Isometric Twitch Contraction Research Articles

Urea loading enhances postfreeze performance of frog skeletal muscle

The wood frog (Rana sylvatica) is a terrestrial hibernator that can accumulate urea as an osmoprotectant in autumn and winter. This study tested the hypothesis that elevated urea can also function as a cryoprotectant in this freeze-tolerant species. Performance characteristics (threshold stimulus voltage, maximal isometric twitch and tetanic contraction forces, and (1/2) fatigue time) of isolated gastrocnemius muscles were measured before and after experimental freezing at -1.5 degrees C for 18 h, followed by thawing. Frozen/thawed muscles exhibited reduced function relative to baseline (prefreeze) levels; however, muscles preincubated in a saline solution containing urea (80 mmol l(-1)) performed substantially better in some tests than muscles incubated without urea. Concentrations of urea in these treated muscles, approximately 65 mmol l(-1), were within the physiological range in winter R. sylvatica. Reducing tissue urea levels to approximately 33 mmol l(-1) resulted in a similar pattern of response, although the differences between urea-incubated and saline-incubated muscles were not statistically significant. Tests of cryoprotective efficacy were also performed on gastrocnemius muscles from R. pipiens, a closely related, but freeze-intolerant species that hibernates aquatically and thus has little need to accumulate urea. Urea-treated muscles from this species performed no better than muscles incubated in saline, attesting that freeze tolerance cannot be conferred simply by augmenting cryoprotectant levels. Overall, these results bolster an earlier report that urea accumulated in response to low moisture availability can serve a cryoprotective role in freeze-tolerant ectotherms.

Carbonic anhydrases IV and IX: subcellular localization and functional role in mouse skeletal muscle

The subcellular localization of carbonic anhydrase (CA) IV and CA IX in mouse skeletal muscle fibers has been studied immunohistochemically by confocal laser scanning microscopy. CA IV has been found to be located on the plasma membrane as well as on the sarcoplasmic reticulum (SR) membrane. CA IX is not localized in the plasma membrane but in the region of the t-tubular (TT)/terminal SR membrane. CA IV contributes 20% and CA IX 60% to the total CA activity of SR membrane vesicles isolated from mouse skeletal muscles. Our aim was to examine whether SR CA IV and TT/SR CA IX affect muscle contraction. Isolated fiber bundles of fast-twitch extensor digitorum longus and slow-twitch soleus muscle from mouse were investigated for isometric twitch and tetanic contractions and by a fatigue test. The muscle functions of CA IV knockout (KO) fibers and of CA IX KO fibers do not differ from the function of wild-type (WT) fibers. Muscle function of CA IV/XIV double KO mice unexpectedly shows a decrease in rise and relaxation time and in force of single twitches. In contrast, the CA inhibitor dorzolamide, whether applied to WT or to double KO muscle fibers, leads to a significant increase in rise time and force of twitches. It is concluded that the function of mouse skeletal muscle fibers expressing three membrane-associated CAs, IV, IX, and XIV, is not affected by the lack of one isoform but is possibly affected by the lack of all three CAs, as indicated by the inhibition studies.

Muscle contractile properties in children with spastic diplegia

The aim of the present study was to evaluate contractile properties of the plantarflexor muscles in children with spastic diplegia (SD) in comparison of age-matched healthy children. Twelve prepubertal children with SD aged 11g12 years (6 girls and 6 boys) and 12 age- and gender-matched healthy control children (6 girls and 6 boys) participated in this study. Subjects were seated in a custom-made dynamometric chair with the dominant leg flexed 90° at the knee and ankle joints. Twitch contraction characteristics of the plantarflexor muscles were measured by supramaximal electrical stimulation of posterior tibial nerve in popliteal fossa using square-wave pulses of 1 ms duration at rest and after a brief (5 s) isometric maximal voluntary contraction (MVC), i.e., during post-activation potentiation (PAP). Children with SD had significantly lower ( p < 0.05) MVC force, twitch contraction peak force (PF), PAP of twitch force, and twitch maximal rates of force development and relaxation compared to control group. Twitch contraction PF:MVC force ratio was higher ( p < 0.05) in children with SD than in the control group. However, no significant differences in twitch contraction and half-relaxation times were observed between the measured groups. It was concluded that prepubertal children with SD in comparison of normal children are characterized by markedly reduced isometric voluntary and electrically evoked twitch contraction maximal force, capacity for twitch PAP, and rates of twitch force production and relaxation of the plantarflexor muscles. The time–course characteristics of isometric twitch contraction were similar in children with SD and normal children.

