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This investigation was designed to determine whether repeated exposure to high sustained +Gz acceleration induces persisting changes in the myocardial energetic metabolism. Rats were exposed to three plateaus of 30 s at 10 +Gz, four times a week, for 4 weeks. Myocardial concentrations of high-energy phosphorylated compounds were evaluated by 31P-nuclear magnetic resonance (NMR) spectroscopy on isolated hearts submitted to isovolumic aortic perfusion. Heart performances were recorded using the intraventricular balloon method. Compared to the hearts of control rats (n = 5), the hearts of centrifuged rats (n = 5) had higher concentrations of inorganic phosphate (Pi:1.40 +/- 0.33 nM vs. 0.36 +/- 0.07 mM; p < 0.01), decreased phosphocreatine concentrations (PC:15 +/- 0.39 mM vs. 15.69 +/- 0.19 mM; p < 0.01), and a lower left ventricular developed pressure (LVDP) (21 +/- 1 mmHg vs. 34 +/- 2 mmHg; p < 0.01). The workload was increased by sequential augmentation of calcium in the perfusion medium. The relationship between LVDP and the Pi/PC ratio showed that the cost of the cardiac work was greater for the centrifuged rats.

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The effect of calcium activation on energy production was investigated in isolated perfused hearts from rats treated with triiodothyronine (T3) during 15 days (0.2 mg/kg/day) and in hearts of rats allowed to recover after T3-treatment during 15 days. Changes in phosphorylated compound concentrations were followed in the isolated hearts perfused with a glucose-pyruvate medium by 31P-NMR spectroscopy, when the external calcium concentration was increased from 0.5-1, 1.5 and 2 mM. As expected, T3-treatment resulted in the hypertrophy of the heart (50% increase in HW/BW) that was nearly reversible 15 days after discontinuation of the treatment. When compared to controls, creatine, phosphocreatine (PCr) and glycogen contents were lower (58, 24 and 17% decrease respectively) in the hypertrophied hearts and higher (10, 14 and 18% respectively) after regression of hypertrophy. Intracellular pH, ATP, inorganic phosphate concentrations and the phosphorylation potential were not altered under T3-treatment and after regression of hypertrophy, while calculated free ADP concentration was lower in hypertrophied hearts (control: 40 +/- 2 microM, T3-treatment: 21 +/- 1 microM, regression: 37 +/- 1 microM). Increasing the calcium concentration induced a similar increase in left ventricular developed pressure in the three groups of hearts, with inorganic phosphate concentration increasing with cardiac work. The PCr concentration slightly decreased while the ATP concentration did not change. In spite of different initial PCr concentrations, the evolutions of PCr and Pi concentrations for each stepwise increase in external calcium were similar in the three groups. It is concluded that, in spite of the well-known decrease in efficiency induced by the drug, the mechanisms of PCr (ATP) production remain able to respond to an acute moderate increase in energy demand provoked by a physiological stimulus. This adaptation also persists after the treatment when the energy metabolism balance is apparently improved.

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Rat hearts, arrested in situ after intracaval injection of a cardioplegic solution, were preserved for 15 hours at 4 degrees C either by simple immersion or by low-flow (0.3 mL/min) perfusion. After preservation under both conditions, the left ventricular pressure developed by the reperfused hearts reached 8% and 43% of the control value (80 mm Hg), respectively. The addition of trimetazidine (TMZ; 10(-6) M) to the cardioplegic solution induced an improvement in functional recovery (by 2.4 and 1.5, respectively). This effect of TMZ was accompanied by a better energy profile illustrated by a 2-fold increase in the adenosine triphosphate to inorganic phosphate ratio and a reduction of intracellular acidosis as determined by 31P nuclear magnetic resonance spectroscopy. The function of the mitochondria (state 3, reduced nicotinamide-adenine dinucleotide [NADH] formation) isolated from the preserved hearts was significantly depressed in the stored hearts. The addition of TMZ to the cardioplegic solution partially protected oxoglutarate (and succinate) mitochondrial respiration and induced an increase in Ca2+ triggered NADH formation. These results show that the bioenergy status of the myocardial cell in isolated arrested stored rat heart is improved by the presence of TMZ in the preservation solution. Moreover, the experiments demonstrate that this effect includes protection of mitochondrial function and suggest that the drug could exert some control in the Ca2+ regulation of mitochondria.

