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J C Rüegg

Publications and source records attributed to J C Rüegg.

At least 55 records · Page 3Linked to original sources
PubMed1991

Modulation of crossbridge kinetics by myosin isoenzymes in skinned human heart fibers.

Skinned fibers from the normal human heart with the beta-myosin heavy chain (ventricular fibers) revealed both a higher force generation per cross section and a higher Ca2+ sensitivity than skinned fibers with the alpha-myosin heavy chain (atrial fibers). The relation between isometric ATPase activity and isometric tension of atrial fibers was higher than that of ventricular fibers. Since the ATPase-tension relation equals the rate constant for the transition from force-generating into non-force-generating crossbridge states (g(app)), myosin heavy chain isoenzymes seem to have different crossbridge turnover kinetics. Modulation of g(app) by myosin heavy chain isoenzymes could explain the different contractile behavior of atrial and ventricular fibers. g(app) was independent of Ca2+.

Adenosine Triphosphatases↗
PubMed1991

Contribution of cAMP-phosphodiesterase inhibition and sensitization of the contractile proteins for calcium to the inotropic effect of pimobendan in the failing human myocardium.

Previous studies have shown reduced effects of cAMP-dependent positive inotropic agents in the failing human myocardium; thus other cAMP-independent mechanisms of action may be useful to increase force of contraction in this condition. The purpose of this investigation was to determine whether a positive inotropic effect of the cAMP-phosphodiesterase (PDE) inhibitor pimobendan is observed in the failing human myocardium and to study whether other factors, such as an increase in the Ca2+ sensitivity of myofilaments, play a functional role in the increase in force of contraction. Pimobendan produced a positive inotropic effect in isolated preparations from nonfailing donor hearts; however, in moderately (New York Heart Association class II-III, NYHA II-III) and severely (NYHA IV) failing myocardium, this effect was reduced. In addition, in NYHA IV specimens pimobendan inhibited the crude cAMP-PDE (crude PDE) and the isoenzymes I-III (PDE I-III) in a concentration-dependent way. As judged from the IC50 values found in this tissue for the inhibition of PDE III and of crude PDE, the potency of the compound was 18.1 times greater on PDE III. Consistent with a cAMP-PDE-dependent mechanism of action, the positive inotropic effect was potentiated by isoproterenol and inhibited by adenosine in failing myocardium. In failing myocardium, pimobendan also increased the sensitivity of skinned cardiac fibers to Ca2+ and shifted the Ca(2+)-tension relation to the left. This sensitizing effect began at 0.01 mumol/l in NYHA II-III and NYHA IV and rose to about 200% at 300 mumol/l in both groups. In contrast, the demethylated metabolite UD-CG 212 Cl failed to produce positive inotropic effects in failing myocardium alone, but in the presence of isoproterenol, it exerted an increase in force of contraction. The potency of UD-CG 212 Cl for PDE III inhibition in NYHA IV was greater than that of pimobendan. The metabolite pronouncedly decreased the sensitivity of skinned cardiac fibers to Ca2+ at 30-300 mumol/l in NYHA II-III and NYHA IV. It is concluded that in the failing human heart pimobendan inhibited PDE III and sensitized contractile proteins for Ca2+. Both effects appear to be involved in the positive inotropic effect of the compound, because its metabolite, UD-CG 212 Cl, had no effect on force of contraction and on the Ca2+ sensitivity of skinned cardiac fibers but inhibited PDE III even more potently than pimobendan.(ABSTRACT TRUNCATED AT 400 WORDS)

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1991

Higher Ca2+ sensitivity of triton-skinned guinea pig mesenteric microarteries as compared with large arteries.

Small mesenteric resistance arteries and the main branch of the mesenteric artery (outer in situ diameter 115 +/- 3 microns [n = 76] and greater than 1,000 microns, respectively) were skinned with 1% Triton X-100. Both preparations were mounted as rings for circumferential force measurement in an EGTA solution (free Ca2+, less than 10 nM; calmodulin, 0.3 microM; pH 6.7). Force-pCa curves were obtained by increasing free Ca2+ (0.05 to 30 microM). The resulting dose-dependent contractions, after normalization to maximal force development (arteries, 21.4 +/- 2.4 [n = 3]; arterioles, 15.3 +/- 2.1 mN/mm2 [n = 5]) were fitted to sigmoidal force-pCa curves. Values of ED50 and of the cooperativity factor h were 6.08 and 2.39 in arterioles and 5.64 and 1.64 in arteries. The higher Ca2+ sensitivity of arteriolar preparations remained at pH 7.0 at higher calmodulin concentrations and after inhibition of smooth muscle phosphatase with okadaic acid. Total myosin light chain kinase activity in crude arteriolar extracts (using [gamma-32P] ATP and isolated gizzard light chains as substrates) was approximately 25% of arterial kinase. Both kinase preparations had identical Ca2+ sensitivities. Likewise, total arteriolar phosphatase activity (using 32P-labeled gizzard light chains) was approximately 25% of the arterial activity; both phosphatases had an identical sensitivity toward okadaic acid. The ratio of kinase/phosphatase activities was identical in both tissues. Extracts of both tissues contained two isozymes of the myosin heavy chain as determined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Temporal relationship between force, ATPase activity, and myosin phosphorylation during a contraction/relaxation cycle in a skinned smooth muscle.