Twitch contractile properties of plantarflexor muscles in young and middle-aged recreationally physically active and non-active women

This study examined the effect of recreational physical activity on the contractile properties of skeletal muscles in middle-aged vs young women. A total 74 young (20-29-year-old) and middle-aged (45-54-year-old) women participated. The subjects were distributed into four groups: 1) young recreationally physically active (RPA) (n=19), 2) young recreationally physically non-active (RPN) (n=21), 3) middle-aged RPA (n=23) and 4) middle- aged RPN (n=16). RPA women exercised regularly in groups of recreational gymnastics 2-3 times per week. Isometric twitch of the plantarflexor muscles was evoked by supramaximal electrical stimulation of the tibial nerve in resting and post-activation potentiation state. A greater resting twitch maximal rate of force development (RFD), and potentiated twitch peak force (PF) and maximal rate of relaxation (RR) were observed in the young RPA women compared with the two middle-aged women groups. In young RPN women, these characteristics were greater than in the middle-aged RPN women, whereas they did not differ significantly when compared with the middle-aged RPA women. A shorter resting and potentiated twitch contraction time, and a greater potentiated twitch maximal RFD were found in the young compared with the middle-aged groups. There were no significant differences in twitch characteristics between RPA and RPN women of similar age. The recreational gymnastic type of physical activity did not have a marked effect on twitch contractile properties in young and middle-aged women. A reduced speed of isometric twitch contraction was found in middle-aged women, which was more pronounced in the post-activation potentiation state.

In vivo behavior of muscle fascicles and tendinous tissues in human tibialis anterior muscle during twitch contraction

In this study we investigated the time course of length and velocity of muscle fascicles and tendinous tissues (TT) during isometric twitch contraction, and examined how their interaction relates to the time course of external torque and muscle fascicle force generation. From seven males, supra-maximal twitch contractions (singlet) of the tibialis anterior muscle were induced at 30 degrees , 10 degrees and -10 degrees plantar flexed positions. The length and velocity of fascicles and TT were determined from a series of their transverse ultrasound images. The maximal external torque appeared when the shortening velocity of fascicles was zero. The fascicle and TT length, and external torque showed a 10-30 ms delay of each onset, with a significant difference in half relaxation times at -10 degrees . The time course of TT elongation, and fascicle and tendinous velocities did not differ between joint angles. Curvilinear length-force properties, whose slope of quasi-linear part was ranged from -15.0 to -5.9 N/mm for fascicles and 5.4 to 14.3N/mm for TT, and a loop-like pattern of velocity-force properties, in which the mean power was ranged from 0.14 to 0.80 W for fascicles, and 0.14 to 0.81 W for TT were also observed. These results were attributed to the muscle-tendon interaction, depending on the slack and non-linearity of length-force relationship of compliant TT. We conclude that the mechanical interaction between fascicles and TT, are significant determinants of twitch force and time characteristics.