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The dynamics of the changes in myocardial phosphorylated compound contents (inorganic phosphate: Pi; phosphocreatine: PCr; ATP) induced by 10(-6)M isoprenaline administration was studied, using 31P-NMR spectroscopy, in hearts isolated from rats adapted for three weeks to normobaric hypoxia (10% of oxygen). When compared with the behaviour of control hearts, the inotropic response to Ca2+ and isoprenaline was larger in the hearts from hypoxic rats, while the oxygen consumption was similar. During administration of isoprenaline, a decrease in the myocardial contents of high energy phosphates (ATP and PCr) and an accumulation of Pi was observed in both groups. After action of isoprenaline, the hearts from hypoxic animals showed significant overshoot of PCr, that was not seen in hearts from normoxic rats. The mechanisms of these alterations are analysed and the phosphocreatine overshoot, as well as the increased rate pressure product to oxygen consumption ratio, are assumed to indicate more efficient energy conversion in the heart from animals adapted to chronic hypoxia.

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OBJECTIVE: The aim was to test the hypothesis that aging may change the function and energetics of isolated hearts subjected to an increased work load induced by varying the calcium concentration ([Ca2+]) in the perfusion fluid from 0.5 to 1.0, 1.5, or 2.0 mM. METHODS: Female Wistar rats aged 4, 12-14, and 22-24 months were used. Their hearts were perfused through the aorta and changes in myocardial phosphorylated compound concentration were measured by means of 31P-nuclear magnetic resonance spectroscopy and biochemical analysis. Myocardial contractility was measured in situ in closed heart anaesthetised animals and was followed in vitro during perfusion. RESULTS: The contractile indices measured in situ revealed a decrease with aging of the left ventricular developed pressure and dP/dtmax, while heart rate did not show any significant difference. In all age groups the stepwise increase in [Ca2+] caused a graded increase in left ventricular pressure in the perfused hearts. In aged rats, the left ventricular pressure associated with the different concentrations of Ca2+ was significantly lower than in young rats. In all three groups the myocardial content of inorganic phosphorus (Pi) increased in response to a rise in [Ca2+] and left ventricular pressure. The ATP content of the hearts remained constant in all three groups at each value of [Ca2+] induced left ventricular pressure. However, both ATP and total adenine nucleotide contents of the hearts were lower in aged rats. When the alteration in Pi due to each increase in [Ca2+] was expressed in relation to the rise in left ventricular developed pressure, this relationship was not significantly different in the three groups of hearts. CONCLUSIONS: The reduced mechanical activity of aged rat hearts is not due to a diminished efficiency of the mechanisms transferring high energy phosphates.

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The effect of different chronic blood pressure levels on cardiac energy metabolism was studied by 31P-NMR spectroscopy in perfused hearts from the Lyon strains of hypertensive (LH), normotensive (LN) and hypotensive (LL) rats at the ages of 12 and 21 weeks. The in vivo assessment of haemodynamic parameters measured at 21 weeks in anaesthetized rats with an ultraminiature catheter pressure transducer confirmed that left ventricular systolic pressure and mean aortic pressure were significantly greater (+25%) in LH rats than in LN and LL rats. In the LL animals, left ventricular systolic pressure was slightly reduced (-10%) and cardiac contractility (estimated by LV dP/dtmax) showed a 24% decreased compared to normotensive animals. The energy state of the cardiomyocytes was characterized at different work levels of isolated rat hearts, by determining the concentration of the free phosphorylated compounds at each work level. Changes in workload were induced by varying the calcium concentration in the perfusion fluid. Increasing extracellular calcium concentration resulted in a similar increase in left ventricular developed pressure (LVDP) in all groups studied. Intracellular pH was not influenced by either the age of the animals or the level of cardiac work, in the three groups of animals. ATP content of the LN and LL rats remained constant during the whole perfusion period while the 12 week-old LH rats showed a decreased ATP content with increasing cardiac work. In the older LH rats, ATP content was decreased at the highest work level (corresponding to 2 mM calcium). In response to the increase in work, phosphocreatine (PCr) content diminished and inorganic phosphate (Pi) content increased in both LN, LH and LL animals. PCr degradation and Pi accumulation were higher in the LH rats and less in LL rats compared to the LN. These changes were more important in the younger than in the older hypertensive animals. The relationship between LVDP and [Pi]/[PCr] indicates that oxidative metabolism is maximally activated in the young hypertensive rats and suggests that this maximal activation represents an adaptive phase to the increase in blood pressure. Since the difference between the metabolic pattern of the 21 week-old LH rats and age-matched LN rats was less pronounced, it is likely that a compensatory stage has been reached at that age.

