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Biomedical subjects

D Pette

Publications and source records attributed to D Pette.

At least 253 records · Page 14Linked to original sources
PubMed1979

Metabolic heterogeneity of muscle fibers classified by myosin ATPase.

Muscle fibers are commonly classified histochemically into three types by the staining intensity for myosin ATPase combined with those for metabolic enzymes. Preincubation at pH 4.6 gives rise to three staining intensities of myosin ATPase which are also used for fiber typing. The two classification systems were compared by computer analysis of the individual staining profiles of over 2,500 fibers, and found not to be equivalent. The analysis showed metabolic heterogeneity among the fiber groups distinguished according to their differences in myosin ATPase.

Adenosine Triphosphatases↗
PubMed1979

Turnover rates of hexokinase I, phosphofructokinase, pyruvate kinase and creatine kinase in slow-twitch soleus muscle and heart of the rabbit.

Hexokinase I was purified from rabbit heart to a specific activity of 70 U/mg protein. The purified enzyme was electrophoretically homogeneous with an apparent molecular weight of 102,000. Purified immunoglobulins from sheep were used to titrate the percentage of hexokinase I in various tissues of the rabbit. Precipitating antibodies from sheep were also prepared against rabbit muscle MM-creatine kinase, phosphofructokinase and pyruvate kinase. Apparent turnover rates of these phosphotransferases and of hexokinase I were determined in rabbit heart and soleus muscle by means of the immunoprecipitation technique after single pulse labelling with [U-14Cl]leucine in vivo. Apparent half-lives of phosphofructokinase, pyruvate kinase and hexokinase I were 0.56 d, 0.73 d and 0.93 d in rabbit heart. In slow-twitch soleus muscle half-lives of phosphofructokinase, pyruvate kinase, hexokinase II and creatine kinase were 0.63 d, 0.72 d, 0.85 d and 0.82 d. The similarity of the rate constants of degradation of these enzymes is interpreted as an indication that different tissue concentrations result primarily from different rates of synthesis.

Animals↗
PubMed1977

ATPase activities, Ca2+ transport and phosphoprotein formation in sarcoplasmic reticulum subfractions of fast and slow rabbit muscles.

Subfractionation of sarcoplasmic reticulum from fast-twitch and slow-twitch rabbit skeletal muscles was performed on a sucrose density gradient. Vesicle fractions were characterized by: measurement of (Ca2+,Mg2+)-dependent (extra) ATPase, Mg2+-dependent (basal) ATPase, Ca2+ uptake characteristics, polypeptide patterns in sodium dodecylsulphate polyacrylamide gel electrophoreses, phosphoprotein formation and electronmicroscopy of negatively stained samples. In fast-twitch muscle, low and high density vesicles were separated. The latter showed high activity of (Ca2+,Mg2+)-dependent ATPase, negligible activity of Mg2+-dependent ATPase, high initial rate and high capacity of Ca2+ uptake, high amount of phosphorylated 115000-Mr polypeptide, and appeared morphologically as thin-walled vesicles covered with particles of 4 nm in diameter. Low density vesicles had little (Ca2+,Mg2+)-dependent ATPase but high Mg2+-dependent ATPase. Although the initial rate of Ca2+ uptake was markedly lower, the total capacity of uptake was comparable with that of high density vesicles. Phosphorylated 115000-Mr polypeptide was detectable at low concentrations. Instead, 57000 and 47000-Mr polypeptides were characterized as forming stable phosphoproteins in the presence of ATP and Mg2+. Negatively stained, these vesicles appeared to have smooth surfaces. It is suggested that low density vesicles represent a Ca2+ sequestering system different from that of high density vesicles and that Mg2+-dependent (basal) ATPase as well as the 57000 and 47000-Mr polypeptides are part of the Ca2+ transport system within the low density vesicles. According to the results from slow-twitch muscle, Ca2+ sequestration by the sarcoplasmic reticulum functions in this muscle type only through the low density vesicles.

Adenosine Triphosphatases↗
PubMed1977

Myosin light chain patterns of individual fast and slow-twitch fibres of rabbit muscles.

