摘要:

To establish a murine model that may allow for definition of the precise role of phospholamban in myocardial contractility through selective perturbations in the phospholamban gene, we initiated studies on the role of phospholamban in the murine heart. Intact beating hearts were perfused in the absence or presence of isoproterenol, and quantitative measurements of cardiac performance were obtained. Isoproterenol stimulation was associated with increases in the affinity of the sarcoplasmic reticulum Ca2+pump for Ca2+that were due to phospholamban phosphorylation. To assess the regulation of phospholamban gene expression during murine development, Northern blot and polymerase chain reaction analyses were used. Phospholamban mRNA was first detected in murine embryos on the ninth day of development (the time when the cardiac tube begins to contract). In murine embryoid bodies, which have been shown to recapitulate several aspects of cardiogenesis, phospholamban mRNA was detected on the seventh day (the time when spontaneous contractions are first observed). Only those embryoid bodies that exhibited contractions expressed phospholamban transcripts, and these were accompanied by expression of the protein, as revealed by immunofluorescence microscopy. Sequence analysis of the cDNA encoding phospholamban in embryoid bodies indicated complete homology to that in adult hearts. The deduced amino acid sequence of murine phospholamban was identical to rabbit cardiac phospholamban but different from dog cardiac and human cardiac phospholamban by one amino acid. These data suggest that phospholamban, the regulator of the Ca2+. ATPase in cardiac sarcoplasmic reticulum, is present very early in murine cardiogenesis in utero and in vitro, and this may constitute an important determinant for proper development of myocardial contractility.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Changes in cardiac myosin heavy chain (MHC) gene expression and isozyme transitions have been shown to be caused by developmental changes, hemodynamic overload, or the activity of various hormones. In this study, to examine whether caffeine, which has teratogenic effects on the fetal cardiovascular system, causes the distribution of cardiac MHC phenotype and, if so, to evaluate the mechanisms of the distribution of cardiac MHC phenotype by caffeine, we examined the effects of caffeine, theophylline, and cAMP on the cardiac MHC isoform transitions at the gene and protein levels using hypothyroid adult rats. Furthermore, we examined the expression of α- and β-MHC gene in cardiac muscles of fetuses whose dams had received caffeine. The results showed that caffeine, theophylline, and cAMP caused accumulations of α-MHC mRNA and MHC isozyme V1. Furthermore, in the fetal hearts, it was recognized that caffeine induced an accumulation of α-MHC gene expression, as was also seen in the dams. However, this effect of caffeine on the heart was stronger in the fetus than in the dam. Intracellular cAMP concentration was increased by the administration of caffeine, theophylline, or cAMP, and the levels showed a positive correlation with those of α-MHC mRNA. These results suggest that the induction of α-MHC mRNA expression by the administration of caffeine may be induced by an increase in intracellular cAMP concentration.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

To investigate the cellular mechanism for the synthesis and secretion of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), we examined the effects of vasoactive agents on the secretion rates and gene expression of ANP and BNP in cultured rat atrial cells. Endothelin (10−7M, +61%), 12-O-tetrade-canoylphorbol 13-acetate (TPA, 10−6M, +62%), the calcium ionophore A23187 (10−6M, +95%), and Bay K 8644 (10−6M, +34%) (p<0.05 each) all increased the secretion of ANP into the culture media in a dose-dependent fashion. On the other hand, endothelin (10−7M, +57%) and TPA (10−6M, +55%) (p<0.01 each) increased the secretion of BNP in a dose-dependent manner, whereas A23187 (10−6M, −45%,p<0.001) suppressed the secretion of BNP in a dose-dependent manner, and Bay K 8644 caused no significant effects on BNP secretion. The molecular forms of intracellular ANP were exclusively γ-ANP, whereas those of BNP were γ-BNP and its carboxy terminal 45-amino-acid peptide, BNP-45. The ratio of media to cell contents was much higher in BNP than in ANP. Northern blot analysis revealed that both ANP mRNA and BNP mRNA levels were significantly increased by 10−7M endothelin (ANP mRNA, +52%; BNP mRNA, +36%;p<0.05 each) and 5×10−5M 1-oleoyl-2-acetylglycerol (ANP mRNA, +296%; BNP mRNA, +133%;p<0.01 each) but not by 10−6M A23187. Thus, the secretion of ANP is stimulated by both the elevation of [Ca2+]iand the activation of protein kinase C, whereas its synthesis is increased mainly by the activation of protein kinase C. The synthesis and secretion of BNP are augmented by the activation of protein kinase C rather than the elevation of [Ca2+]i. Furthermore, the processing and secretion of ANP and BNP may be regulated in different manners.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