The temporal relationship between myosin phosphorylation, contractile force and ATPase activity was studied in skinned preparations from the guinea-pig Taenia coli. When free Calcium concentration [( Ca2+]) was increased from pCa (-log[Ca2+]) 9 to pCa 4.5 at low calmodulin concentration (0.05 microM), ATPase activity and myosin light-chain phosphorylation rose quickly, while the increase in force and stiffness was delayed. The time-course of tension increase was faster at higher calmodulin concentrations (5 microM), although the maximal level of phosphorylation was unchanged. Lowering the calcium concentration from pCa 4.5 to pCa 9 at the plateau of contraction caused a rapid decrease in ATPase activity and in myosin phosphorylation, while force and stiffness decayed more slowly. The force decay could be accelerated by inorganic phosphate. These results suggest that, during contraction, force may be produced actively by phosphorylated and ATP-splitting crossbridges, but may be maintained by dephosphorylated crossbridges which cycle slowly. However, force could also be modulated by calmodulin and inorganic phosphate in a manner not involving an alteration in the extent of myosin phosphorylation.

Adenosine Triphosphatases↗
PubMed1990

Stretch-induced increase in the Ca2+ sensitivity of myofibrillar ATPase activity in skinned fibres from pig ventricles.

The increase in force development in the heart with increase in end-diastolic pressure (Frank-Starling mechanism) has been ascribed to an increase in contractile responsiveness of the myofibrils to calcium. We now show that this calcium sensitization is also associated with an increase in calcium responsiveness of the myofibrillar ATPase. Thus, at submaximal Ca activation (pCa 6.0), the ATPase activity of skinned fibres from pig right ventricles is increased from 57.9 +/- 4.4% to 70.6 +/- 4.4% of the maximal Ca2+ activation of ATPase by stretching (by 15% lo). At maximal Ca2+ activation, ATPase was barely altered by stretching. The relationship between ATPase activity of skinned trabecula of pig right ventricle and ATPase-Ca2+ concentrations is shifted (by 0.1 pCa unit) to higher pCa values after a stretch-induced increase of the sarcomere length from 2.1 microns to 2.4 microns. The relationship between force and pCa was affected in a similar way by extension. This increased calcium sensitivity is, however, not associated with an alteration in the relationship between ATPase activity and force development (tension cost). In accordance with Brenner's hypothesis, we propose therefore that stretch activation of ATPase is associated with an increase in the apparent rate constant of crossbridge attachment rather than with a decrease in the apparent rate constant of crossbridge detachment.

Adenosine Triphosphatases↗
PubMed1990

Phosphorylation and thiophosphorylation by myosin light chain kinase: different effects on mechanical properties of chemically skinned ventricular fibers from the pig.

The influence of myosin light chain phosphorylation (treatment with myosin light chain kinase = MLCK, calmodulin and ATP) and thiophosphorylation (incubation with MLCK, calmodulin and ATP gamma S) on the maximal shortening velocity (Vmax) and Ca2+ sensitivity of chemically-skinned ventricular fibers from the pig has been studied. Vmax was determined by the slack-test method and by extrapolation of the force-velocity relation by the isotonic quick release method. Vmax was 1.53 muscle length/s (L/s) and 1.94 L/s using the force-velocity relation and the slack-test, respectively. Phosphorylation increased the Ca2+ sensitivity for isometric force development of skinned fibers but had no influence on Vmax. Thiophosphorylation decreased Vmax but had no influence on Ca2+ sensitivity. Phosphorylation pattern of the myosin light chains of the skinned fibers was studied using [gamma-32P]ATP or [gamma-P35S]ATP (250 muCi each) and autoradiography. Incubation of skinned fibers with labeled ATP led to a phosphate incorporation into the 18-kDa myosin light chain (MPLC or regulatory light chain) while incubation with labeled ATP gamma S led to an incorporation of thiophosphate into the 28-kDa myosin light chain (alkali light chain) and tropomyosin. We suggest that the difference in mechanical behavior between phosphorylated and thiophosphorylated skinned fibers are due to differences in the phosphorylation profiles of myofibrillar regulatory proteins.