Genetic CLC-1 chloride channel deficiency modifies diaphragm muscle isometric contractile properties

Genetic deficiency of the muscle chloride channel CLC-1 leads to myotonia congenita in humans as well as myotonia in mice and goats. The hallmark of myotonia is delayed muscle relaxation due to persistent electrical discharges in the muscle. The present study tested the hypothesis that performance of CLC-1 deficient diaphragm muscle is also altered during the contractile phase of the contraction-relaxation cycle. Diaphragm of CLC-1 deficient and wild type mice underwent in vitro isometric contractility testing. Myotonia was easily demonstrable during contractions elicited by train stimulation, but was not seen during twitch stimulation or during train stimulation preceded by a series of twitch stimulations. Twitch force was reduced from 16.7+/-2.5 N/cm(2) in normal muscle to 7.2+/-1.9 N/cm(2) in CLC-1 deficient muscle (P<0.002). Isometric twitch contraction time was shortened from 19.6+/-0.9 to 15.7+/-1.0 ms (P<0.002). During repetitive 25 Hz stimulation, force/area was lower for diseased than normal muscle, whereas force as a percent of initial values declined at a faster rate for normal than diseased muscle. The latter could be accounted for by a rightward shift in the force-frequency relationship of CLC-1 deficient relative to normal muscle, as use of stimulation frequencies which elicited comparable force levels as a percentage of maximum 100 Hz tetanic force led to similar rates of fatigue. These findings indicate that genetic CLC-1 deficiency not only affects muscle relaxation (myotonia) but also modulates diaphragm performance during the contractile phase of the contraction-relaxation cycle.

Mechanism of the Frank–Starling law—A simulation study with a novel cardiac muscle contraction model that includes titin and troponin I

A stretch-induced increase of active tension is one of the most important properties of the heart, known as the Frank–Starling law. Although a variation of myofilament Ca 2+ sensitivity with sarcomere length (SL) change was found to be involved, the underlying molecular mechanisms are not fully clarified. Some recent experimental studies indicate that a reduction of the lattice spacing between thin and thick filaments, through the increase of passive tension caused by the sarcomeric protein titin with an increase in SL within the physiological range, promotes formation of force-generating crossbridges (Xbs). However, the mechanism by which the Xb concentration determines the degree of cooperativity for a given SL has so far evaded experimental elucidation. In this simulation study, a novel, rather simple molecular-based cardiac contraction model, appropriate for integration into a ventricular cell model, was designed, being the first model to introduce experimental data on titin-based radial tension to account for the SL-dependent modulation of the interfilament lattice spacing and to include a conformational change of troponin I (TnI). Simulation results for the isometric twitch contraction time course, the length-tension and the force–[Ca 2+] relationships are comparable to experimental data. A complete potential Frank–Starling mechanism was analyzed by this simulation study. The SL-dependent modulation of the myosin binding rate through titin's passive tension determines the Xb concentration which then alters the degree of positive cooperativity affecting the rate of the TnI conformation change and causing the Hill coefficient to be SL-dependent.

Effect of dopamine on rat diaphragm apoptosis and muscle performance

The purpose of this study was to determine whether dopamine (DA) decreases diaphragm apoptosis and attenuates the decline in diaphragmatic contractile performance associated with repetitive isometric contraction using an in vitro diaphragm preparation. Strenuous diaphragm contractions produce free radicals and muscle apoptosis. Dopamine is a free radical scavenger and, at higher concentrations, increases muscle contractility by simulating beta2-adrenoreceptors. A total of 47 male Sprague-Dawley rats weighing 330-450 g were used in a prospective, randomized, controlled in vitro study. Following animal anaesthetization, diaphragms were excised, and muscle strips prepared and placed in a temperature-controlled isolated tissue bath containing Krebs-Ringer solution (KR) or KR plus 100 microm DA. The solutions were equilibrated with oxygen (O2) at 10, 21 or 95% and 5% carbon dioxide, with the balance being nitrogen. Diaphragm isometric twitch and subtetanic contractions were measured intermittently over 65 min. The diaphragms were then removed and, using a nuclear differential dye uptake method, the percentages of normal, apoptotic and necrotic nuclei were determined using fluorescent microscopy. There were significantly fewer apoptotic nuclei in the DA group diaphragms than in the KR-only group diaphragms in 10 and 21% O2 following either twitch or subtetanic contractions. Dopamine at 100 microm produced only modest increases in muscle performance in both 10 and 21% O2. The attenuation of apoptosis by DA was markedly greater than the effect of DA on muscle performance. Dopamine decreased diaphragmatic apoptosis, perhaps by preventing the activation of intricate apoptotic pathways, stimulating antiapoptotic mechanisms and/or scavenging free radicals.

