摘要:

&NA;By use of gene targeting and/or transgenesis, it is now possible to make defined changes in genes whose functions underlie mammalian cardiovascular function. Because of technical and economic considerations, these experiments are largely confined to the mouse. Genetic modification of the loci responsible for aspects of cardiac development, differentiation, and function via gene targeting, as well as modulation of the cardiac protein complement using transgenesis, has begun to provide mouse models of cardiac hypertrophy, dilated cardiomyopathy, and hypertrophic cardiomyopathies. In order to use these animal models fully and explore their phenotypes at the whole organ and whole animal levels, the extension of cardiovascular physiological methodologies to the mouse is imperative. Techniques for exploring aspects of cardiovascular function are well developed for larger animal models, but their modification for the small size of the mouse heart and for the animal's rapid cardiac cycle has proven to be a formidable challenge, requiring the combined efforts of the molecular biology, physiology, and cardiology communities. We review here the ability of present‐day technology to obtain reproducible data on murine cardiac function at the whole organ and animal levels. (Circ Res. 1998;82:407‐415.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;Recently, we developed a transgenic mouse with cardiac‐specific G (s) alpha overexpression (TG mouse), which exhibits enhanced postsynaptic beta‐adrenergic receptor signaling, ultimately developing a cardiomyopathy. The goal of the present study was to determine whether cardiac Gsalpha overexpression alters autonomic cardiovascular control, which could shed light on the mechanism responsible for the later development of cardiomyopathy. Mean arterial pressure was increased (P<.05) in conscious, chronically instrumented TG mice (123 +/‐ 1 mm Hg) compared with age‐matched wild‐type (WT) control mice (103 +/‐ 1 mm Hg). Respiratory frequency was increased (P<.05) in TG mice (269 +/‐ 26/min) compared with WT mice (210 +/‐ 20/min). By use of telemetric techniques, baseline heart rate (HR) was elevated (P<.05) in conscious, untethered TG mice (696 +/‐ 13 bpm) compared with WT mice (568 +/‐ 28 bpm). Intrinsic HR, after propranolol and atropine or after ganglionic blockade with hexamethonium, was not different between TG and WT mice. Both the normal minute‐to‐minute and circadian variations of HR observed in WT mice were markedly blunted in TG mice. HR variability was assessed by the time‐domain and frequency‐domain methods. At baseline, time‐domain analysis indices were reduced (P<.05) in TG mice compared with WT mice. Although the low frequency (LF) component was higher (P<.05) than the high frequency (HF) component in WT mice, the LF component was less (P<.05) than the HF component in TG mice. In addition, arterial baroreflex regulation of HR was markedly blunted in TG mice in response to both nitroglycerin‐induced hypotension and phenylephrine‐induced hypertension. The reduced LF/HF ratio in TG mice was surprising in view of enhanced beta‐adrenergic signaling and may be due to reduced neural tone secondary to the elevated arterial pressure or alterations in arterial baroreflex control. Dobutamine infusion in WT mice also resulted in depressed HR variability. The combination of elevated baseline HR, arterial pressure, and respiratory frequency suggests that enhanced beta‐adrenergic signaling in TG mice results in reduced HR variability, in terms of both minute‐to‐minute variability and the lack of circadian variations in HR. The lack of normal HR variability in general and the failure of HR to decline, even during sleep, may actually be critical mechanisms contributing to the ultimate development of cardiomyopathy in these animals. (Circ Res. 1998;82:416‐423.