The effect of perfusate [Ca2+] on the function of cardiac sarcoplasmic reticulum (CSR) was assessed by the oxalate-supported Ca2+uptake rate of ventricular homogenates of isolated rat hearts maintained in a modified Langendorif preparation. The total Ca2+pumping activity of the CSR was determined by using 20 FM ruthenium red or 625 μM ryanodine to close the CSR Ca2+release channel. The homogenate Ca2+uptake rate in the absence of ruthenium red or ryanodine decreased progressively with increasing perfusate [Ca2+] (25.7±1.2, 21.4±1.5, 17.2±1.1, and 163±1.2 [mean±SEMI nmol Ca2+· min−1· mg−1for hearts perfused for 5 minutes with 0.2, 1.4, 2.8, and 5.6 mM Ca2+, respectively;p=0.0001;n=8). This depression was not observed when Ca2+uptake was assayed in the presence of ryanodine or ruthenium red. Since the Ca2+uptake in the presence of ryanodine or ruthenium red is determined by the Ca2+-ATPase, this result suggests that perfusion with varying [Ca2+] did not affect the Ca2+-ATPase. The observed decrease in Ca2+uptake in the absence of ryanodine or ruthenium red is caused by an increased efflux through the ryanodine-sensitive Ca2+ release channel. When hearts perfused for 5 minutes with 0.2 or 5.6 mM Ca2+were reperfused for 10 minutes with 1.4 mM Ca2+, homogenate Ca2+uptake rates were restored to near control levels. These effects of perfusate Ca2+were not direct effects, because changes in the [Ca2+] of the homogenization medium did not alter the homogenate Ca2+uptake activity in either the presence or absence of ryanodine. The homogenate Ca2+uptake rates were unaffected by prior active loading of the CSR with Ca2+. These results suggest a regulatory role of perfusate Ca2+in increasing the open state of the ryanodine-sensitive Ca2+release channel that is distinct from the beat-to-beat regulation of Ca2+release from the CSR by Ca2+(Ca2+-induced Ca2+release).

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Intracerebroventricularly infused hypertonic saline elicits sympathoexcitatory and pressor effects. To clarify the mechanisms mediating these effects, we evaluated blood pressure (BP), heart rate (HR), and renal sympathetic nerve activity (RSNA) responses to intracerebroventricular administration of 0.3 M NaCl, ouabain, and rat hypothalamic and pituitary extracts containing ouabain-like activity (OLA) in conscious Wistar rats, before and after intracerebroventricular preinjection of digoxin-specific antibody Fab (DAF) fragments. To exclude modulatory effects of arginine vasopressin (AVP), treatment with DAF fragments was in all experiments preceded by intravenous injection of the AVP antagonist [β-mercapto-β,β-cyclopentamethylenepropionyl1,o-Me-Tyr2,Arg8]AVP. After AVP antagonist pretreatment, 0.3 M NaCl i.c.v. at 3.8 μl/min for 10 minutes caused simultaneous increases in BP, RSNA, and HR. After AVP antagonist pretreatment, intracerebroventricular injections of 0.3 and 1.0 μ/l μl ouabain or the OLA equivalent to 1 μg ouabain/2 μl elicited similar significant increases in BP, HR, and RSNA. After pretreatment with AVP antagonist and DAF fragments (66 μg/4 μl i.c.v.), BP, HR, and RSNA responses to 0.3 M NaCl, ouabain, and OLA were all significantly diminished. In contrast, combined AVP blockade and DAF fragments did not affect the BP response to intracerebroventricular angiotensin II, the BP, HR, and RSNA responses to intracerebroventricular carbachol and to air stress, or the HR and RSNA responses to intravenous sodium nitroprusside. Intracerebroventricularly injected γ-globulins (66 μg/4 μl) did not affect the responses to 0.3 M NaCl, ouabain, or OLA. These data demonstrate that the effects of intracerebroventricularly infused hypertonic saline share, at least in part, a common mechanism with the effects of intracerebroventricular ouabain/OLA; i.e., brain OLA appears to be involved in the sympathoexcitatory and pressor effects of intracerebroventricular hypertonic saline.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