Estimating in Vitro Mitochondrial Oxygen Consumption During Muscle Contraction and Recovery: A Novel Approach that Accounts for Diffusion

A deconvolution algorithm, based on a Bayesian statistical framework and smoothing spline technique, is applied to reconstructing input functions from noisy measurements in biological systems. Deconvolution is usually ill-posed. However, placing a Bayesian prior distribution on the input function can make the problem well-posed. Using this algorithm and a computational model of diffusional oxygen transport in an approximately cylindrical muscle (about 0.5-mm diameter and 10-mm long mouse leg muscle), the time course of muscle oxygen uptake and mitochondrial oxygen consumption, both during isometric twitch contractions (at various frequencies) and the recovery period, is estimated from polarographic measurements of oxygen concentration on the muscle surface. An important feature of our experimental protocol is the availability of data for the apparatus characteristics. From these time courses, the actual mitochondrial consumption rates during resting and exercise states can be estimated. Mitochondrial oxygen consumption rate increased during stimulation to a maximum steady state value approximately five times of the resting value of 0.63 nmol/s/g wet weight for the stimulation conditions studied. Diffusion slowed the kinetic responses to the contraction but not the steady state fluxes during the stimulation interval.

Enhanced arteriolar alpha-adrenergic constriction impairs dilator responses and skeletal muscle perfusion in obese Zucker rats.

The present study tested the hypothesis that enhanced vascular alpha-adrenergic constriction in obese Zucker rats (OZR) impairs arteriolar dilation and perfusion of skeletal muscle at rest and with increased metabolic demand. In lean Zucker rats (LZR) and OZR, isolated gracilis arterioles were viewed via television microscopy, and the contralateral cremaster muscle or gastrocnemius muscle was prepared for study in situ. Gracilis and cremasteric arterioles were challenged with dilator stimuli under control conditions and after blockade of alpha-adrenoreceptors with prazosin, phentolamine, or yohimbine. Gastrocnemius muscles performed isometric twitch contractions of increasing frequency, and perfusion was continuously monitored. In OZR, dilator responses of arterioles to hypoxia (gracilis), wall shear rate (cremaster), acetylcholine, and iloprost (both) were impaired vs. LZR. Treatment with prazosin and phentolamine (and in cremasteric arterioles only, yohimbine) improved arteriolar reactivity to these stimuli in OZR, although responses remained impaired vs. LZR. Gastrocnemius muscle blood flow was reduced at rest in OZR; this was corrected with intravenous infusion of phentolamine or prazosin. At all contraction frequencies, blood flow was reduced in OZR vs. LZR; this was improved by infusion of phentolamine or prazosin at low-moderate metabolic demand only (1 and 3 Hz). At 5 Hz, adrenoreceptor blockade did not alter blood flow in OZR from levels in untreated rats. These results suggest that enhanced alpha-adrenergic constriction of arterioles of OZR contributes to impaired dilator responses and reduced muscle blood flow at rest and with mild-moderate (although not with large) elevations in metabolic demand.

Leaf extract of Caesalpinia bonduc Roxb. (Caesalpiniaceae) induces an increase of contractile force in rat skeletal muscle in situ.