Adenosine Triphosphate↗
PubMed1990

Calcium sensitivity and myosin light chain pattern of atrial and ventricular skinned cardiac fibers from patients with various kinds of cardiac disease.

In the present study, the Ca2(+)-sensitivity and myosin light chain patterns of skinned fibers of right atrium and left papillary muscles of 27 patients suffering from mitral valve disease (MVD, moderate heart failure), ischemic cardiomyopathy (ICM, severe heart failure), dilated cardiomyopathy (DCM, severe heart failure), and coronary heart disease (CHD, no heart failure, no atrial hypertrophy) were investigated. Myosin light chains of both chemically skinned and intact samples were studied by two-dimensional gel electrophoresis (2D-PAGE). Ca2(+)-sensitivity of ventricular fibers was about 0.14 pCa-units higher than that of atrial fibers in all groups except dilated cardiomyopathy where this difference was markedly diminished (only 0.06 pCa-units). Generally, Ca2(+)-sensitivity of skinned ventricular fibers was the same among the different heart diseases. Skinned atrial fibers from patients with dilated cardiomyopathy, however, were significantly (about 0.08 pCa-units) more sensitive for Ca2+ than those of the other groups (coronary heart disease, mitral valve disease or ischemic cardiomyopathy) which showed similar Ca2(+)-tension relationships. Ventricle-specific P-light chain forms could be observed in atrial samples from patients of all groups, whereas no atrium-specific light chain forms were detectable in any ventricular sample. It is concluded that there is no difference in Ca2(+)-sensitivity of the ventricular contractile elements of the human heart in different heart diseases. In atrial myocardium, there is an increased Ca2(+)-sensitivity of skinned fibers from hearts with dilated cardiomyopathy which is probably related to an elevation of right atrial pressure.

Calcium↗
PubMed1990

Regulation of myosin heavy chain expression in the hearts of hypertensive rats by testosterone.

Stroke-prone spontaneously hypertensive rats were used for our investigation of the influence of prepubertal gonadectomy and testosterone substitution on blood pressure, cardiac hypertrophy, and the expression of ventricular myosin heavy chain (MHC) isoenzymes at different developmental stages. Blood pressure and the degree of cardiac hypertrophy were decreased by castration and increased by testosterone substitution. We found the same relative distributions of MHC isoforms on the protein level (investigated by pyrophosphate electrophoresis) and on the messenger RNA level (investigated by the polymerase chain reaction). Castration favored the expression of the beta-MHC form, and testosterone substitution enhanced the expression of the alpha-MHC form. These effects were more pronounced in 8-week-old than in 14-16-week-old animals. We conclude that testosterone regulates cardiac MHC expression on a pretranslational level. This regulation is independent of hemodynamic load or cardiac hypertrophy.

Animals↗
PubMed1990

Towards a molecular understanding of contractility.

The contractile performance of the heart depends on the molecular force generators, the myosin crossbridges between thick and thin filaments. These are oscillating between two states, strong and weak, or force-generating and non-force-generating. Calcium activation of the contractile machinery increases the probability of forming force-generating crossbridges, whereas the speed of contraction depends on the probability of returning force-generating crossbridges into weak ones. It is inversely related to the energetic cost of contraction. The probabilities or rate constants describing the transitional states of the force generators (and denoted as g and f) describe not only the kinetic properties of the contractile system but also its responsiveness to the calcium activator. The development of force depends not only on the free calcium ion concentration in the myoplasm, but also on calcium responsiveness which is dependent both on crossbridge kinetics and the calcium affinity of the regulatory protein, troponin. In future, it will be important to find out how all these molecular properties of the force-generators are affected in diseased states as well by pharmacological interventions, such as the application of novel cardiotonic drugs.

Calcium↗
PubMed1989

Calcium sensitivity and unloaded shortening velocity of hypertrophied and non-hypertrophied skinned human atrial fibres.