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It was the aim of this study to evaluate the effects of hyperthyroidism on heart function and cardiac energy metabolism of spontaneously hypertensive (SHR) rats. Hyperthyroidism was induced by daily injections of T3 (0.2 mg/kg s.c.) for 14 days. The hearts were then isolated and perfused in the Langendorff mode. ATP, phosphocreatine (PCr), and inorganic phosphate (Pi) were measured continuously by means of 31P-nuclear magnetic resonance (NMR) spectroscopy. Work load was altered by varying stepwise the Ca++ concentration in the perfusion fluid from 0.5 to 1.0, 1.5, and 2.0 mM, respectively. At every elevation of the Ca++ concentration, the increase in left ventricular developed pressure (LVDP) was higher in the hyperthyroid SHR than in the untreated SHR hearts. The ATP and PCr concentrations were lower in the hyperthyroid SHR compared to the untreated SHR hearts throughout the perfusion period. PCr decreased at every Ca++ elevation in both the untreated and hyperthyroid SHR hearts. The PCr/ATP ratio was not altered at any Ca++ concentration neither in the untreated SHR nor in the hyperthyroid SHR hearts. The Ca(++)-induced stepwise elevation in LVDP was higher at any given PCr/Pi ratio in the hyperthyroid SHR than in the untreated SHR hearts. Thus, the Ca(++)-inducible contractile reserve was greater in the hyperthyroid SHR heart.

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Rat hearts, arrested in situ after intracaval injection of a simple mineral cardioplegic solution (St. Thomas), were preserved for 15 h at 4 degrees C either by simple immersion in the cardioplegic solution or low-flow perfusion by the same liquid (0.3 ml/min). Trimetazidine (TMZ) was added to the cardioplegic solution at a concentration of 10(-6) M in two additional groups corresponding to both preservation conditions. The biochemically determined ventricular ATP content was increased in immersed hearts by TMZ (+ 78%). The [ATP]/[Pi] ratio, as calculated from 31P nuclear magnetic resonance (NMR) spectroscopy, was significantly increased in both preservation conditions by TMZ: 0.08 +/- 0.03 versus 0.04 +/- 0.01 in immersed hearts and 0.53 +/- 0.13 versus 0.26 +/- 0.16 in perfused hearts (mean +/- SD). Intracellular pH was increased when TMZ was administered in perfused hearts (7.02 +/- 0.14 vs. 6.67 +/- 0.14, mean +/- SD). Functional recovery, estimated by the pressure developed by the left ventricle (intraventricular isovolumic balloon) when hearts were reperfused with a physiologic solution, was improved by TMZ in both preservation conditions: +137% in immersed hearts and +54% in perfused hearts. These results demonstrate that both the bioenergetic status of the myocardial cell and the functional capabilities of isolated, arrested, stored rat heart can be significantly improved by addition of the antiischemic drug TMZ to the preservation solution.

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We report studies on the isolated hearts of rats treated with triiodothyronine (0.2 mg/kg daily) for 14 days, on spontaneously hypertensive rats (12 and 21 weeks old, Lyon strain) and on their respective controls. A 30% increase in cardiac weight was developed with triiodothyronine and a 40% increase in heart weight in the presence of spontaneous hypertension. The hearts were perfused in the presence of 2 mM pyruvate and the intracellular content of phosphocreatine, inorganic phosphate and ATP measured by nuclear magnetic resonance spectroscopy with 31P. The left ventricular developed pressure was measured with an intraventricular balloon. Changes in contractile strength were induced by stepwise modifications of the extracellular concentration of calcium from 0.5 mM to 1.0, 1.5 and 2.0 mM. In all experimental groups, each increase in the extracellular calcium induced an increase in the developed pressure, together with a decrease in phosphocreatine and an increase in inorganic phosphate; the ATP level remained unchanged. These metabolic changes increased progressively with the increase in developed pressure. In the hearts of animals treated with triiodothyronine and of the 21 weeks old hypertensive rats, the extent of changes in phosphocreatine and inorganic phosphate was the same as in the controls; but, in the hearts of 12 weeks old hypertensive rats, the changes were significantly greater than in their controls. These observations suggest that, during the development of cardiac hypertrophy from spontaneous hypertension, there is a transitory deficiency in the capacity for aerobic ATP production relative to the rate of hydrolysis of ATP induced by an inotropic effect.