We studied product inhibition of the actin-activated ATPase of myosin subfragment-1 (S-1) from the three types of muscle tissue: skeletal, cardiac, and smooth. Increasing levels of [MgADP] in the 0–1-mM range caused significant inhibition of the actin-activated MgATPase activity of cardiac and gizzard but not skeletal muscle S-1. When total nucleotide concentration ([ATP] + [ADP]) was kept constant at 1 mM, ATPase activity was inhibited by 50% at an ADP/ATP ratio of 6:1 for cardiac S-1 and 3:1 for gizzard S-1. For skeletal S-1, however, even a 19:1 ratio did not cause 50% inhibition of ATPase activity. The observed effect was not due to changes in pH or inorganic phosphate concentration, nor could it be explained by substrate (ATP) depletion. In the absence of actin, ADP had little or no inhibitory effect on the ATPase activity of S-1, and these observations imply that ADP is competing directly for the ATP binding site of the actin-Si complexes of cardiac and smooth muscle S-1. ADP has previously been shown to be a weak competitive inhibitor of the ATPase activity in skeletal muscle. The current data imply that ADP is a very effective competitive inhibitor for the actin-activated ATPase activity of cardiac and gizzard S-1 and, therefore, that ADP may be a physiologically important modulator of contractile activity in cardiac and smooth muscle.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