The pharmacological properties of Caesalpinia bonduc Roxb. (Caesalpiniaceae) are not well known, but it is used traditionally to treat snake bite (Bellomaria and Kacou, 1995; Schaffner, 1997). In the present study, the mechanism through which Caesalpinia bonduc extract (Cebo) affects gallamine-induced relaxation in rat tibial muscle contractility were studied via measurement of isometric-tension-anesthetized, 10-12-week-old, male rats. Isometric twitch contractions of the indirectly-stimulated anterior tibia muscle of the right hindleg were recorded in situ. Cebo administered intravenously (i.v.) increased twitch contractions in a dose-dependent manner. The ED50 value is 2.75 x 10(-4) g/kg body wt. Similar results were obtained using the anticholinesterase neostigmine. In contrast, gallamine (a non-depolarizing muscle relaxant) or the venom of the puff adder Bitis arietans reduced the force of contraction. Treatment with Cebo or neostigmine, however, reversed the relaxation induced by either gallamine or puff adder venom. In conclusion, Cebo stimulates the muscle contractile activity, an effect which may be due to an activation of the cholinergic mechanism.

Chronic Transcutaneous Electrical Stimulation of Calf Muscles Improves Functional Capacity without Inducing Systemic Inflammation in Claudicants

Objectives. To assess whether electrical stimulation of ischaemic calf muscles in claudicants causes a systemic inflammatory response and to evaluate effects of its chronic application on muscle function and walking ability.Design. Prospective randomised controlled trial of calf muscle stimulation.Materials and methods. Stable claudicants were randomised to receive either active chronic low frequency (6 Hz) motor stimulation (n=15) or, as a control treatment, submotor transcutaneous electrical nerve (TENS) stimulation (n=15) of calf muscles in one leg, 3×20 min per day for four weeks. Leucocyte activation was quantified by changes in cell morphology, vascular permeability by urinary albumin:creatinine ratio (ACR), calf muscle function by isometric twitch contractions and walking ability by treadmill performance pre- and post-intervention.Results. Acute active muscle stimulation activated leucocytes less (28% increase) than a standard treadmill test (81% increase) and did not increase ACR. Chronic calf muscle stimulation significantly increased pain-free walking distance by 35 m (95% CI 17, 52, P<0.001) and maximum walking distance by 39 m (95% CI 7, 70, P<0.05) while control treatment had no effect. Active stimulation prevented fatigue of calf muscles during isometric electrically evoked contractions by abolishing the slowing of relaxation that was responsible for loss of force.Conclusions. Chronic electrical muscle stimulation is an effective treatment for alleviating intermittent claudication which, by targeted activation of a small muscle mass, does not engender a significant systemic inflammatory response.

Sternohyoid muscle fatigue properties of dy/dy dystrophic mice, an animal model of merosin-deficient congenital muscular dystrophy.

Humans with merosin-deficient congenital muscular dystrophy have both sucking problems during infancy and sleep-disordered breathing during childhood. We hypothesized that merosin-deficient pharyngeal muscles fatigue faster than normal muscles. This was tested in vitro using sternohyoid muscle from an animal model of this disease, the dy/dy dystrophic mouse. Isometric twitch contraction and half-relaxation times were similar for dy/dy and normal sternohyoid. However, rate of force loss during repetitive 25-Hz train stimulation was markedly diminished in dystrophic compared with normal sternohyoid muscle. Furthermore, force potentiation, which occurred during the early portion of the fatigue-inducing stimulation, had a longer duration in dystrophic compared with normal muscle (approximately 60 versus 20 s). As a result of these two processes, at the end of 2 min of stimulation, force of dystrophic muscle had decreased by 8 +/- 5% and that of normal muscle by 69 +/- 4% (p < 0.0001). The potassium-channel blocker, 3,4-diaminopyridine, increased force of dy/dy sternohyoid muscle during twitch and 25-Hz contractions by 148 +/- 20% (p < 0.00001) and 109 +/- 18% (p < 0.00002), respectively. During repetitive 25-Hz stimulation, force of 3,4-diaminopyridine-treated dystrophic muscle remained significantly higher than that of untreated muscle, despite the early force potentiation being eliminated and fatigue being accelerated. Thus, merosin deficiency reduces fatigue and prolongs the duration of force potentiation. The latter alterations may partially preserve the integrity of upper airway muscle function, without which the severity of pharyngeal complications (feeding problems, sleep-related respiratory dysfunction) might be even more pronounced in the human merosin-deficient congenital muscular dystrophies.