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Twenty millimolar and 2 mM uridine triphosphate and 2 mM uridine were injected intra-arterially into rat leg muscles during a 20 min period of intense exercise and during the recovery phase (20 min). Administration of 20 mM uridine triphosphate during exercise, provoked a complete depression of muscle contractility. On the contrary, supply of 2 mM uridine triphosphate or 20 mM uridine induced a reduction in the decrease of muscular developed tension during the exercise period of time and favoured functional recovery. This positive effect of pyrimidine compounds on muscular functional parameters did not seem to be correlated with metabolic effects. Indeed, 2 mM uridine supply did not modify the evolution of intramuscular glycogen and lactate concentrations and made worsened the adenine nucleotide degradation induced by exercise.

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The potential role of phosphorylated compounds in the control of myocardial cell respiration was investigated by means of 31P-nuclear magnetic resonance (NMR) spectroscopy. Isolated isovolumic rat hearts, perfused with a 9 mM glucose, 2 mM pyruvate medium at a constant beating rate (6 Hz) and temperature (37 degrees C), were subjected to changes in work load by varying the calcium concentration ([Ca2+]) in the perfusion fluid from 0.5 to 1.0, 1.5, or 2.0 mM. Each change in left ventricular developed pressure (LVDP) induced by the [Ca2+] change was accompanied by alterations in the inorganic phosphate-to-creatine phosphate ratio ([Pi]/[PCr]), with the ATP level remaining constant. The relationship between [Pi]/[PCr] and LVDP followed a Michaelis-Menten pattern with an apparent Michaelis constant (Km) of 0.09 and a maximal LVDP of 91 mmHg. This Km corresponded to intracellular concentrations of 1.2 mM for Pi and 13.0 mM for PCr. The calculated [ADP] and phosphorylation potential corresponding to these values were 44 microM and 151,000 M-1, respectively. All these values are close to those estimated under in situ physiological conditions. These results support the assumption that in the rat heart, as in skeletal muscle, mitochondrial activity could be controlled by changes in phosphorylated compound concentrations under normoxic conditions.

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Comparison of rat heart preservation by simple storage in a cardioplegic solution at 4 degrees C (6 hr for group I; 15 hr for group II) and by hypothermic low-flow perfusion of the same solution (0.3 ml min-1, 15 hr: group III) was performed by measuring biochemical and functional parameters and by collecting 31P-NMR spectroscopy data. When compared to control values, adenine nucleotide levels remained unchanged in group I hearts, while glycogen was 45% hydrolyzed and lactate level increased by 700%. Extension of heart immersion to 15 hr (group II) led to breakdown of ATP (-77%), of the sum of adenine nucleotides (-27%), and of glycogen (-77%), whereas lactate accumulation reached 900% of the control value. Functional recovery, measured at the end of a 60-min reperfusion was less than 10% in group II hearts when compared to group I hearts. This dramatic development was completely avoided by hypothermic low-flow perfusion (group III). 31P-NMR data showed that phosphocreatine was completely degraded in all groups of preserved hearts. Low-flow perfusion limited cellular acidosis. The ATP/Pi (Pi = inorganic phosphate) ratio calculated from NMR data was lower for group II hearts (0.04 +/- 0.01, n = 6) than for group I hearts (0.29 +/- 0.12; n = 6) or group III hearts (0.19 +/- 0.09; n = 6) and could constitute a convenient bioenergetic index to predict the capability of the heart to recover satisfactory contractility following a preservation period.

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The phosphorylation of 5 microM cytidine by the isolated perfused rat heart was increased by 50 and 120% respectively, when the myocardial UTP content was increased by 30%, 1 hr. after the treatment of animals with isoprenaline (5 mg/kg; s.c.) and by 50%, 6 hrs. after the treatment of animals with galactosamine (430 mg/kg; i.p.). On the other hand, this rate of synthesis of cytidylic nucleotides from cytidine was not decreased when the intracellular UTP and cytidylic nucleotide levels were increased through 130 and 90% respectively (12 hrs. after isoprenaline treatment).