The mechanical properties of myocardium of different animals are modified by a chronic increase in haemodynamic load. In this study differences in calcium sensitivity and maximum unloaded shortening velocity of hypertrophic and non-hypertrophic chemically skinned human atrial fibres are characterized. Investigating right atria of 34 patients, possible correlations are studied between preoperative atrial pressure, degree of hypertrophy (estimated from the muscle fibre diameter), calcium responsiveness (pCa50 eliciting half-maximum contraction) and Vmax (unloaded shortening velocity). Hypertrophic fibres from atrial appendages of patients having an increased right atrial pressure (RAP 8.5 +/- 1.6 mm Hg) and suffering from mitral valve disease (stenosis and insufficiency combined) had a fibre diameter of 18.0 +/- 0.9 microns. They also had a higher calcium sensitivity (pCa50 5.65 +/- 0.08) and a lower unloaded shortening velocity (1.7 +/- 0.1 muscle lengths/s) than non-hypertrophic fibres from the appendages of patients with normal right atrial pressure (RAP 3.2 +/- 0.5 mm Hg) and coronary heart disease (CHD: pCa50 5.45 +/- 0.04; Vmax = 3.4 +/- 0.2 muscle lengths/s; fibre diameter 12.8 +/- 0.4 microns). Thus non-hypertrophic fibres from control CHD patients differed significantly (p less than 0.01) from hypertrophied atrial fibres of patients with mitral valve disease and with combined valve disease (MAV, pCa50 = 5.58 +/- 0.05, Vmax 2.0 +/- 0.3 muscle lengths/s, fibre diameter 14.6 +/- 0.9 microns) or aortic valve disease (stenosis combined with insufficiency, fibre diameter 14.8 +/- 1.4 microns, pCa50 5.56 +/- 0.03, Vmax 2.0 +/- 0.24 muscle length/s; RAP 11.0 +/- 2.6 mm Hg).(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium↗
PubMed1989

An oil-well method for time-resolved microfluorescence assays.

A simple fluorescence assay method has been devised for direct time-resolved recordings of NADH-coupled enzyme reactions in a microliter sample volume in an "oil-well". The apparatus allows practically instantaneous mixing and stirring of two droplets containing the enzyme and substrate, respectively, and which are viewed with a microscope spectrophotometer. This procedure can be used for instance for enzyme kinetic studies of ATPases on a microscale. Thus, by monitoring NADH-fluorescence changes we determined the time course of ATP splitting in nanogram quantities of myosin subfragment 1. In principle, the method can also be used to determine the time course of any other reaction causing fluorescence changes or luminescence changes.

Calibration↗
PubMed1989

A calmodulin-binding peptide relaxes skinned muscle from guinea-pig taenia coli.

During smooth muscle activation the calcium calmodulin complex interacts with myosin light chain kinase (MLCK) whereby activating it. A synthetic peptide analogue (RS20) corresponding to the calmodulin recognition sequence of MLCK has been synthesized and previously found to inhibit the calmodulin stimulated light chain kinase activity. Here we studied the effect of this peptide on skinned fibers from guinea pig taenia coli. Maximal contractions induced by 30 microM Ca2+ at 0.1 microM calmodulin could be completely relaxed by the peptide at 1 microM. The inhibitory effect was accompanied by partial dephosphorylation only of the regulatory myosin light chain. Relaxation could be reversed by addition of calmodulin which also increased the extent of light chain phosphorylation. The calmodulin concentration required for reversing the inhibition depended on the concentration of the inhibitory peptide suggesting that the peptide competed with MLCK for the calmodulin binding site. As the calcium-calmodulin-peptide mixture constitutes a calmodulin buffer, our results suggest, that the peptide is a calmodulin antagonist unique in terms of its potency and that less than nanomolar concentrations of free calmodulin may be required for inducing smooth muscle contractions.

Animals↗
PubMed1989

Inhibition of TnI-TnC interaction and contraction of skinned muscle fibres by the synthetic peptide TnI [104-115].

Circular dichroism was used to study the induction of helix in TnC or TnI-TnC by the TnI peptide [104-115] at various Ca2+ concentrations. The increase in negative ellipticity and pCa2+ values for the peptide-TnC complex, indicates that binding of the peptide to TnC, induces a small helical conformational change in TnC. This results in an increase in the Ca2+ binding constant and the pCa50 value required to induce 50% of Ca2+ -dependent helix in TnC. The introduction of the peptide to a preformed mixture of TnI-TnC resulted in an increase in negative ellipticity and a decrease in the pCa50 and the apparent Ca2+ binding constant towards the values obtained for the TnI peptide-TnC complex and away from those of TnI-TnC. This demonstrates that the TnI peptide can successfully compete with TnI for TnC and thereby inhibit the TnI-TnC interaction. The addition of the TnI peptide to skinned rabbit psoas or porcine cardiac fibres resulted in the inhibition of the force development and a decrease in the pCa50 values required for 50% Ca2+ activation. The magnitude of the inhibition of tension development and the shift in the Ca2+ sensitivity for skinned cardiac muscle fibres was approximately half that observed with skeletal muscle fibres. In view of the CD findings, these skinned fibre results can be accounted for by the peptide inhibiting the TnI interaction with TnC. However, it is possible that the TnI peptide also has a direct inhibitory effect on TM-actin.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1989

Calcium-sensitivity modulation of cardiac myofibrillar proteins.

