and 2.35 μm. At each SL increasing [dextran] caused D 1,0 to decrease and both I 1,1/ I 1,0 and chord K to increase, indicating increased weak Xbr binding. The results suggest that in intact cardiac muscle increasing SL and decreasing lattice spacing could lead to increased force by increasing the probability of initial weak Xbr binding.

Regulation of Force Development Studied by Photolysis of Caged ADP in Rabbit Skinned Psoas Fibers

The present study examined the effects of Ca 2+ and strongly bound cross-bridges on tension development induced by changes in the concentration of MgADP. Addition of MgADP to the bath increased isometric tension over a wide range of [Ca 2+] in skinned fibers from rabbit psoas muscle. Tension-pCa (pCa is −log [Ca 2+]) relationships and stiffness measurements indicated that MgADP increased mean force per cross-bridge at maximal Ca 2+ and increased recruitment of cross-bridges at submaximal Ca 2+. Photolysis of caged ADP to cause a 0.5 mM MgADP jump initiated an increase in isometric tension under all conditions examined, even at pCa 6.4 where there was no active tension before ADP release. Tension increased monophasically with an observed rate constant, k ADP, which was similar in rate and Ca 2+ sensitivity to the rate constant of tension re-development, k tr, measured in the same fibers by a release-re-stretch protocol. The amplitude of the caged ADP tension transient had a bell-shaped dependence on Ca 2+, reaching a maximum at intermediate Ca 2+ (pCa 6). The role of strong binding cross-bridges in the ADP response was tested by treatment of fibers with a strong binding derivative of myosin subfragment 1 (NEM-S1). In the presence of NEM-S1, the rate and amplitude of the caged ADP response were no longer sensitive to variations in the level of activator Ca 2+. The results are consistent with a model in which ADP-bound cross-bridges cooperatively activate the thin filament regulatory system at submaximal Ca 2+. This cooperative interaction influences both the magnitude and kinetics of force generation in skeletal muscle.

Electron paramagnetic resonance reveals age-related myosin structural changes in rat skeletal muscle fibers.

We tested the hypothesis that low specific tension (force/cross-sectional area) in skeletal muscle from aged animals results from structural changes in myosin that occur with aging. Permeabilized semimembranosus fibers from young adult and aged rats were spin labeled site specifically at myosin SH1 (Cys-707). Electron paramagnetic resonance (EPR) was then used to resolve and quantify the structural states of the myosin head to determine the fraction of myosin heads in the strong-binding (force generating) structural state during maximal isometric contraction. Fibers from aged rats generated 27 +/- 0.8% less specific tension than fibers from younger rats (P < 0.001). EPR spectral analyses showed that, during contraction, 31.6 +/- 2.1% of myosin heads were in the strong-binding structural state in fibers from young adult animals but only 22.1 +/- 1.3% of myosin heads in fibers from aged animals were in that state (P = 0.004). Biochemical assays indicated that the age-related change in myosin structure could be due to protein oxidation, as indicated by a decrease in the number of free cysteine residues. We conclude that myosin structural changes can provide a molecular explanation for age-related decline in skeletal muscle force generation.

Conformation of the myosin motor during force generation in skeletal muscle.

Myosin motors drive muscle contraction, cytokinesis and cell locomotion, and members of the myosin superfamily have been implicated in an increasingly diverse range of cell functions. Myosin can displace a bound actin filament several nanometers in a single interaction. Crystallographic studies suggest that this 'working stroke' involves bending of the myosin head between its light chain and catalytic domains. Here we used X-ray fiber diffraction to test the crystallographic model and measure the interdomain bending during force generation in an intact single muscle fiber. The observed bending has two components: an elastic distortion and an active rotation that generates force. The average bend of the force-generating myosin heads in a muscle fiber is intermediate between those in crystal structures with different bound nucleotides, and the C-terminus of the head is displaced by 7 nm along the actin filament axis compared with the in vitro conformation seen in the absence of nucleotide.

Effects of protein kinase A inhibition on rat diaphragm force generation

Although protein kinases are known to play a role in modulating a variety of intracellular functions, the direct effect of inhibition of these enzymes on skeletal muscle force production has not been studied. The purpose of the present study was to examine this issue by determining the effects produced on diaphragm force generation by two protein kinase inhibitors: (a) H7, an inhibitor of both cAMP-dependent protein kinase (PKA) and of protein kinase C, and (b) H89, a selective inhibitor of PKA. Experiments ( n=15) were performed using isolated, arterially perfused, electrically stimulated rat diaphragms. Perfusate temperature was adjusted to maintain muscle temperature at 27°C and arterial pressure was kept at 150 Torr. Animals were divided into three groups: (a) a control group perfused with Krebs-Henselheit solution equilibrated with 95% O 2/5% CO 2, (b) a group in which H7 (2 μM) was added to the perfusate, and (c) a group perfused with solution containing H89 (4 μM). In all three groups, we assessed diaphragm twitch kinetics, force-frequency relationships and in vitro fatiguability. We found that both H7 and H89 administration slowed twitch relaxation, augmented force generation in response to low frequency stimulation, and increased the rate of development of fatigue. Specifically, for control, H7 and H89 groups, respectively, we found: (a) 1/2 relaxation time averaged 64±2 S.E.M., 87±6 and 90±2 ms, P<0.003, (b) force production during 10-Hz stimulation averaged 12.6±1.1, 20.1±2.3, and 20.3±2.1 N/cm 2, P<0.035, and (c) force fell to 14.3±2.0, 9.5±0.5 and 8.7±0.2% of its initial value after 20 min of fatiguing stimulation, P<0.035. These data show that it is possible to produce large increases in low frequency skeletal muscle force generation by directly inhibiting PKA. We speculate that it may be possible to pharmacologically augment respiratory muscle force and pressure generation in clinical medicine by administration of PKA inhibitors.

Effects of in vivo-like activation frequency on the length-dependent force generation of skeletal muscle fibre bundles.

It is known that a range of firing frequencies can be observed during in vivo muscle activity, yet information is lacking as to how different in vivo-like frequencies may affect force generation of skeletal muscle. This study examined the effects of constant (CSF, constant within one contraction) and decreasing stimulation frequencies (DSF) on mean sarcomere length-force characteristics of rat gastrocnemius medialis fibre bundles. The CSF resulted in an optimal mean sarcomere length (lso) of 2.30 (SEM 0.02), 2.46 (SEM 0.03), 2.76 (SEM 0.03) and more than 2.99 (SEM 0.07) lm, for 100, 50, 30 and 15 Hz, respectively. Compared to 100-Hz stimulation, both lso and the ascending limb of the relationship significantly shifted to higher lengths with lower frequencies. No shift was encountered for the initial part of the descending limb. The DSF reduced the frequency-induced shift to higher mean lengths [lso 2.33 (SEM 0.02), 2.52 (SEM 0.08) and more than 2.92 (SEM 0.10) microm, respectively, for 50, 30 and 15 Hz]. No effect of activation time on length-force characteristics was observed. It was concluded from these studies that the frequency and history of stimulation is a major determinant of the length-force characteristics of muscle fibre bundles, and should be taken into account when analysing animal and human locomotion. The previously observed frequency-induced shift in whole muscle length-force relationship resides mainly at the level of fibre bundles.