RESUMO
Genetic modification in the mouse heart has provided new and important insights into many aspects of ATP synthesis, supply and utilization and how this changes in the failing heart. Here, three topics based on recent literature will be reviewed: direct manipulation of precursor pool for creatine; the path linking creatine, purine nucleotides and metabolic remodeling; and long-term reprogramming of ATP synthesis pathways. These examples illustrate the value of deleting, over-expressing and mutating specific proteins in our quest for understanding the energetics of the failing heart.
Assuntos
Insuficiência Cardíaca/genética , Insuficiência Cardíaca/metabolismo , 5'-Nucleotidase/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Creatina Quinase/metabolismo , Regulação da Expressão Gênica , Camundongos , Contração Miocárdica , Miocárdio/metabolismo , Purinas/metabolismoRESUMO
Heme oxygenase (HO)-1 degrades the pro-oxidant heme and generates carbon monoxide and antioxidant bilirubin. We have previously shown that in response to hypoxia, HO-1-null mice develop infarcts in the right ventricle of their hearts and that their cardiomyocytes are damaged by oxidative stress. To test whether HO-1 protects against oxidative injury in the heart, we generated cardiac-specific transgenic mice overexpressing different levels of HO-1. By use of a Langendorff preparation, hearts from transgenic mice showed improved recovery of contractile performance during reperfusion after ischemia in an HO-1 dose-dependent manner. In vivo, myocardial ischemia and reperfusion experiments showed that infarct size was only 14.7% of the area at risk in transgenic mice compared with 56.5% in wild-type mice. Hearts from these transgenic animals had reduced inflammatory cell infiltration and oxidative damage. Our data demonstrate that overexpression of HO-1 in the cardiomyocyte protects against ischemia and reperfusion injury, thus improving the recovery of cardiac function.
Assuntos
Heme Oxigenase (Desciclizante)/genética , Isquemia Miocárdica/prevenção & controle , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/enzimologia , Animais , Regulação Enzimológica da Expressão Gênica/fisiologia , Genótipo , Coração/fisiopatologia , Heme Oxigenase (Desciclizante)/metabolismo , Heme Oxigenase-1 , Humanos , Proteínas de Membrana , Camundongos , Camundongos Transgênicos , Infarto do Miocárdio/enzimologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/prevenção & controle , Isquemia Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/patologiaRESUMO
Endogenous nitric oxide (eNO) modulates tissue respiration. To test whether eNO modulates myocardial O2 consumption (MVO2), ATP synthesis, and metabolic efficiency, we used isolated isovolumic guinea pig hearts perfused at a constant flow. N(omega)-nitro-L-arginine (L-NNA; 5 x 10(-5) mol/l) was used to inhibit eNO production. MVO2 was measured at different levels of cardiac work, estimated as the rate-pressure product (RPP). ATP content and synthesis rate were determined using (31)P NMR and magnetization transfer during high cardiac work. L-NNA increased coronary vascular resistance (19 +/- 3%, P < 0.05) and MVO2 (12 +/- 3%, P < 0.05) without an increase in the RPP. In contrast, vehicle infusion resulted in insignificant changes in coronary vascular resistance (3 +/- 2%, P > 0.05) and MVO2 (-2 +/- 1%, P > 0.05). Compared with vehicle, L-NNA caused a higher MVO2 both during KCl arrest (L-NNA 5.6 +/- 0.5 vs. vehicle 3.0 +/- 0.4 micromol x min(-1) x mg x dry wt(-1), P < 0.05) and during increased cardiac work elicited by elevating perfusate Ca2+, indicating an upward shift in the relationship between contractile performance (measured as RPP) and MVO2. However, neither ATP contents nor ATP synthesis rates were different in the two groups during high cardiac work. Thus, because inhibition of eNO production by L-NNA increased MVO2 without a change in the ATP synthesis rate, these data suggest that eNO increases myocardial metabolic efficiency by reducing MVO2 in the heart.