The purpose of this study was to investigate whether sympathetic stimulation modulated the rise in extracellular K+concentration ([K+]0) evoked by acute myocardial ischemia. In 35 α-chloralose-anesthetized dogs, we measured changes in [K+]Oduring acute myocardial ischemia in the presence and absence of sympathetic stimulation or norepinephrine infusion. A series of four 5 -minute occlusions of the distal left anterior descending coronary artery (LAD) was completed in 18 dogs. Thirty minutes of reperfusion separated each LAD occlusion. Four to five K+-sensitive electrodes were inserted into the left ventricular midmyocardium that was perfused by the distal LAD. Lead II of the electrocardiogram, arterial pressure, and [K+]0were recorded, and the right atrium was paced at a constant cycle length. The first, second, and fourth LAD occlusions were done in the absence of sympathetic stimulation or norepinephrine infusion. The changes in [K+]Oevoked by the first LAD occlusion differed (p<0.05) from those elicited by the second and fourth occlusions. However, the changes in [K+]0during the second and fourth LAD occlusions were similar (p>0.2) and served as controls for the responses obtained during the third occlusion. Two minutes before the third LAD occlusion, sympathetic stimulation (4 Hz) or norepinephrine infusion (0.25–0.5 μg/kg per minute i.v.) was begun and was continued until 2 minutes after reperfusion. We found that sympathetic stimulation and norepinephrine infusion increased (p<0.05) myocardial blood flow in both normal and ischemic tissue. The mean response recorded by 23 K+-sensitive electrodes in 11 dogs showed that sympathetic stimulation increased (p<0.001) the [K+]0at 1,2,3,4, and 5 minutes after the onset of LAD occlusion compared with the second and fourth occlusions. In contrast, the mean response recorded by 20 K+-sensitive electrodes in seven dogs showed that norepinephrine infusion reduced (p<0.02) the [K+]0at 4 and 5 minutes after the onset of LAD occlusion. These data show that sympathetic stimulation increased the [K+]0evoked by acute myocardial ischemia, an effect that was not mimicked by the intravenous administration of norepinephrine.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Stimulation of thymidine incorporation by basic fibroblast growth factor or epidermal growth factor treatment of cultured quiescent smooth muscle cells (rat and human) was attenuated by the concomitant treatment with interleukin-1β in the presence of indomethacin. Platelet-derived growth factor-AB and -BB-induced thymidine incorporation was not inhibited by the presence of the cytokine under similar experimental conditions. Elevation of nitrite levels in the conditioned medium of cultures exposed to interleukin-1β correlated with the inhibition of thymidine incorporation. Platelet-derived growth factor-AB and -BB inhibited the production of nitric oxide (measured as nitrite levels in conditioned medium) by cells treated simultaneously with interleukin-1β and growth factor. However, platelet-derived growth factor-AA neither affected nitrite production nor thymidine incorporation by smooth muscle cells. Levels of cytokine-stimulated nitrite production by smooth muscle cells were increased synergistically by the presence of fibroblast growth factors or epidermal growth factor. The inhibition of thymidine incorporation and concomitant elevation of nitrite production was abolished in the presence of nitro-l-arginine. Cultures maintained in the presence of low levels of the cytokine for 9 days were growth-inhibited, and this was reversed when culture medium was supplemented with nitro-l-arginine. The treatment of smooth muscle cells, which were grown in coculture inserts with the cytokine to induce nitric oxide production, before their combination with other quiescent layers of cells resulted in the inhibition of thymidine incorporation by this second layer of cells regardless of the growth factor used for stimulation. Nitric oxide may act as an endogenous inhibitor of smooth muscle cell proliferation in the vessel wall, and impairment of its production may be one action of potent vascular mitogens such as platelet-derived growth factor.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Adenosine has been shown to protect the ischemic and reperfused myocardium. To examine whether the protective effect of the nucleoside is mediated by modulation of oxidative stress, isolated rat hearts were perfused for 30 minutes with 100 μM H2O2or an exogenous free radical-generating system consisting of purine (3.06 mM) and xanthine oxidase (10 units/l) in the presence or absence of drugs acting on adenosine A1or A2receptors. H2O2alone produced a greater than 90% loss in contractility concomitant with a threefold elevation in resting tension, although these effects occurred in the absence of ultrastructural damage. Two A1receptor agonistsN6-cyclopentyladenosine (CPA, 1 μM) andR(−)-N6-(2-phenylisopropyl)adenosine (R-PIA, 1 μM) significantly attenuated the cardiodepressant effects of H2O2and depressed the elevation in resting tension; however, only the effect of CPA was found to be significant with regard to the latter parameter. A similar concentration ofS(+)-N6-(2-phenylisopropyl) adenosine (S-PIA), a markedly less potent A1receptor agonist, was found to be without beneficial effect. However, a significant protective effect against both the reduction in contractility and the elevation in resting tension was seen with a 10-fold elevation in the concentration of S-PIA (10 μM). The protective effects on functional parameters were associated with preservation of high-energy phosphate and adenine nucleotide contents after 30 minutes of H2O2treatment. The salutary effects of all drugs were reversed in the presence of the A1receptor antagonist 1,3-dipropyl-8-cyclopentylxanthine (0.5 μM). An A2receptor agonist 2-[p-(carboxyethyl)phenethylamino]-5'-N-ethylcarboxamidoadenosine, termed CGS 21680 (1 μM), failed to alter the cardiac response to H2O2with regard to all parameters studied. Neither a 50% reduction in external CaCl2concentration nor treatment with 10 μM dl-propranolol exerted salutary effects against H2O2-induced dysfunction. None of the A1receptor agonists modulated the response to purine plus xanthine oxidase. Our results demonstrate a selective protective effect of adenosine A1receptor activation against the cardiac toxicity of H2O2and provide, at least in part, a basis for the cardioprotective actions of adenosine and its analogues.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Ventricular fibrillation (VF) is known to produce alterations in myocardial energetics, but the mechanism of these changes remains unclear. To investigate energy metabolism during VF, phosphorus nuclear magnetic resonance spectroscopy and magnetization transfer were applied to isolated perfused ferret hearts. VF was induced either by perfusion with digitalis (strophanthidin, 30 μM) or by high-frequency electrical stimulation. We measured the flux in two critical reactions: from inorganic phosphate (Pi) to ATP (ATP synthesis rate) and from phosphocreatine (PCr) to ATP (energy transfer capacity). During digitalis-induced VF, energy-related phosphates showed changes similar to those during hypoxia: myocardial [Pi] increased and [PCr] decreased. Concomitantly, the ATP synthesis rate increased to levels about threefold higher than control, whereas oxygen consumption increased by only 16%. The ATP synthesis rate exhibited a strong negative correlation with left ventricular pressure during VF (r= −0.95,n=5,p<0.02), whereas oxygen consumption did not (r=0.19,p>0.05). On the other hand, energy transfer capacity catalyzed by creatine kinase was significantly smaller during VF than in the control condition but still higher than the simultaneous ATP synthesis rate. In contrast to the marked energetic deterioration during VF induced by digitalis, electrically induced VF led to only a small increase in [Pi] and a small decrease in [PCr], and there were no significant changes in the ATP synthesis rate, energy transfer capacity, or O2consumption. These results indicate that the rundown in energy metabolism during VF induced by digitalis was mainly attributable to a limitation of energy production through oxidative phosphorylation as well as to a marked increase in energy consumption. In contrast, myocardial energy generation remained unimpaired during VF induced by electrical stimulation. Intracellular calcium overload is more severe during VF induced by digitalis than during electrically induced VF (Circ Res1991;68:1378–1389); severe calcium overload would be expected to compromise the capacity for energy generation by mitochondria. Thus, we propose that known differences in cellular calcium loading underlie the discrepant energetic patterns of the two types of VF.

ISSN:0009-7330    

出版商:OVID    

年代:1992

数据来源: OVID