Etruscan shrew muscle: the consequences of being small.

The skeletal muscles of the smallest mammal, the Etruscan shrew Suncus etruscus, are functionally and structurally adapted to the requirements of an enormously high energy turnover. Isometric twitch contractions of the extensor digitorum longus (EDL) and soleus muscles are shorter than in any other mammal, allowing these muscles to contract at outstandingly high frequencies. The skeletal muscles of S. etruscus contract at up to 900 min(-1) for respiration, up to 780 min(-1) for running and up to 3500 min(-1) for shivering. All skeletal muscles investigated lack slow-twitch type I fibres and consist only of fast-twitch type IID fibres. These fibres are optimally equipped with properties enabling a high rate of almost purely oxidative metabolism: they have a small diameter, their citrate synthase activity is higher and their lactate dehydrogenase activity is lower than in the muscles of any other mammal and they have a rapid shortening velocity. Differences in isometric twitch contraction times between different muscles are, at least in part, probably due to differences in cytosolic creatine kinase activities.

Effects of pH on the length-dependent twitch potentiation in skeletal muscle.

When muscle is elongated, there is a length dependence of twitch potentiation and an increased Ca(2+) sensitivity of the myofilaments. Changes in the charge potential of myofilaments, induced by a decrease in pH, are known to abolish the length dependence of Ca(2+) sensitivity. This study was aimed at testing the hypothesis that a decrease in pH, and the concomitant loss of length dependence of Ca(2+) sensitivity, depresses the length dependence of staircase potentiation. In vitro, isometric twitch contractions of fiber bundles dissected from the mouse extensor digitorum longus, performed before and after 10 s of 10-Hz stimulation (i.e., the staircase potentiation protocol) were analyzed at five different lengths, ranging from optimal length for maximal force production (L(o); = 12 +/- 0.7 mm) to L(o) + 1.2 mm (L(o) + 10%). These measurements were made at an extracellular pH of 6.6, 7.4, and 7.8 (pH changes induced by altering the CO(2) concentration of the bath solution). At pH 7.4 and 7.8, the degree of potentiation after 10-Hz stimulation showed a linear decrease with increased fiber bundle length (r(2) = 0.95 and r(2) = 0.99, respectively). At pH 6.6, the length dependence of potentiation was abolished, and the slope of the length-potentiation relationship was not different from zero (r(2) = 0.05). The results of this study indicate that length dependence of potentiation in intact skeletal muscle is abolished by lowering the pH. Because decreasing the pH decreases Ca(2+) sensitivity and changes the charge potential of the filaments, the mechanism of length-dependent potentiation may be closely related to the length dependence of Ca(2+) sensitivity, and changes in the charge potential of the myofilaments may be important in regulating this relationship.

Nitric oxide inhibits smooth muscle responses evoked by cholinergic nerve stimulation in the guinea pig gastric fundus.

In circular smooth muscle tissues of the guinea pig gastric fundus, transmural nerve stimulation (TNS) evoked an atropine-sensitive cholinergic excitatory junction potential (e.j.p.) and, after inhibiting the e.j.p. with atropine, an apamin-sensitive nonadrenergic noncholinergic (NANC) inhibitory junction potential (i.j.p.). The amplitude of e.j.p.s was similar when the frequency of TNS was low (<0.5 Hz), but it decreased successively (depression phenomenon) when the frequency was high (>1 Hz). The depression phenomenon was attenuated after inhibiting the production of nitric oxide (NO) with N(omega)-nitro-L-arginine (NOLA), but was not altered by inhibiting the i.j.p. with apamin. The e.j.p.s were increased in amplitude by the inhibition of cholinesterase activity, but they were decreased by NO produced from SNP with no alteration of their depression phenomenon. Isometric twitch contractions were depressed during high-frequency TNS. NOLA caused an increase in the amplitude of twitch contractions and the attenuation of their depression that changed the transient contraction produced by high-frequency TNS (1 Hz) to a tetanic one. SNP reduced the amplitude of twitch contractions, with no alteration of the depression phenomena. Contractions produced by low concentrations of acetylcholine, but not by high concentrations, were attenuated by SNP, with no alteration of the membrane depolarization. The results suggest that NO produced during TNS has inhibitory actions on cholinergic transmission; the depression of e.j.p.s is mainly prejunctional events, and the depression of mechanical responses is mainly postjunctional events.