Assuntos
Trifosfato de Adenosina/biossíntese , Coração/fisiologia , Óxido Nítrico/fisiologia , Oxigênio/fisiologia , Animais , Inibidores Enzimáticos/farmacologia , Cobaias , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/fisiologia , Óxido Nítrico Sintase Tipo III , Nitroarginina/farmacologia , Consumo de Oxigênio/fisiologiaRESUMO
In situations such as severe low-flow ischemia, where myocardial work output is low and dependence on anaerobic glycolysis is high, increasing the myocardial supply of glucose and insulin is cardioprotective. Our goal was to determine whether this strategy of "metabolic support" would also be cardioprotective in the moderately hypoperfused heart receiving inotropic stimulation, i.e. when myocardial work was near normal, and reliance on anaerobic glycolysis was minimal. Isovolumic left ventricular performance and cardiac energetics (31P-NMR spectroscopy) were measured in 20 isolated rat hearts perfused with red blood cell containing perfusate (hematocrit 40%) with either normal (5 m M, 15 microU/ml) or increased (19.5 m M, 250 microU/ml) glucose and insulin in addition to normal levels of lactate and free fatty acids. Lowering global coronary flow to 30% of normal decreased left ventricle developed pressure by 50%. Administering dobutamine for 40 min restored developed pressure to 95+/-13% of baseline but caused diastolic pressure to increase by 23+/-6 mmHg and [ATP] to decrease by 44+/-6%. Glucose and insulin prevented the increase in end-diastolic pressure, and [ATP] fell by only 14+/-3%. Despite these improvements in cardiac energetics and diastolic function, left ventricle developed pressure was not improved by increased glucose and insulin during, or after the hypoperfusion. We conclude that inotropic support of the hypoperfused heart can cause new diastolic dysfunction, but that this diastolic dysfunction can be eliminated by preserving myocardial high-energy phosphates with increased glucose and insulin.
Assuntos
Coração/fisiologia , Isquemia Miocárdica , Miocárdio/metabolismo , Perfusão , Função Ventricular Esquerda/fisiologia , Trifosfato de Adenosina/metabolismo , Animais , Velocidade do Fluxo Sanguíneo/efeitos dos fármacos , Pressão Sanguínea/efeitos dos fármacos , Dobutamina/farmacologia , Eletrofisiologia , Ácidos Graxos/farmacologia , Glucose/farmacologia , Concentração de Íons de Hidrogênio , Insulina/farmacologia , Ácido Láctico/farmacologia , Espectroscopia de Ressonância Magnética , Masculino , Ratos , Ratos Sprague-Dawley , Termodinâmica , Fatores de Tempo , Função Ventricular Esquerda/efeitos dos fármacosRESUMO
Type 2 iodothyronine deiodinase (D(2)) catalyzes intracellular 3, 5, 3' triiodothyronine (T(3)) production from thyroxine (T(4)), and its messenger RNA mRNA is highly expressed in human, but not rodent, myocardium. The goal of this study was to identify the effects of D(2) expression in the mouse myocardium on cardiac function and gene expression. We prepared transgenic (TG) mice in which human D(2) expression was driven by the alpha-MHC promoter. Despite high myocardial D(2) activity, myocardial T(3) was, at most, minimally increased in TG myocardium. Although, plasma T(3) and T(4), growth rate as well as the heart weight was not affected by TG expression, there was a significant increase in heart rate of the isolated perfused hearts, from 284 +/-12 to 350 +/- 7 beats/min. This was accompanied by an increase in pacemaker channel (HCN2) but not alpha-MHC or SERCA II messenger RNA levels. Biochemical studies and (31)P-NMR spectroscopy showed significantly lower levels of phosphocreatine and creatine in TG hearts. These results suggest that even mild chronic myocardial thyrotoxicosis, such as may occur in human hyperthyroidism, can cause tachycardia and associated changes in high energy phosphate compounds independent of an increase in SERCA II and alpha-MHC.
