Chronic L-arginine treatment increases cardiac cyclic guanosine 5'-monophosphate in rats with aortic stenosis: effects on left ventricular mass and beta-adrenergic contractile reserve

OBJECTIVES: We tested the hypothesis that nitric oxide (NO) cyclic guanosine 5'-monophosphate (GMP) signaling is deficient in pressure overload hypertrophy due to ascending aortic stenosis, and that long-term L-arginine treatment will increase cardiac cyclic GMP production and modify left ventricular (LV) pressure overload hypertrophy and beta-adrenergic contractile response. BACKGROUND: Nitric oxide cyclic GMP signaling is postulated to depress vascular growth, but its effects on cardiac hypertrophic growth are controversial. METHODS: Forty control rats and 40 rats with aortic stenosis left ventricular hypertrophy ([LVH] group) were randomized to receive either L-arginine (0.40 g/kg/day) or no drug for 6 weeks. RESULTS: The dose of L-arginine did not alter systemic blood pressure. Animals with LVH had similar LV constitutive nitric oxide synthase (cNOS) mRNA and protein levels, and LV cyclic GMP levels as compared with age-matched controls. In rats with LVH L-arginine treatment led to a 35% increase in cNOS protein levels (p = 0.09 vs untreated animals with LVH) and a 1.7-fold increase in LV cyclic GMP levels (p < 0.05 vs untreated animals with LVH). However, L-arginine treatment did not suppress LVH in the animals with aortic stenosis. In contrast, in vivo LV systolic pressure was depressed in L-arginine treated versus untreated rats with LVH (163 +/- 16 vs 198 +/- 10 mm Hg, p < 0.05). In addition, the contractile response to isoproterenol was blunted in both isolated intact hearts and isolated myocytes from L-arginine treated rats with LVH compared with untreated rats with LVH. This effect was mediated by a blunted increase in peak systolic intracellular calcium in response to beta-adrenergic stimulation. CONCLUSIONS: Left ventricular hypertrophy due to chronic mechanical systolic pressure overload is not characterized by a deficiency of LV cNOS and cyclic GMP levels. In rats with aortic stenosis, L-arginine treatment increased cardiac levels of cyclic GMP, but it did not modify cardiac mass in rats with aortic stenosis. However, long-term stimulation of NO-cyclic GMP signaling depressed in vivo LV systolic function in LVH rats and markedly blunted the contractile response to beta-adrenergic stimulation.     

Cellular basis of contractile derangements of hypertrophied feline ventricular myocytes

The objective of this study was to further explore the cellular basis of the reduced rate and magnitude of contraction of feline left ventricular myocytes with severe hypertrophy induced by slow progressive pressure overload. A 3.0 mm internal diameter band was placed around the ascending aorta of 12 young (8-10 weeks old) cats, and sham operations were performed in 13 others. This caused no major pressure overload initially, but 15 weeks later there was a significant pressure gradient across the band (56+/-14 mmHg) and the heart weight to body weight ratio had increased from 4.2-7 gm/kg. Contraction rates and magnitudes of myocytes isolated from the hearts with hypertrophy (LVH) were significantly slower and smaller, respectively, than those from control (C) animals. Indo-1 fluorescence transients in LVH myocytes were significantly smaller in magnitude and longer in duration than in C, suggesting that contractile defects result from Ca2+ derangements. Elevation of bath Ca2+ increased the peak Indo-1 fluorescence and the rate and magnitude of contraction in all myocytes. At the bath Ca2+ which had maximal inotropic effects there were no differences in the peak Indo-1 fluorescence in LVH and C myocytes, but contraction magnitude remained significantly smaller in LVH. This suggests that there are Ca2+-independent contractile derangements in LVH. In support of this hypothesis, the relationship between contraction magnitude and the peak Indo-1 fluorescence (index of myofibrillar Ca2+ sensitivity) was significantly shifted in LVH myocytes, suggesting that myofibrillar Ca2+ sensitivity was reduced. There was also a significant shift of the terminal portions of hysteresis loops of cell length v indo-1 fluorescence ratio, providing additional support for this idea. Experiments with isoproterenol suggest that it can reduce myofibrillar Ca2+ sensitivity in C, but not LVH myocytes. The idea that increased internal resistance to shortening (internal load) is responsible for the contractile defects of LVH myocytes was examined by defining the relationship between the rate of relengthening and the magnitude of shortening. There was no significant difference in this relation between C and LVH myocytes. In addition, colchicine (which depolymerizes microtubular tubulin) had no significant effect on contraction magnitude in either C or LVH myocytes. These results suggest that the contractile properties of feline LVH myocytes result from changes in cellular Ca2+ regulation and myofibrillar Ca2+ sensitivity, but not from changes in the internal loading.     