Length dependence of staircase potentiation: interactions with caffeine and dantrolene sodium

In skeletal muscle, there is a length dependence of staircase potentiation for which the mechanism is unclear. In this study we tested the hypothesis that abolition of this length dependence by caffeine is effected by a mechanism independent of enhanced Ca2+ release. To test this hypothesis we have used caffeine, which abolishes length dependence of potentiation, and dantrolene sodium, which inhibits Ca2+ release. In situ isometric twitch contractions of rat gastrocnemius muscle before and after 20 s of repetitive stimulation at 5 Hz were analyzed at optimal length (Lo), Lo - 10%, and Lo + 10%. Potentiation was observed to be length dependent, with an increase in developed tension (DT) of 78 +/- 12, 51 +/- 5, and 34 +/- 9% (mean +/- SEM), at Lo - 10%, Lo, and Lo + 10%, respectively. Caffeine diminished the length dependence of activation and suppressed the length dependence of staircase potentiation, giving increases in DT of 65+/-13, 53 +/- 11, and 45 +/- 12% for Lo - 10%, Lo, and Lo + 10%, respectively. Dantrolene administered after caffeine did not reverse this effect. Dantrolene alone depressed the potentiation response, but did not affect the length dependence of staircase potentiation, with increases in DT of 58 +/- 17, 26 +/- 8, and 18 +/- 7%, respectively. This study confirms that there is a length dependence of staircase potentiation in mammalian skeletal muscle which is suppressed by caffeine. Since dantrolene did not alter this suppression of the length dependence of potentiation by caffeine, it is apparently not directly modulated by Ca2+ availability in the myoplasm.

Possible mechanism of selective inotropic activity of the n-butanolic fraction from Berberis aristata fruit

Berberis aristata is an edible plant employed in South Asian traditional medicine; in particular, its fruit is used as a tonic remedy for liver and heart. In isolated cardiac tissues, Berberis aristata fruit extract exhibits a positive inotropic action. Activity-directed fractionation using organic solvents revealed that the cardiotonic activity is concentrated in the n-butanolic fraction (BF). The cardiac action of BF was investigated in spontaneously beating right atria and in electrically driven right ventricular strips and left atria obtained from reserpinized guinea pigs. The results show that this fraction produces a dose-dependent positive inotropic action with little effect on heart rate. To study its possible mode of action, guinea pig atria were pretreated with propranolol, a β-adrenoceptor blocking agent. This treatment abolished the cardiotonic effect of isoprenaline, whereas the cardiotonic effect of BF remained unaltered, suggesting that this effect does not involve stimulation of β-adrenoceptors. On the other hand, application of carbachol reverses only part of the BF-induced increase in ventricular force of contraction, indicating that besides a cyclic AMP (cAMP)-dependent mechanism, a cAMP-independent mechanism underlies the inotropic action of BF. This is in line with the observation that the dynamics of isometric twitch contractions are not significantly altered by BF. Investigations in skinned myocardial preparations showed that BF modulates the calcium-dependent interaction of actin and myosin, apparently by reducing the cooperativity of the calcium-dependent binding of myosin to actin, i.e., there is enhanced calcium activation at low to physiological intracellular calcium, and reduced calcium activation at high intracellular calcium concentrations as present, for example, in ischemic calcium overload. These data indicate that the edible plant, Berberis aristata, contains active principle(s) that cause(s) a selective inotropic effect, involving—in the form of the modulatory effect on actin myosin cooperativity—a novel mechanism of action. Further phytochemical and pharmacological studies may lead to isolation and structural identification of an attractive, new cardiotonic agent from Berberis aristata fruit.