Assuntos
Coração/fisiopatologia , Iodeto Peroxidase/metabolismo , Proteínas Musculares , Tireotoxicose/genética , Tireotoxicose/fisiopatologia , Nucleotídeos de Adenina/metabolismo , Animais , ATPases Transportadoras de Cálcio/genética , Creatina Quinase/metabolismo , Metabolismo Energético , Coração/fisiologia , Humanos , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização , Iodeto Peroxidase/genética , Canais Iônicos/genética , Isoenzimas/genética , Isoenzimas/metabolismo , Camundongos , Camundongos Transgênicos , Miocárdio/metabolismo , Cadeias Pesadas de Miosina/genética , Tamanho do Órgão , Canais de Potássio , Regiões Promotoras Genéticas , RNA Mensageiro/genética , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático , Tiroxina/sangue , Tiroxina/metabolismo , Transcrição Gênica , Tri-Iodotironina/sangueRESUMO
To understand why the adult human heart expresses three isoforms of the sodium pump, we generated transgenic mice (TGM) with 2.3- to 5. 5-fold overexpression of the human alpha(3)-isoform of Na-K-ATPase in the heart. Hearts from the TGM had increased maximal Na-K-ATPase activity and ouabain affinity compared with control hearts, even though the density of Na-K-ATPase pump sites (of all isoforms) was similar to that of control mice. In perfused hearts, contractility both at baseline and in the presence of ouabain tended to be greater in TGM than in controls. Surface electrocardiograms in anesthetized TGM had a steeper dependence of Q-T on sinus cycle length, and Q-T intervals measured during atrial pacing were significantly longer in TGM. Q-T dispersion during sinus rhythm also tended to be longer in TGM. Thus TGM overexpressing human alpha(3)-isoform have several of the phenotypical features of human long Q-T syndrome, despite the absence of previously described mutations in Na(+) or K(+) channels.
Assuntos
Síndrome do QT Longo/enzimologia , Síndrome do QT Longo/genética , Miocárdio/enzimologia , ATPase Trocadora de Sódio-Potássio/biossíntese , Animais , Estimulação Cardíaca Artificial , Células Cultivadas , Modelos Animais de Doenças , Eletrocardiografia/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Humanos , Técnicas In Vitro , Isoenzimas/biossíntese , Isoenzimas/genética , Modelos Lineares , Camundongos , Camundongos Transgênicos , Contração Miocárdica/efeitos dos fármacos , Contração Miocárdica/genética , Ouabaína/farmacologia , Fenótipo , ATPase Trocadora de Sódio-Potássio/antagonistas & inibidores , ATPase Trocadora de Sódio-Potássio/genética , Taquicardia Ventricular/etiologiaRESUMO
Creatine kinase (CK) exists as a family of isoenzymes in excitable tissue. We studied isolated perfused hearts from mice lacking genes for either the main muscle isoform of CK (M-CK) or both M-CK and the main mitochondrial isoform (Mt-CK) to determine 1) the biological significance of CK isoenzyme shifts, 2) the necessity of maintaining a high CK reaction rate, and 3) the role of CK isoenzymes in establishing the thermodynamics of ATP hydrolysis. (31)P NMR was used to measure [ATP], [PCr], [P(i)], [ADP], pH, as well as the unidirectional reaction rate of PCr--> [gamma-P]ATP. Developmental changes in the main fetal isoform of CK (BB-CK) were unaffected by loss of other CK isoenzymes. In hearts lacking both M- and Mt-CK, the rate of ATP synthesis from PCr was only 9% of the rate of ATP synthesis from oxidative phosphorylation demonstrating a lack of any high energy phosphate shuttle. We also found that the intrinsic activities of the BB-CK and the MM-CK isoenzymes were equivalent. Finally, combined loss of M- and Mt-CK (but not loss of only M-CK) prevented the amount of free energy released from ATP hydrolysis from increasing when pyruvate was provided as a substrate for oxidative phosphorylation.