Decompensation of pressure-overload hypertrophy in G alpha q-overexpressing mice

BACKGROUND: Receptor-mediated activation of myocardial Gq signaling is postulated as a biochemical mechanism transducing pressure-overload hypertrophy. The specific effects of Gq activation on the functional and morphological adaptations to pressure overload are not known. METHODS AND RESULTS: To determine the effects of intrinsic myocyte G alpha q signaling on the left ventricular hypertrophic response to experimental pressure overload, transgenic mice overexpressing G alpha q specifically in the heart (G alpha q-25) and nontransgenic siblings underwent microsurgical creation of transverse aortic coarctation and the morphometric, functional, and molecular characteristics of these pressure-overloaded hearts were compared at increasing times after surgery. Before aortic banding, isolated G alpha q-25 ventricular myocytes exhibited contractile depression (depressed +dl/dt and -dl/dt) and G alpha q-25 hearts showed a pattern of fetal gene expression similar to the known characteristics of nontransgenic pressure-overloaded mice. Three weeks after transverse aortic banding, G alpha q-25 left ventricles hypertrophied to a similar extent (approximately 30% increase) as nontransgenic mice. However, whereas nontransgenic mice exhibited concentric left ventricular remodeling with maintained ejection performance (compensated hypertrophy), G alpha q-25 left ventricles developed eccentric hypertrophy and ejection performance deteriorated, ultimately resulting in left heart failure (decompensated hypertrophy). The signature hypertrophy-associated progress of fetal cardiac gene expression observed at baseline in G alpha q-25 developed after aortic banding of nontransgenic mice but did not significantly change in aortic-banded G alpha q-25 mice. CONCLUSIONS: Intrinsic cardiac myocyte G alpha q activation stimulates fetal gene expression and depresses cardiac myocyte contractility. Superimposition of the hemodynamic stress of pressure overload on G alpha q overexpression stimulates a maladaptive form of eccentric hypertrophy that leads to rapid functional decompensation. Therefore G alpha q-stimulated cardiac hypertrophy is functionally deleterious and compromises the ability of the heart to adapt to increased mechanical load. This finding supports a reevaluation of accepted concepts regarding the mechanisms for compensation and decompensation in pressure-overload hypertrophy.     

MRI evaluation of right ventricular pressure overload in chronic obstructive pulmonary disease

In chronic obstructive pulmonary disease (COPD), the development of pulmonary hypertension is common. This study was performed to assess the signs of right ventricular (RV) pressure overload and RV failure in COPD. In 8 COPD patients without primary cardiac disease, RV wall thickness, mass, and end-diastolic volume were measured by cardiac-triggered cine MRI. MR phase-contrast velocity quantification was used to measure stroke volume and the patterns of flow into and out of the RV. Data of patients were tested versus those of a control group matched for age (n = 8). Results showed that the RV wall thickness was increased (.6 +/- 0.1 vs 0.4 +/- 0.1 cm, P < .001). RV mass was increased (67 +/- 11 vs 57 +/- 5 g, P < .005). RV stroke volume was decreased (57 +/- 13 vs 71 +/- 13 ml, P < .01), but RV ejection fraction was not different. In the main pulmonary artery flow, the quotient of acceleration time divided by ejection time was decreased (33 +/- 5% vs 38 +/- 4%, P < .05), which is indicative of pulmonary hypertension. In conclusion, this MRI protocol provides a tool to assess the effects of RV pressure overload in COPD before heart failure has become manifest.     

Contractile properties of aged avian muscle after stretch-overload

The effects of ageing on muscle contractile adaptations to stretch-overload was examined in the anterior latissimus dorsi (ALD) muscle of 12 old (90 weeks of age) and 12 young adult (10 weeks of age) Japanese quails. A weight corresponding to 12% of the birds' body weight was attached to one wing for 30 days, while the contralateral wing served as the intra-animal control. In vitro contractile measurements were made at 25 degrees C by indirect stimulation of the ALD by its nerve (pulse 0.2 ms). Compared with young adult twitch characteristics, aged muscles had significantly greater contraction time (149 +/- 9 ms vs. 174 +/- 16 ms). Stretch-overload increased contraction time to 162 +/- 7 ms in young muscles and 215 +/- 14 ms in old muscles. Ageing and overload resulted in a greater fusing of twitches at stimulation frequencies of 5 and 10 Hz which resulted in a leftward shift of the force-frequency curve at these frequencies, relative to young adult control muscles. Maximal shortening velocity (Vmax) decreased from 2.6 +/- 0.3 to 1.2 +/- 0.1 muscle lengths/s in young muscles after overload. Vmax in old control muscles was similar to young muscles after stretch, but stretch further decreased Vmax in old muscles to 0.8 muscle lengths/s. Maximal tetanic force and specific force were similar in young and old muscles, both before and after stretch. These data indicate that ageing induces a slowing of both twitch contractile characteristics and shortening velocity in the ALD, without affecting maximal force capabilities.     