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;The goal of this study was to examine the relationship between Ca (2+) entry through L‐type Ca2+channels and local [Ca2+]itransients (Ca2+sparks) in single rat cardiac ventricular cells. L‐type Ca2+channels were activated by depolarization of cell‐attached membrane patches, and [Ca2+] (i) was measured simultaneously as fluo 3 fluorescence using laser scanning conforcal microscopy. Patch depolarization with Ca2+as the charge carrier (10 or 110 mmol [center dot] L‐1) significantly increased the probability of the occurrence of Ca2+sparks (Ca2+spark rate) only in the volume of cytoplasm located immediately beneath the membrane patch (basal Ca2+spark rate, 119 Ca2+sparks [center dot] cell‐1[center dot] s‐1; patch depolarization Ca2+spark rate, 610 Ca2+sparks [center dot] cell‐1[center dot] s‐1; P<.005). With Ba2+in the pipette solution (10 mmol [center dot] L‐1), patch depolarization was not associated with an increased Ca2+spark rate at the position of the pipette or at any other sites distant from the pipette. Therefore, Ca2+entry and not voltage per se was a necessary event for the occurrence of Ca2+sparks. Under identical experimental conditions, patch depolarization experiments opened single L‐type Ca2+channels with a single‐channel conductance of 19 pS with Ba2+as the charge carrier. Although single‐channel openings could not be resolved when Ca2+was the charge carrier, ensemble averages yielded an inward current of up to 0.75 pA. The results suggest that voltage‐activated Ca2+entry through one or a small number of L type Ca2+channels triggers the release of Ca2+only from the sarcoplasmic reticulum in direct proximity to those L‐type Ca2+channels. The relatively low probability of triggering Ca2+sparks may have resulted from some alteration of excitation‐contraction coupling associated with the technique of the cell‐attached patch clamp. (Circ Res. 1998;82:424‐429.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;We have previously reported that stretching of cardiomyocytes activates the phosphorylation cascade of protein kinases, including Raf‐1 kinase and mitogen‐activated protein (MAP) kinases, followed by an increase in protein synthesis partly through enhanced secretion of angiotensin II and endothelin‐1. Membrane proteins, such as ion channels and exchangers, have been postulated to first receive extracellular stimuli and evoke intracellular signals. The present study was performed to determine whether mechanosensitive ion channels and exchangers are involved in stretch‐induced hypertrophic responses. Neonatal rat cardiomyocytes cultured on expandable silicone dishes were stretched after pretreatment with a specific inhibitor of stretch‐sensitive cation channels (gadolinium and streptomycin), of ATP‐sensitive K+channels (glibenclamide), of hyperpolarization‐activated inward channels (CsCl), or of the Na+‐H+exchanger (HOE 694). Pretreatment with gadolinium, streptomycin, glibenclamide, and CsCl did not show any inhibitory effects on MAP kinase activation by mechanical stretch. HOE 694, however, markedly attenuated stretch‐induced activation of Raf‐1 kinase and MAP kinases by [=approximate]50% and 60%, respectively, and attenuated stretch‐induced increase in phenylalanine incorporation into proteins. In contrast, HOE 694 did not inhibit angiotensin II‐and endothelin‐1‐induced Raf‐1 kinase and MAP kinase activation. These results suggest that among many mechanosensitive ion channels and exchangers, the Na+‐H+exchanger plays a critical role in mechanical stress‐induced cardiomyocyte hypertrophy. (Circ Res. 1998;82:430‐437.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;Cytotoxic T lymphocytes (CTLs) that infiltrate the heart are important immune effectors implicated in heart transplant rejection, myocarditis, and other cardiomyopathies. To investigate the mechanism(s) underlying CTL damage to the myocardium through activation of the Fas receptor (Fas/CD95/Apo‐1) by the Fas ligand, we explored the interaction between peritoneal exudate CTLs (PELs), derived from perforin gene‐knockout (P‐/‐) mice, and murine ventricular myocytes. Fas expression on isolated ventricular myocytes was demonstrated immunohistochemically. Action potentials, [Ca2+]itransients, and contractions of myocytes conjugated to P‐/‐ PELs or treated with the apoptosis‐inducing anti‐Fas monoclonal antibody Jo2 were recorded. Action potential characteristics of nonconjugated myocytes and myocytes conjugated with P‐/‐ PELs were, respectively, as follows: Vm, ‐73.2 +/‐ 1.5 and ‐53.6 +/‐ 6.4 mV (mean +/‐ SEM); action potential amplitude, 117.9 +/‐ 3.9 and 74.3 +/‐ 21.2 mV; and action potential duration at 80% repolarization, 17 +/‐ 6 and 42 +/‐ 13 milliseconds (all P<.05). P‐/‐ PELs also induced early and delayed afterdepolarizations as well as arrhythmogenic activity. Diastolic [Ca2+]iincreased during the cytocidal interaction with P‐/‐ PELs, from a fluorescence ratio of 0.82 +/‐ 0.05 (n=7) to 1.98 +/‐ 0.09 (n=13) (P<.05). All of the effects caused by P‐/‐ PELs were reproduced by incubating the myocytes with Jo2. Heparin (50 [micro sign]g/mL), an antagonist of inositol trisphosphate (IP3)‐operated sarcoplasmic reticulum Ca2+channels, or U‐73122 (2 [micro sign]mol/L), a phospholipase C inhibitor, but not the inactive agonist U‐73343, prevented Fas‐mediated myocyte dysfunction. Additionally, intracellular application (through the patch pipette) of the active IP3analogue, inositol 1,4,5‐trisphosphate, but not the inactive analogue, inositol 1,3,4‐trisphosphate, caused electrophysiological changes resembling those resulting from P‐/‐ PELs and Jo2, suggesting that CTL‐induced Fas‐based myocyte dysfunction is mediated by IP (3). We conclude that a Fas‐based perforin‐independent mechanism of CTL action can account for the immunopathology seen in the allotransplanted heart, myocarditis, and dilated cardiomyopathy. (Circ Res. 1998;82:438‐450.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;The aim of this study was to investigate whether treatment with the protein kinase C (PKC) agonist 1,2‐dioctanoyl‐sn‐glycerol (1,2DOG) can protect isolated adult Wistar rat cardiomyocytes against simulated ischemia and reoxygenation. Cytosolic Ca2+(assessed by fura 2 fluorescence), pHi(assessed by BCECF fluorescence), and cell length were measured during 80 minutes of simulated ischemia (anoxia, pH (o) 6.4) and 20 minutes of reoxygenation (pHo7.4) and compared between control cells and cells treated with 20 [micro sign]mol/L 1,2DOG before anoxia (10‐minute treatment and 10‐minute washout), before and during anoxia (two‐step treatment), or only during anoxia. Treatment before anoxia attenuated rigor contracture but did not influence anoxic Ca2+overload. In contrast, two‐step treatment before and during anoxia accelerated rigor contracture but reduced the rate of anoxic Ca2+accumulation. During reoxygenation, control cells developed irreversible hypercontracture (reduction of cell length to 43 +/‐ 2% of the initial cell length, n=62), which was accompanied by spontaneous oscillations of cytosolic Ca2+(19.6 +/‐ 1.6 per minute). Two‐step treatment with 1,2DOG before and during anoxia significantly reduced hypercontracture (reduction of cell length to 60 +/‐ 2%, P<.01 versus control, n=41) and suppressed spontaneous Ca2+oscillations (2.8 +/‐ 0.9 per minute, P<.01 versus control). These effects could not be reproduced by treatment with 1,2DOG before anoxia or during anoxia or by a two‐step treatment with the PKC‐inactive 1,3‐dioctanoyl‐sn‐glycerol and were fully abolished with 1 [micro sign]mol/L bisindolylmaleimide (PKC inhibitor). We conclude that a two‐step activation of PKC before and during anoxia is required for effective protection of cardiomyocytes against anoxic Ca2+overload and reoxygenation‐induced hypercontracture. (Circ Res. 1998;82:451‐457.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;One possible mechanism for neurohumoral activation after myocardial infarction may be the generation of an immune response against cardiac self‐antigens. We hypothesize that if there is a T cell‐mediated reaction to self‐antigens, the transfer of splenic lymphocytes from postinfarct rats into syngeneic rats with normal hearts should result in a T cell‐mediated autoimmune myocarditis in the healthy syngeneic rats. Rats were killed 6 weeks after coronary ligation. Splenocytes from animals with large and small infarcts were purified from spleens, activated with concanavalin A, and injected in varying doses into normal syngeneic rats. These recipient rats were killed 6 weeks later, and histopathological studies were performed. Our results demonstrate in vivo evidence of lymphocyte‐mediated myocardial injury by adoptive transfer of sensitized lymphocytes from rats who developed congestive heart failure after acute myocardial infarction. The amount of infiltrate and necrosis in the recipient rats appeared directly related to the size of the infarct from the donor rats. This suggests that larger infarcts lead to a greater inflammatory response