Assuntos
Creatina Quinase/química , Creatina Quinase/genética , Coração/embriologia , Miocárdio/enzimologia , Trifosfato de Adenosina/metabolismo , Fatores Etários , Animais , Animais Recém-Nascidos , Cromatografia Líquida de Alta Pressão , Creatina Quinase/fisiologia , Glucose/metabolismo , Concentração de Íons de Hidrogênio , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/fisiologia , Cinética , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos Mutantes , Oxigênio/metabolismo , Perfusão , Ácido Pirúvico/metabolismo , Termodinâmica , Fatores de TempoRESUMO
Comparisons of myocardium remodeled by the 2 most common causes of left ventricular hypertrophy (LVH), hypertension and aortic constriction, are limited. We hypothesized that important differences may exist in the myocardium of hearts with these 2 origins of "pressure overload" LVH. Accordingly, we studied isolated hearts from 3 groups of Dahl salt-sensitive rats, controls, and hearts with matched amounts of LVH secondary to either hypertension or aortic constriction. Isovolumic LV function and myocardial energetics ((31)P nuclear magnetic resonance spectroscopy) were measured as coronary flow was lowered to 16% of baseline for 48 minutes. During this low-flow ischemia, isovolumic end-diastolic pressure, a measure of LV stiffness, increased to 52+/-4 mm Hg in controls and 51+/-6 mm Hg in aortic banded hearts but to only 35+/-5 mm Hg in hearts with hypertensive LVH. In all hearts, the P(i) resonance in the (31)P nuclear magnetic resonance spectrum, whose position indicates myocardial pH, split into 2 peaks during low-flow ischemia, which indicates distinct regions of pH 6.9 (moderate acidosis) and pH 6.2 (severe acidosis). Concentrations of ATP, PCr, P(i), and H(+) of the moderately acidotic region were not different among groups. However, the size of the severely acidotic region was smallest in the hypertensive LVH hearts, and in all 3 groups, the size of this region correlated (r(2)=0.65 to 0.80) with the degree of LV stiffening. We conclude that in Dahl rats, LVH secondary to hypertension protects against ischemia-induced diastolic dysfunction by minimizing the size of the region of severe acidosis.
Assuntos
Estenose da Valva Aórtica/fisiopatologia , Pressão Sanguínea , Hipertensão/fisiopatologia , Hipertrofia Ventricular Esquerda/fisiopatologia , Animais , Estenose da Valva Aórtica/patologia , Hipertensão/patologia , Hipertrofia Ventricular Esquerda/patologia , Contração Miocárdica , RatosRESUMO
BACKGROUND: Our goals were to (1) simulate the degree of low-flow ischemia and mixed anaerobic and aerobic metabolism of an acutely infarcting region; (2) define changes in anaerobic glycolysis, oxidative phosphorylation, and the creatine kinase (CK) reaction velocity; and (3) determine whether and how increased glycolytic substrate alters the energetic profile, function, and recovery of the ischemic myocardium in the isolated blood-perfused rat heart. METHODS AND RESULTS: Hearts had 60 minutes of low-flow ischemia (10% of baseline coronary flow) and 30 minutes of reperfusion with either control or high glucose and insulin (G+I) as substrate. In controls, during ischemia, rate-pressure product and oxygen consumption decreased by 84%. CK velocity decreased by 64%; ATP and phosphocreatine (PCr) concentrations decreased by 51% and 63%, respectively; inorganic phosphate (P(i)) concentration increased by 300%; and free [ADP] did not increase. During ischemia, relative to controls, the G+I group had similar CK velocity, oxygen consumption, and tissue acidosis but increased glycolysis, higher [ATP] and [PCr], and lower [P(i)] and therefore had a greater free energy yield from ATP hydrolysis. Ischemic systolic and diastolic function and postischemic recovery were better. CONCLUSIONS: During low-flow ischemia simulating an acute myocardial infarction region, oxidative phosphorylation accounted for 90% of ATP synthesis. The CK velocity fell by 66%, and CK did not completely use available PCr to slow ATP depletion. G+I, by increasing glycolysis, slowed ATP depletion, maintained lower [P(i)], and maintained a higher free energy from ATP hydrolysis. This improved energetic profile resulted in better systolic and diastolic function during ischemia and reperfusion. These results support the clinical use of G+I in acute MI.