Contraction parameters, myosin composition and metabolic enzymes of the skeletal muscles of the etruscan shrew Suncus etruscus and of the common European white-toothed shrew Crocidura russula (Insectivora: soricidae).

In the Etruscan shrew, the isometric twitch contraction times of extensor digitorum longus (EDL) and soleus muscles are shorter than in any other mammal, allowing these muscles to contract at outstandingly high contraction frequencies. This species has the highest mass-specific metabolic rate of all mammals and requires fast skeletal muscles not only for locomotion but also for effective heat production and for an extremely high ventilation rate. No differences could be detected in the fibre type pattern, the myosin heavy and light chain composition, or in the activity of the metabolic enzymes lactate dehydrogenase and citrate synthase of the two limb muscles, the EDL and the soleus, which in larger mammalian species exhibit distinct differences in contractile proteins and metabolic enzymes. All properties determined in EDL and soleus muscles of Suncus etruscus, as well as in the larger Crocidura russula, are typical for fast-oxidative fibres, and the same holds for several other skeletal muscles including the diaphragm muscle of S. etruscus. Nevertheless, the EDL and soleus muscles showed different mechanical properties in the two shrew species. Relaxation times and, in C. russula, time to peak force are shorter in the EDL than in the soleus muscle. This is in accordance with the time course of the Ca(2+) transients in these muscles. Such a result could be due to different parvalbumin concentrations, to a different volume fraction of the sarcoplasmic reticulum in the two muscles or to different Ca(2+)-ATPase activities. Alternatively, the lower content of cytosolic creatine kinase (CK) in the soleus compared with the EDL muscle could indicate that the observed difference in contraction times between these shrew muscles is due to the CK-controlled activity of their sarcoplasmic reticulum Ca(2+)-ATPase.

Effects of Isoproterenol on Twitch Contraction of Wild Type and Phospholamban-deficient Murine Ventricular Myocardium

Ablation of the gene for phospholamban (PLB), a transmembrane peptide regulator for the cardiac sarcoplasmic reticulum Ca2+pump, in mice brings about a complete loss of the myocardial responses toβ-adrenergic agonists (e.gLuoet alCirc. Res.1994;75: 401). We have evaluated the functional significance of PLB-independent mechanisms in the myocardial responses toβ-adrenergic stimulation in isolated intact ventricular myocardium. We compared the effects of (−)-isoproterenol (ISO) on isometric twitch contraction of paced right ventricular muscle strips of wild type (WT) and PLB-deficient (PLBKO) mice. At 37°C, frequent spontaneous contractions in both types of muscles required the inclusion of lidocaine, an antiarrhythmic, in the bathing medium. Thus the experiments were also performed at two lower temperatures, 30°C and 25°C, at which lidocaine was not needed. Under three conditions, in the absence of ISO, PLBKO ventricular muscles exhibited substantially shortened time to peak tension (TPT) and half relaxation time (TR1/2), compared with the WT muscles. In both WT and PLBKO muscles ISO increased the peak developed tension and decreased TPT and TR1/2in a dose-dependent manner although the effects were generally smaller in PLBKO than in WT muscles. One micromolar ISO caused TPT and TR1/2to decrease by 7.3±1.2% (mean±SEM) and 7.5±1.2% in PLBKO vs. 22.8±0.7% and 29.1±1.7% in WT at 37°C; by 13.5±0.4% and 14.1±1.2% in PLBKO vs. 31.3±0.8%, and 44.8±1.3% in WT at 30°C; by 15.0±2.3% and 21.1±4.9% in PLBKO vs. 25.8±1.9% and 54.0±1.9% in WT at 25°C. These findings strongly suggest that PLB-independent mechanisms play a significant role in mediating the positive inotropic and lusitropic effects ofβ-adrenergic agonists on ventricular myocardium.