Role of altered beta-adrenoceptor signal transduction in the pathogenesis of cirrhotic cardiomyopathy in rats

BACKGROUND & AIMS: Attenuated ventricular contractility has been documented in cirrhosis, but the pathogenesis remains unclear. The beta-adrenergic receptor system is critical in modulating cardiac contraction. Therefore, the aim of this study was to clarify beta-adrenoceptor signaling function in a rat model of cirrhosis. METHODS: Cirrhosis was induced by bile duct ligation, whereas controls underwent a sham operation. Myocardial contractility was studied by measuring isolated left ventricular papillary muscle contraction under isoproterenol stimulation. Beta-Adrenoceptor signaling was evaluated by measuring adenosine 3',5'-cyclic monophosphate generation after stimulation with isoproterenol, sodium fluoride, and forskolin. Guanosine triphosphate-binding protein expression from ventricular plasma membranes was determined by Western blots to measure G(s)alpha, Gi2alpha, and G(common)beta, respectively. RESULTS: Maximum papillary muscle contractile responses in control and cirrhotic rats were 113% +/- 3% and 70% +/- 2% of basal, respectively (P<0.01), with no significant differences in the dose-inducing half-maximal response. Adenosine 3', 5'-cyclic monophosphate generation after stimulation with all three agents was significantly lower in cirrhotic compared with control rat membranes. G(s)alpha and Gi2alpha expression was significantly reduced in cirrhotics compared with controls, but G(common)beta expression remained unchanged. CONCLUSIONS: These data showed cardiac contractile impairment in cirrhosis, associated with altered beta-adrenergic receptor signaling function and guanine nucleotide-binding protein expression. These factors may play an important role in the pathogenesis of cirrhotic cardiomyopathy.     

Heart muscle after heart infarct. Function of surviving heart muscle following acute myocardial infarct

In the initial phase of the infarction, there is a decrease of ventricular function due to loss of contractile activity. In addition, a negative effect of the paradoxical movement of the infarcted area on the hemodynamics of the ventricle is noted. The stiffening of the infarcted area in the early stage has a favorable influence on ventricular function, leading to a change in the elastic properties of the ventricle. The loss of ventricular compliance persists after the infarction, and its severity depends on the extent of myocardial destruction. In the non-compliant ventricle, the end-diastolic pressure rises without a proportionate increase in volume. Consequently, the ventricular function curve shows a shift downward and to the right, making it difficult to distinguish between the loss of contractile function or compliance in the heart in situ. Evaluation of the contractile properties of the surviving cardiac muscle in situ is, however, hardly possible due to the changed geometry and the additonal elastic elements functioning in series with the surviving muscle. To exclude these factors, a study of the contractile properties of the surviving cardiac muscle in the isolated state was carried out following experimental myocardial infarction in cats. By ligating several coronary branches, infarctions in the area of the left ventricle were caused; to avoid the ischaemic border zone of the infarction, right ventricular papillary muscles were studied. Haemodynamic investigations showed an increase in right ventricular end-diastolic pressure which persisted 6 weeks after infarction. As early as 48 hours but, more significantly 1 week after infarction, there was a decrease of actively developed force in the surviving cardiac muscle due to a lower rate of force development. The resting length tension curve of the surviving cardiac muscle after infarction showed no alterations; and 6 weeks following infarction, almost normal contractility parameters were observed. As a result of the infarction, a decrease in contractility in the surviving cardiac muscle is observed during the early stage, which regresses after complete recuperation.     

Targeted overexpression of the sarcoplasmic reticulum Ca2+-ATPase increases cardiac contractility in transgenic mouse hearts

Cardiac hypertrophy and heart failure are known to be associated with a reduction in Ca2+-ATPase pump levels of the sarcoplasmic reticulum (SR). To determine whether, and to what extent, alterations in Ca2+ pump numbers can affect contraction and relaxation parameters of the heart, we have overexpressed the cardiac SR Ca2+-ATPase specifically in the mouse heart using the alpha-myosin heavy chain promoter. Analysis of 2 independent transgenic lines demonstrated that sarco(endo)plasmic reticulum Ca2+-ATPase isoform (SERCA2a) mRNA levels were increased 3.88+/-0. 4-fold and 7.90+/-0.2-fold over those of the control mice. SERCA2a protein levels were increased by 1.31+/-0.05-fold and 1.54+/-0. 05-fold in these lines despite high levels of mRNA, suggesting that complex regulatory mechanisms may determine the SERCA2a pump levels. The maximum velocity of Ca2+ uptake (Vmax) was increased by 37%, demonstrating that increased pump levels result in increased SR Ca2+ uptake function. However, the apparent affinity of the SR Ca2+-ATPase for Ca2+ remains unchanged in transgenic hearts. To evaluate the effects of overexpression of the SR Ca2+ pump on cardiac contractility, we used the isolated perfused work-performing heart model. The transgenic hearts showed significantly higher myocardial contractile function, as indicated by increased maximal rates of pressure development for contraction (+dP/dt) and relaxation (-dP/dt), together with shortening of the normalized time to peak pressure and time to half relaxation. Measurements of intracellular free calcium concentration and contractile force in trabeculae revealed a doubling of Ca2+ transient amplitude, with a concomitant boost in contractility. The present study demonstrates that increases in SERCA2a pump levels can directly enhance contractile function of the heart by increasing SR Ca2+ transport.     