Single muscle fibres were isolated by microdissection from freeze-dried samples of rabbit psoas and soleus muscles. The individual fibres were typed according to qualitative histochemical reactions for succinate dehydrogenase or NADH-tetrazolium reductase and for alkaline Ca2+-activated myofibrilla myosin ATPase after acid or alkaline preincubation. Methods are described for electrophoretic analysis by means of polyacrylamide disc electrophoresis in the presence of SDS of total myofibrilla proteins in single fibres after pre-extraction of soluble proteins. Fast-twitch white fibres revealed a myosin light chain pattern characteristic of "fast- type" myosin with three light chains of apparent molecular weights of 22,300 (LC1) 18,400 (LC2) and 16,000 (LC3). Fast-twitch red fibres were indistinguishable in this respect from fast-twist white fibres and showed an identical pattern of myosin light chains. Slow-twitch fibres could be characterized by a myosin light chain pattern typical of myosin of slow-twitch muscles with peptides of the apparent molecular weights of 23,500 (LC1Sa), 23,000 (LC1Sb) and 18,500 (LS2S). Slow-twitch fibres isolated from soleus as well as from psoas muscle were indistinguishable with regard to their myosin light chain patterns, thus suggesting that fibres of the same histochemical type correspond in their myosin light chain patterns irrespective of their origin from different muscles.

Adenosine Triphosphatases↗
PubMed1977

Activity patterns of phosphofructokinase, glyceraldehydephosphate dehydrogenase, lactate dehydrogenase and malate dehydrogenase in microdissected fast and slow fibres from rabbit psoas and soleus muscle.

Methods for standardized determination of phosphofructokinase (PFK), glyceraldehydephosphate dehydrogenase (GAPDH), lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) activities in nanogram samples of microdissected single fibres of rabbit psoas and soleus muscle are described. Fast and slow fibres in soleus muscle show lower absolute activities of these enzymes than the respective fibre types in psoas muscle. Slow fibres represent a more uniform population in the two muscles according to absolute and relative activities of the enzymes investigated. Slow fibres are characterized by high activities of MDH and relatively low activities of glycolytic enzymes. Fast fibres in the soleus muscle represent a population with high activities of MDH and glycolytic enzymes. Fast fibres in psoas muscle represent a heterogeneous population with high activities of glycolytic enzymes and extremely variable activity of MDH. More than 10-fold differences exist in the MDH activities of the extreme types of this fibre population. Differences in the activity levels of MDH in single fast type fibres but also in the activities of glycolytic enzymes between fast and slow fibres are greater than those reported between extreme white and red rabbit muscles.

Animals↗
PubMed1977

Regulation of glycogen synthase interconversion in cultured muscle cells: actions of insulin, calcium, ionophore A 23187 and cytochalasin B.

The glycogen content of cultured chick embryo breast muscle cells changes during their development and can be reduced by starvation. It is demonstrated that the rate of glucose incorporation into glycogen and the degree of interconversion of glycogen synthase are controlled by the actual glycogen content. Stimulation of both corresponding activities by insulin is found in fused and in unfused cells. The insulin response depends on the extracellular calcium concentration and can be mimicked by the ionophore A 23187. These metabolic effects as well as calcium efflux data confirm the hypothesis that insulin acts on its enzyme target via increased cytoplasmic calcium concentration. Cytochalasin B is shown to inhibit the interconversion but does not interfere with the insulin-induced increase of the mitochondrial calcium pool or with the acceleration of the calcium efflux out of 45C-preloaded myotubes.

Animals↗
PubMed1976

Time dependent effects on contractile properties, fibre population, myosin light chains and enzymes of energy metabolism in intermittently and continuously stimulated fast twitch muscles of the rabbit.

Fast-twitch tibialis anterior and extensor digitorum longus rabbit muscles were subjected to long-term intermittent (8 h daily) or continuous (24 h daily) indirect stimulation with a frequency pattern resembling that of a slow motoneuron. Increases in time to peak of isometric twitch contraction were observed without parallel changes in the pattern of myosin light chains or alterations in the distribution of slow and fast fibres as discernible by the histochemical ATPase reaction. However, changes in the fibre population and in the myosin light chain pattern were observed after intermittent stimulation periods exceeding 40 days or continuous stimulation periods longer than 20 days. Under these conditions even higher increases were found in contraction time. In one animal a complete change in fbire population was observed. In this case myosin light chains of the slow (LCS1, LCS2) and of the fast type (LCf1) were obviously synthetized simultaneously within the same fibre. Early changes in the enzyme activity pattern of energy metabolism indicated a conversion of the fibres including their mitochondrial population. These changes and the earlier reported changes in the sarcoplasmic reticulum are probably responsible for the early changes in contractile properties which occur before the transformation of the myosin.

Adenosine Triphosphatases↗
PubMed1976

Insulin action on glucose transport and calcium fluxes in developing muscle cells in vitro.