as well as a greater propensity for alteration of cardiac surface antigens or the emergence of previously sequestered antigens. None of the other organs (kidney, liver, lung, or brain) had evidence of infiltrates. Two‐dimensional echocardiography did not reveal systolic dysfunction. This study provides direct evidence of autoimmune myocardial injury produced by adoptive transfer of concanavalin A‐activated splenocytes after myocardial infarction. We propose that neurohumoral activation early in the postinfarction period triggers a series of specific inflammatory and immunological events that lead to formation of specific clones of T cells. When these are activated and transferred into normal rats, cardiac‐specific cellular infiltration occurs, occasionally accompanied by myocardial necrosis. This model should help to further explore the link between neurohumoral activation after myocardial infarction and the subsequent immune alterations that might be associated with the development and/or progression of congestive heart failure. Additionally, this might be a useful model in which to study other immune‐mediated cardiomyopathies. (Circ Res. 1998;82:458‐463.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;The outflow tract (OFT) provides the structural components forming the ventriculoarterial connection. The prevailing concept that this junction “rotates” to acquire its definitive topography also requires a concept of “counterrotation” and is difficult to reconcile with cell‐marking studies. Rats between 10 embryonic days (EDs) and 2 postnatal days were stained immunohistochemically and by in situ hybridization. DNA replication was determined by incorporation of bromodeoxyuridine and apoptosis by the annexin V binding and terminal deoxynucleotidyl transferase‐mediated dUTP‐X nick end labeling (TUNEL) assays. Starting at ED12, cardiomyocytes in the distal (truncal) part of the OFT begin to shed their myocardial phenotype without proceeding into apoptosis, suggesting transdifferentiation. Myocardial regression is most pronounced on the dextroposterior side and continues until after birth, as revealed by the disappearance of the myocardial cuff surrounding the coronary roots and semilunar sinuses and by the establishment of fibrous continuity between mitral and aortic semilunar valves. Fusion of the endocardial ridges of the truncus on late ED13 is accompanied by the organization of alpha‐smooth muscle actin‐and nonmuscle myosin heavy chain‐positive myofibroblasts into a central whorl and the appearance of the semilunar valve anlagen at their definitive topographical position within the proximal portion of the truncus. After fusion of the proximal (conal) portion of the endocardial ridges, many of the resident myofibroblasts undergo apoptosis and are replaced by cardiomyocytes. The distal myocardial boundary of the OFT is not a stable landmark but moves proximally over the spiraling course of the aortic and pulmonary routes, so that the semilunar valves develop at their definitive topographic position. After septation, the distal boundary of the OFT continues to regress, particularly in its subaortic portion. The myocardializing conus septum, on the other hand, becomes largely incorporated into the right ventricle. These opposite developments account for the pronounced asymmetry of the subaortic and subpulmonary outlets in the formed heart. (Circ Res. 1998;82:464‐472.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;The effect of angiotensin II (Ang II) on the activity of the cardiac Na+‐independentCl‐‐HCO3‐exchanger (anionic exchanger [AE]) was explored in cat papillary muscles. pHiwas measured by epifluorescence with BCECF‐AM. Ang II (500 nmol/L) induced a 5‐(N‐ethyl‐N‐isopropyl)amiloride‐sensitive increase in pHiin the absence of external HCO3‐(HEPES buffer), consistent with its stimulatory action on Na+‐H+exchange (NHE). This alkalinizing effect was not detected in the presence of a CO2‐HCO3‐buffer (pH (i) 7.07 +/‐ 0.02 and 7.08 +/‐ 0.02 before and after Ang II, respectively; n=17). Moreover, in Na+‐freeHCO3‐‐bufferedmedium, in which neither NHE nor Na+‐HCO3‐cotransport are acting, Ang II decreased pHi, and this effect was canceled by previous treatment with SITS. These findings suggested that the Ang II‐induced activation of NHE was