Assuntos
Trifosfato de Adenosina/biossíntese , Isquemia Miocárdica/metabolismo , Miocárdio/metabolismo , Animais , Creatina Quinase/metabolismo , Modelos Animais de Doenças , Glucose/metabolismo , Glucose/fisiologia , Hemodinâmica , Insulina/fisiologia , Espectroscopia de Ressonância Magnética , Masculino , Infarto do Miocárdio/metabolismo , Isquemia Miocárdica/fisiopatologia , Fosforilação Oxidativa , Consumo de Oxigênio , Fosfocreatina/metabolismo , Ratos , Ratos WistarRESUMO
Glucose enters the heart via GLUT1 and GLUT4 glucose transporters. GLUT4-deficient mice develop striking cardiac hypertrophy and die prematurely. Whether their cardiac changes are caused primarily by GLUT4 deficiency in cardiomyocytes or by metabolic changes resulting from the absence of GLUT4 in skeletal muscle and adipose tissue is unclear. To determine the role of GLUT4 in the heart we used cre-loxP recombination to generate G4H(-/-) mice in which GLUT4 expression is abolished in the heart but is present in skeletal muscle and adipose tissue. Life span and serum concentrations of insulin, glucose, FFAs, lactate, and beta-hydroxybutyrate were normal. Basal cardiac glucose transport and GLUT1 expression were both increased approximately 3-fold in G4H(-/-) mice, but insulin-stimulated glucose uptake was abolished. G4H(-/-) mice develop modest cardiac hypertrophy associated with increased myocyte size and induction of atrial natriuretic and brain natriuretic peptide gene expression in the ventricles. Myocardial fibrosis did not occur. Basal and isoproterenol-stimulated isovolumic contractile performance was preserved. Thus, selective ablation of GLUT4 in the heart initiates a series of events that results in compensated cardiac hypertrophy.
Assuntos
Cardiomegalia/etiologia , Proteínas de Transporte de Monossacarídeos/fisiologia , Proteínas Musculares , Contração Miocárdica , Animais , Fator Natriurético Atrial/genética , Cardiomegalia/fisiopatologia , Feminino , Glucose/metabolismo , Transportador de Glucose Tipo 1 , Transportador de Glucose Tipo 4 , Masculino , Camundongos , Camundongos Transgênicos , Proteínas de Transporte de Monossacarídeos/genética , Miocárdio/metabolismo , Peptídeo Natriurético Encefálico/genética , Tamanho do ÓrgãoRESUMO
Activation of protein kinase C (PKC) protects the heart from ischemic injury; however, its mechanism of action is unknown, in part because no model for chronic activation of PKC has been available. To test whether chronic, mild elevation of PKC activity in adult mouse hearts results in myocardial protection during ischemia or reperfusion, hearts isolated from transgenic mice expressing a low level of activated PKCbeta throughout adulthood (beta-Tx) were compared with control hearts before ischemia, during 12 or 28 min of no-flow ischemia, and during reperfusion. Left-ventricular-developed pressure in isolated isovolumic hearts, normalized to heart weight, was similar in the two groups at baseline. However, recovery of contractile function was markedly improved in beta-Tx hearts after either 12 (97 +/- 3% vs. 69 +/- 4%) or 28 min of ischemia (76 +/- 8% vs. 48 +/- 3%). Chelerythrine, a PKC inhibitor, abolished the difference between the two groups, indicating that the beneficial effect was PKC-mediated. (31)P NMR spectroscopy was used to test whether modification of intracellular pH and/or preservation of high-energy phosphate levels during ischemia contributed to the cardioprotection in beta-Tx hearts. No difference in intracellular pH or high-energy phosphate levels was found between the beta-Tx and control hearts at baseline or during ischemia. Thus, long-term modest increase in PKC activity in adult mouse hearts did not alter baseline function but did lead to improved postischemic recovery. Furthermore, our results suggest that mechanisms other than reduced acidification and preservation of high-energy phosphate levels during ischemia contribute to the improved recovery.
Assuntos
Isquemia Miocárdica/fisiopatologia , Miocárdio/enzimologia , Proteína Quinase C/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Concentração de Íons de Hidrogênio , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos Transgênicos , Isquemia Miocárdica/enzimologia , Isquemia Miocárdica/metabolismo , Miocárdio/metabolismo , Fosfocreatina/metabolismoRESUMO
BACKGROUND: Whether myocardial ATP content falls in heart failure is a long-standing and controversial issue. The mechanism(s) to explain any decrease in ATP content during heart failure have not been identified. METHODS AND RESULTS: Cardiac dysfunction, heart failure, and a prolonged steady state of heart failure were induced by chronic right ventricular pacing for 1 to 2 weeks, 3 to 4 weeks, and 7 to 9 weeks in dogs. Cardiac function and myocardial O(2) consumption (Mf1.gif" BORDER="0">O(2)) were measured with the dogs in the conscious state. ATP, total purine, and creatine were measured in biopsy specimens obtained at each stage. ATP and the total purine pool progressively fell at rates of 0.12 and 0.15 nmol. mg protein(-1). d(-1), despite an increase in Mf1.gif" BORDER="0">O(2). The rate of loss of creatine was 1.06 nmol. mg protein(-1). d(-1), 7 times faster than the depletion of total purine. CONCLUSIONS: (1) ATP contents progressively decreased during heart failure as a result of a loss of the total purine pool. The loss of purines may be due to inhibition of de novo purine synthesis. (2) Loss of creatine is an early marker of heart failure and may serve as a compensatory mechanism minimizing the reduction of the total purine pool in the failing heart.