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These experiments were designed to determine through the study of uridine and cytidine kinase activity, the precise mechanisms of plasma nucleoside salvage leading to pyrimidine nucleotide synthesis in the rat heart. The kinetic parameters were: Km = 10 microM, V = 4 nmol g-1 min-1 for cytidine kinase activity and Km = 43 microM and V = 18 nmol g-1 min-1 for uridine kinase activity. Competing activity as concerns the two nucleosides was shown to occur, suggesting that in the rat myocardium as in other cells, one and the same enzyme phosphorylates both uridine and cytidine. UTP and CTP were shown to exert a potent inhibitory action on nucleoside phosphorylation; two factors thus exert a joint influence on the control of pyrimidine nucleotide synthesis in the rat heart: the extracellular concentration of precursor and the intracellular level of UTP and CTP. The kinetic parameters for kinase activities are discussed, taking into account the actual concentration of plasmatic nucleosides. Comparison of these data with respectively those for incorporation of nucleosides into the pyrimidine nucleotides of isolated rat heart and with nucleotide turnover rates in vivo suggests that, under physiological conditions, the utilization of plasma cytidine is crucial to the synthesis of myocardial pyrimidine synthesis.

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The rate of synthesis of myocardial proteins and ribosomal ribonucleic acid (rRNA) was measured during the development of cardiac hypertrophy in rats using a continuous intracardiac infusion of 14C-tyrosine and 3H-uridine in unanaesthetised animals. Cardiac overload was induced by abdominal aortic stenosis. Left ventricular weight and total myocardial RNA concentration were significantly increased on day 4 after aortic stenosis (+19% and +18% respectively). On day 8 left ventricular weight reached +52% whereas RNA concentration had not increased further (+13%). The fractional turnover rates were calculated using the specific activities of intracellular free tyrosine and free uracil nucleotides (precursors) and those of protein bound tyrosine and 28S rRNA bound uridine monophosphate (products) respectively. The fractional rate of synthesis of proteins and rRNA (expressed as percentage per day) increased from 24% to 45% for proteins and from 25% to 34% for rRNA and peaked by day 2. The RNA activity, expressed as gram of protein synthesised per day and per gram of total RNA, was unchanged on day 1 and reached a maximal value on day 2 (+107%). These results suggest that the pre-existing ribosomal RNA could be underutilized under control conditions and that the boosting of RNA transcription, associated with that of protein translation, is a complementary process rather than a prerequisite for the transition period leading to hypertrophy.

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The rate of incorporation of exogenous cytidine into the pyrimidine nucleotides of isolated perfused rat hearts bears a Michaelis-Menten kinetic relationship to the extracellular concentration of the precursor, with an apparent Km of 10 microM and a maximal rate of incorporation of about 60 nmol per gram (ww) per 30 minutes. The myocardial cytosine nucleotide content was significantly increased by a 30 minute supply of 15 microM cytidine and more than doubled after a 2 hour administration period of 50 microM cytidine. In the latter case, the rate of cytidine incorporation fell by 30% (with respect to the initial uptake) during the last 30 minutes of administration. These results suggest that the uptake of cytidine can play an important part in the myocardial cytosine nucleotide metabolism under physiological conditions.

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[14C]inosine in a range of concentrations of 20 microM to 1 mM was administered to the isolated perfused rat heart for 30 min. The incorporation of the nucleoside into myocardial adenine nucleotides increased for extracellular concentrations of the precursor up to 50 microM, reaching a plateau at 60 nmol . g-1 X 30 min-1 with concentrations ranging between 50 and 200 microM. The supply of 500 microM and 1 mM of inosine induced a further increase in cardiac adenine nucleotide synthesis to about 200 nmol . g-1 X 30 min-1. When supplied during low flow ischaemia (0.5 mL . min-1, 30 min.), 1 mM of inosine protected the heart against ATP degradation, while 100 microM of inosine was inefficacious. In the presence of 1 mM of inosine on reperfusion the adenine nucleotide content of the heart was similar to that observed in the absence of the nucleoside. The incorporation of [14C]inosine into adenine nucleotides was, in this last condition, below the value measured before ischaemia. Inosine administration was effective in protecting the heart against ischaemic breakdown of glycogen and favoured postischaemic restoration of glycogen stores.

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In this work, we compared the electrophysiological and metabolic parameters of a volume overload model of cardiac hypertrophy (aorto-caval fistula) with those of two other models of hypertrophy (aortic stenosis and isoproterenol pretreatment). In these last models, a prolongation of action potential and a decrease of myocardial ATP content are observed. However, these alterations are not shown in the aorto-caval fistulated animals while their heart are well hypertrophied. The amplitude increase of phase 3 AP seemed to be a common factor of these models of cardiac hypertrophy.

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