masked, in the presence of the physiological buffer, by a HCO3‐‐dependentacidifying mechanism, probably the AE. This hypothesis was confirmed on papillary muscles bathed with HCO3‐buffer that were first exposed to 1 [micro sign]mol/L S20787, a specific inhibitor of AE activity in cardiac tissue, and then to 500 nmol/L Ang II (n=4). Under this condition, Ang II increased pHifrom 7.05 +/‐ 0.05 to 7.22 +/‐ 0.05 (P<.05). The effect of Ang II on AE activity was further explored by measuring the velocity of myocardial pHirecovery after the imposition of an intracellular alkali load in a HCO3‐‐containingsolution either with or without Ang II. The rate of myocardial pHirecovery was doubled in the presence of Ang II, suggesting a stimulatory effect on AE. The enhancement of the activity of this exchanger by Ang II was also detected when the AE activity was reversed by the removal of extracellular Cl‐in a Na+‐freesolution. Under this condition, the rate of intracellular alkalinization increased from 0.053 +/‐ 0.016 to 0.108 +/‐ 0.026 pH unit/min (n=6, P<.05) in the presence of Ang II. This effect was canceled either by the presence of the AT1receptor antagonist, losartan, or by the previous inhibition of protein kinase C with chelerythrine or calphostin C. The above results allow us to conclude that Ang II, in addition to its stimulatory effect on alkaline loading mechanisms, activates the AE in ventricular myocardium and that the latter effect is mediated by a protein kinase C‐dependent regulatory pathway linked to the AT1receptors. (Circ Res. 1998;82:473‐481.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID

摘要:

&NA;The development of congestive heart failure (CHF) is associated with left ventricular (LV) dilation and myocardial remodeling. However, fundamental mechanisms that contribute to this remodeling process with the progression of CHF remain unclear. The matrix metalloproteinases (MMPs) have been demonstrated to play a significant role in tissue remodeling in a number of pathological processes. The present project tested the hypothesis that the LV dilation and remodeling during the progression of CHF is associated with early changes in MMP expression and zymographic activity. LV and myocyte function, collagen content, and MMP expression and zymographic activity were serially measured during the progression of CHF caused by pacing‐induced supraventricular tachycardia (SVT) in pigs. After 7 days of SVT, LV end‐diastolic dimension and myocyte length both increased by 15% from control values, and LV fractional shortening fell by 20%. At the level of the myocyte, percent shortening fell by 16% after 7 days of SVT, with no change in the steady‐state velocity of shortening. Longer durations of SVT caused progressive LV dilation, LV pump failure, and myocyte contractile dysfunction. Specifically, 21 days of SVT resulted in a >50% increase in LV dimension, a 56% fall in LV fractional shortening, and a 33% decline in myocyte velocity of shortening. The decline in LV and myocyte function with 21 days of SVT was accompanied by signs and symptoms of CHF. Thus, SVT causes time‐dependent changes in LV geometry and function and the subsequent development of CHF. LV myocardial collagen content and confluence fell by >25% after 7 days of SVT and were accompanied by an 80% increase in LV myocardial MMP zymographic activity against the substrate gelatin. After 14 days of SVT, total LV myocardial collagen content was reduced by 24%, and LV myocardial MMP zymographic activity increased by >100% from control values. Interstitial collagenase (MMP‐1), stromelysin (MMP‐3), and 72‐kD gelatinase (MMP‐2) were increased by [=approximate]2‐fold after 7 days of SVT. LV MMP zymographic activity and abundance remained elevated with longer durations of SVT. The results of the present study demonstrated that in this model of CHF, early changes in LV myocardial MMP zymographic activity and protein levels occurred with the initiation and progression of LV dilation and dysfunction. These findings suggest that an early contributory mechanism for the initiation of LV remodeling that occurred in this model of developing CHF is enhanced expression and potentially increased activity of LV myocardial MMPs. (Circ Res. 1998;82:482‐495.)

ISSN:0009-7330    

出版商:OVID    

年代:1998

数据来源: OVID