The glucose transport system of developing muscle cells in vitro is described. Uptake experiments showed stereospecificity but no saturation or competition. Km-values for early myoblasts could not be determined; with progressive development, the absolute amount of uptake increases and becomes insulin sensitive. Km-values of about 13 mM basal and 32 mM for insulin stimulation were determined and discussed with respect to the glucose transport mechanism. Basal and insulin-dependent transport were shown to be dependent on the presence of calcium in the medium. Moreover, the ionophore A 23187 could be shown to mimic the insulin effect on glucose uptake. 45Ca saturation and desaturation experiments showed that insulin enhances the "mitochondrial" calcium pool by 35% and increases the apparent "cytoplasmic" efflux rate constant by 50%. We propose that insulin increases the free cytoplasmic calcium concentration, which may be the intracellular signal for the stimulation of glucose transport.

Animals↗
PubMed1975

Influence of intermittent long-term stimulation on contractile, histochemical and metabolic properties of fibre populations in fast and slow rabbit muscles.

Slow (m.soleus) and fast (m.tibialis anterior) muscles of the rabbit were subjected to indirect long-term intermittent stimulation (3 weeks, 8 hrs daily) with a frequency pattern of 10 imp/sec. Whereas no changes were observed in case of the slow muscle, stimulation induced profound changes in the fast tibialis anterior muscle. These consisted in a rearrangement of the enzyme activity pattern of energy-supplying metabolism, e.g. decrease in glycogenolytic and glycolytic enzyme activities and severalfold increase in key enzymes of aerobic endoxidation of substrates in beta-oxidation and the citric acid cycle. Concomitant with the increase in aerobic oxidative capacity, there was an increased resistance to fatigue. Histochemical studies revealed a strong increase in mitochondria of all fibres. The bimodal distribution of fibre cross-sectional area in the normal tibialis anterior muscle was changed by stimulation into a more homogeneous population of fibres with a smaller cross-sectional area. Despite a 50% increase in time to peak of isometric twitch contraction no changes were observed in the fibre population with regard to myofibrillar ATPase reaction in quantitative evaluation of whole cross-sections of the muscles. The percentage of fibres histochemically classified as slow amounted to 2.8% and 3.1% in control and stimulated tibialis anterior muscle. Nevertheless the data suggest a transformation of the fibre population under the influence of long-term intermittent stimulation.

Adenosine Triphosphatases↗
PubMed1975

Calcium-related changes of enzyme activities in energy metabolism of cultured embryonic chick myoblasts and myotubes.

Changes in activity of enzymes involved in energy metabolism have been determined in unfused, fused as well as in fusion-inhibited chick embryo muscle cells in vitro. Functionally related enzymes which supposedly are coded by "gene clusters" show a similar degree and rate of enzyme activity increase. Hexokinase and glucose-6-phosphate dehydrogenase reveal only slight activity changes during muscle cell development under the conditions studied. The elevation of phosphofructokinase can be distinguished from that of the other glycolytic enzymes by its higher rate of increase and from that of phosphorylase by its time-course of activity change. The Ca2+ dependence of the phosphorylase activity increase runs parallel to myoblast fusion rate. Experiments in which calcium was removed from cultures which had reached the final morphological state of mature myotubes 24 h after onset of fusion show that increases of enzyme activities are irreversible and that these increases proceed at unchanged rates. Experimental evidence suggest that although fusion and enzyme syntheses may be uncoupled, both are similarly triggered by being dependent on Ca2+ concentration.

Animals↗
PubMed1975

Immunofluorescent localization of glycogenolytic and glycolytic enzyme proteins and of malate dehydrogenase isozymes in cross-striated skeletal muscle and heart of the rabbit.

Specific antisera against glycogen phosphorylase, phosphofructokinase, aldolase, glyceraldehyde-phosphate dehydrogenase, enolase, lactate dehydrogenase, cytosolic and mitochondrial malate dehydrogenase from rabbit muscle were obtained from sheep. The gamma-globulins were used for indirect immunofluorescent localization of the respective enzymes in rabbit skeletal muscle and heart. In stretched skeletal muscle a cross-striation like distribution was observed for all enzymes studied. In the case of mitochondrial malate dehydrogenase this pattern is due to the staining of I-band mitochondria. In cross-sections, an intense staining of the sarcolemma and of subsarcolemmal mitochondria was observed. Comparative analyses with polarized light revealed that the cytosolic enzymes under study are distributed in the relaxed muscle fibre predominantly within the isotropic zones. The same distribution holds also for heart. In contracting muscle a decrease in cross-striated fluorescence and a faint staining of the interfibrillar spaces suggests a location also within the interfibrillar space.

Animals↗