Myocardial contractile force may be altered not only by changing intracellular free Ca2+ but also by increasing or decreasing the calcium responsiveness of the myofilaments. The latter effect may be due to interventions that decrease the calcium affinity of troponin C, such as intracellular acidosis or factors that act "downstream" of the calcium-troponin-C interaction in the process of muscle activation. These include inorganic phosphate, which accumulates under hypoxic conditions. Conversely, calcium responsiveness may be increased by drugs that probably increase the calcium affinity of troponin C, such as sulmazole, adibendan, pimobendan, and isomazole. These and related drugs also increase the force development of skinned trabeculae that are submaximally activated by Ca2+ stimulation in ATP salt solution. Thus, we confirmed earlier studies showing that adibendan increased submaximal Ca2+-activated force of pig-heart trabecula skinned by 0.5% lubrol-WX. Qualitatively similar results were obtained with pimobendan. However, calcium responsiveness may not only be modulated by factors influencing troponin, but also by modulation of crossbridge rate constants as well as by other factors that act downstream of troponin C. These include peptides derived from thin-filament proteins and subfragment S1 of myosin.

Animals↗
PubMed1989

Perhexiline increases calcium-activated force in skinned psoas fibres by raising calcium affinity of troponin-C.

We investigated the influence of the calcium-/calmodulin antagonist perhexiline on the calcium sensitivity of skinned muscle fibres from rabbit psoas muscle suspended in MgATP salt solution (pH 7.0; 20 degrees C). Perhexiline strongly sensitized the skinned fibres for calcium; thus the relationship between isometric force development and calcium ion concentration was shifted to the left in the presence of perhexiline. Similar results have been achieved with skinned fibres from pig cardiac (ventricular) muscle fibres. In skinned psoas we also determined the binding of Ca2+ to troponin-C by using the method developed by Güth and Potter (1987). Troponin-C labelled with DANZ (5-dimethyl-aminonaphthalene-2-sulfonyl aziridine) was incorporated into skinned fibres after previous extraction of troponin-C. The fluorescence signal (at 460 nm) as well as force increased when the calcium ion concentration rose to pCa 5.7. Addition of perhexiline increased this force and the fluorescence signal indicating that the increase in calcium responsiveness of the contractile apparatus was, at least partly, due to an increase in calcium occupancy of troponin-C.

Animals↗
PubMed1988

Effects of a protein phosphatase inhibitor, okadaic acid, on membrane currents of isolated guinea-pig cardiac myocytes.

The effects of a protein phosphatase inhibitor, okadaic acid (OA), were studied on membrane currents of isolated myocytes from guinea-pig cardiac ventricle. The whole-cell Ca2+ current (ICa) was recorded as peak inward current in response to test pulse to 0 mV. Extracellular application of OA (5-100 microM) produced an increase of ICa. The effect was markedly enhanced when the myocyte was pretreated with threshold concentrations of isoprenaline. ICa was increased from 11.3 +/- 0.8 microA cm-2 to 19.0 +/- 1.1 microA cm-2 (n = 4) by 5 microM-OA in the presence of 1 nM-isoprenaline. The delayed rectifier current was also slightly increased. Furthermore, the wash-out time of the beta-adrenergic increase of ICa was markedly prolonged by OA. The beta-adrenergic stimulation of cardiac Ca2+ current is thought to be mediated by cAMP-dependent phosphorylation. The present results strongly suggest that the effect of OA on ICa is related to inhibition of endogenous protein phosphatase activity which is responsible for the dephosphorylation process. By the isotope method, the inhibitory effect of OA on different types of phosphatase was compared. OA had a relatively high specificity to type 1-, and type 2A-phosphatases.

Animals↗
PubMed1988

Alteration of calcium sensitivity of skinned frog skeletal muscle fibres by inositol triphosphate and calmodulin antagonists.

We investigated the influence of inositol triphosphate (IP3), trifluoperazine (TFP), and perhexiline on the calcium sensitivity of freeze-dried frog semitendinosus muscle fibres. Further, the effect of IP3 on calcium release from the sarcoplasmic reticulum (SR) of frog semitendinosus fibres skinned by saponin was studied. IP3 decreased the calcium sensitivity of freeze-dried frog skeletal muscle fibres and failed to induce a calcium release from SR of saponin-skinned fibres. Freeze-dried frog skeletal muscle fibres were strongly sensitized for calcium by TFP and perhexiline.

Animals↗