Assuntos
Trifosfato de Adenosina/metabolismo , Creatina/metabolismo , Doenças do Cão/fisiopatologia , Cardiopatias/veterinária , Miocárdio/metabolismo , Purinas/metabolismo , Animais , Cães , Coração/fisiologia , Cardiopatias/fisiopatologia , Hemodinâmica , Modelos CardiovascularesRESUMO
Decreasing coronary perfusion causes an immediate decrease in contractile function via unknown mechanisms. It has long been suspected that this contractile dysfunction is caused by ischemia-induced changes in cardiac energetics. Our goal was to determine whether changes in cardiac energetics necessarily precede the contractile dysfunction as one would expect if a causal relationship exists. In 14 isolated rat hearts, we gradually decreased coronary perfusion using a coronary perfusate with a normal hematocrit and normal concentrations of the major metabolic substrates. Using 31P NMR spectroscopy to measure ATP, phosphocreatine (PCr), Pi, and ADP concentrations ([ATP], [PCr], [Pi], [ADP]), pH, and amount of free energy released from ATP hydrolysis (|DeltaGATP|), we found that none of these variables changed significantly until several minutes after systolic pressure had significantly decreased. Even when developed pressure had decreased by over one-third, only very slight changes in [Pi], pH, and |DeltaGATP| had occurred, with no significant changes in [ATP], [PCr], or [ADP]. Additionally, the rate of high-energy phosphate transfer between ATP and PCr did not decrease enough during hypoperfusion to explain the contractile dysfunction. We conclude that nonenergetic factors are the dominant cause of the initial decrease in systolic function when myocardial perfusion is decreased.
Assuntos
Metabolismo Energético/fisiologia , Contração Miocárdica/fisiologia , Isquemia Miocárdica/metabolismo , Miocárdio/enzimologia , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Pressão Sanguínea/fisiologia , Creatina Quinase/metabolismo , Hibernação/fisiologia , Concentração de Íons de Hidrogênio , Espectroscopia de Ressonância Magnética , Magnetismo , Masculino , Técnicas de Cultura de Órgãos , Fosfatos/metabolismo , Fosfocreatina/metabolismo , Isótopos de Fósforo , Ratos , Ratos Sprague-DawleyRESUMO
To determine whether the decreased contractile performance in diabetic hearts is associated with a reduced energy reserve due to decreased creatine kinase (CK) activity, we measured total CK activity (V(max)) in vitro and CK reaction velocity in vivo using(31)P NMR spectroscopy in isolated perfused rat hearts after 4 and 6 weeks of diabetes. After 4 weeks of diabetes, V(max)decreased by 22% with a larger decrease of CK MB than of CK MM and mitochondrial-CK isoenzymes. There was no further decrease in these parameters after 6 weeks of diabetes. Isovolumic contractile performance of 4 and 6 week diabetic hearts, estimated as rate-pressure product under identical perfusion and loading conditions (EDP set at 6-8 mmHg), was only 50% of that of control. ATP, PCr and total creatine concentrations were not different in control and 4 or 6 weeks diabetic rat hearts. After 4 weeks of diabetes, CK reaction velocity decreased by 22%. This was in proportion to the decline of V(max)and therefore predicted by the rate equation for the CK reaction. However, the further decline in the CK reaction velocity after 6 weeks of diabetes (45%) was greater than that predicted from the CK rate equation (17% decrease), and cannot be explained by substrate control of the enzyme. When hearts were inotropically stimulated by increasing perfusate calcium concentration, CK reaction velocity increased slightly (approximately 15%) in both control and diabetic hearts, thereby maintaining a constant ATP concentration. We conclude that in the diabetic myocardium, the CK reaction velocity decreases but does not limit the availability of high-energy phosphates for contraction over the range of workloads studied. We also conclude that a mechanism(s) in addition to substrate control regulates CK reaction velocity in the 6 week diabetic hearts.
Assuntos
Cardiomiopatias/enzimologia , Creatina Quinase/metabolismo , Diabetes Mellitus Experimental/enzimologia , Animais , Cardiomiopatias/diagnóstico por imagem , Cardiomiopatias/fisiopatologia , Diabetes Mellitus Experimental/diagnóstico por imagem , Diabetes Mellitus Experimental/fisiopatologia , Cinética , Espectroscopia de Ressonância Magnética , Masculino , Radiografia , Ratos , Ratos WistarRESUMO
The free energy release from ATP hydrolysis (|DeltaG approximately p|) is decreased by inhibiting the creatine kinase (CK) reaction, which may limit the thermodynamic driving force for the sarcoplasmic reticulum (SR) Ca2+ pumps and thereby cause a decrease in contractile reserve. To determine whether a decrease in |DeltaG approximately p| results in decreased contractile reserve by impairing Ca2+ handling, we measured left ventricular pressure and cytosolic Ca2+concentration ([Ca2+]c; by indo 1 fluorescence) in isolated perfused rat hearts, with >95% inhibition of CK with 90 micromol iodoacetamide. Iodoacetamide did not directly alter SR Ca2+-ATPase activity, baseline left ventricular developed pressure, or baseline [Ca2+]c. When perfusate Ca2+ concentration was increased from 1.2 to 3.3 mM, LV developed pressure increased from 67 +/- 6 to 119 +/- 8 mmHg in control hearts (P < 0.05) but did not significantly increase in CK-inhibited hearts. Similarly, the amplitude of the [Ca2+]c transient increased from 548 +/- 54 to 852 +/- 140 nM in control hearts (P < 0.05) but did not significantly increase in CK-inhibited hearts. We conclude that decreased |DeltaG approximately p| limits intracellular Ca2+ handling and thereby limits contractile reserve.
Assuntos
Cálcio/metabolismo , Contração Miocárdica/fisiologia , Termodinâmica , Animais , ATPases Transportadoras de Cálcio/metabolismo , Creatina Quinase/antagonistas & inibidores , Creatina Quinase/metabolismo , Citosol/metabolismo , Inibidores Enzimáticos/farmacologia , Iodoacetamida/farmacologia , Masculino , Concentração Osmolar , Perfusão , Pressão , Ratos , Ratos Sprague-Dawley , Retículo Sarcoplasmático/efeitos dos fármacos , Retículo Sarcoplasmático/enzimologia , Função Ventricular Esquerda/fisiologiaRESUMO
A growing body of evidence has been accumulated recently suggesting that growth hormone (GH) and insulin-like growth factor-1 (IGF-1) affect cardiac function, but their mechanism(s) of action is unclear. In the present study, GH and IGF-1 were administered to isolated isovolumic aequorin-loaded rat whole hearts and ferret papillary muscles. Although GH had no effect on the indices of cardiac function, IGF-1 increased isovolumic developed pressure by 24% above baseline. The aequorin transients were abbreviated and demonstrated decreased amplitude. The positive inotropic effects of IGF-1 were not associated with increased intracellular Ca2+ availability to the contractile machinery but to a significant increase of myofilament Ca2+ sensitivity. Accordingly, the Ca2+-force relationship obtained under steady-state conditions in tetanized muscle was shifted significantly to the left (EC50, 0.44+/-0.02 versus 0.52+/-0.03 micromol/L with and without IGF-1 in the perfusate, respectively; P<0.05); maximal Ca2+-activated tetanic pressure was increased significantly by 12% (211+/-3 versus 235+/-2 mm Hg in controls and IGF-1-treated hearts, respectively; P<0.01). The positive inotropic actions of IGF-1 were not associated with changes in either pHi or high-energy phosphate content, as assessed by 31P nuclear magnetic resonance spectroscopy, and were blocked by the phosphatidylinositol 3-kinase inhibitor wortmannin. Concomitant administration of IGF binding protein-3 blocked IGF-1-positive inotropic action in ferret papillary muscles. In conclusion, IGF-1 is an endogenous peptide that through a wortmannin-sensitive pathway displays distinct positive inotropic properties by sensitizing the myofilaments to Ca2+ without increasing myocyte [Ca2+]i.