Adrenomedullin gene expression in the rat heart is stimulated by acute pressure overload: blunted effect in experimental hypertension

The levels of adrenomedullin (ADM), a newly discovered vasodilating and natriuretic peptide, are elevated in plasma and ventricular myocardium in human congestive heart failure suggesting that cardiac synthesis may contribute to the plasma concentrations of ADM. To examine the time course of induction and mechanisms regulating cardiac ADM gene expression, we determined the effect of acute and short-term cardiac overload on ventricular ADM mRNA and immunoreactive ADM (ir-ADM) levels in conscious rats. Acute pressure overload was produced by infusion of arginine8-vasopressin (AVP, 0.05 microg/kg/min, i.v.) for 2 h into 12-week-old hypertensive TGR(mREN-2)27 rats and normotensive Sprague-Dawley (SD) rats. Hypertension and marked left ventricular hypertrophy were associated with 2.2-times higher ir-ADM levels in the left ventricular epicardial layer (178 +/- 36 vs. 81 +/- 23 fmol/g, P<0.05) and 2.6-times higher ir-ADM levels in the left ventricular endocardial layer (213 +/- 23 vs. 83 +/- 22 fmol/g, P<0.01). The infusion of AVP for 2 h in normotensive rats produced rapid increases in the levels of left ventricular ADM mRNA (epicardial layer: 1.6-fold, P<0.05) and ir-ADM (endocardial layer: from 83 +/- 22 to 140 +/- 12 fmol/g, P<0.05), whereas ventricular ADM mRNA and ir-ADM levels did not change significantly in hypertensive rats. Short-term cardiac overload, induced by administration of angiotensin II (33.3 microg/kg/h, s.c., osmotic minipumps) for two weeks in normotensive SD rats resulted in left ventricular hypertrophy (3.05 +/- 0.17 vs. 2.75 +/- 0.3 mg/g, P<0.05) and a 1.5-fold increase (P<0.05) in ventricular ADM mRNA levels. In conclusion, the present results show that pressure overload acutely stimulated ventricular ADM gene expression in conscious normotensive rats suggesting a potential beneficial role for endogenous ADM production in the heart against cardiac overload. Since pressure overload-induced increase in ADM synthesis was attenuated in hypertensive rats, alterations in the ADM system may contribute to the pathogenesis of hypertension in the TGR(mREN-2)27 rat.     

Impaired regional fatty acid uptake and systolic dysfunction in hypertrophied right ventricle

Little information is available regarding the determinants of systolic contractile function of the hypertrophied right ventricle (RV). The purpose of this study was to clarify the relationship between myocardial metabolism and contractile function in the hypertrophied RV due to pulmonary hypertension (PH). METHODS: Iodine-123-labeled 15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid (BMIPP) and 99mTc-sestamibi (MIBI) SPECT were performed to calculate the RV-to-left ventricle (LV) tracer uptake ratio (RV/LV) in 21 patients with PH (6 with primary PH and 15 with chronic thromboembolic PH). The patients also underwent electron-beam CT to assess RV ejection function (RVEF) and percentage systolic wall thickening (%SWT) and right heart catheterization to measure mean pulmonary arterial pressure (mPAP). RESULTS: There were significant positive correlations between mPAP and MIBI-RV/LV (r = 0.89, p < 0.001) and between mPAP and BMIPP-RV/LV (r = 0.86, p < 0.001). However, 8 patients showed lower BMIPP-RV/LV than MIBI-RV/LV, indicating the impairment of myocardial fatty acid uptake in the RV. These patients had lower RVEF and %SWT compared to those with normal myocardial fatty acid uptake (RVEF = 28% +/- 10% compared to 40% +/- 9% and %SWT = 33% +/- 27% compared to 74% +/- 30%, respectively; p < 0.05 for both comparisons). Although mPAP did not differ between the groups, the RVEF-mPAP and %SWT-mPAP regression lines drawn from the patients with impaired myocardial fatty acid uptake were located below the lines from the patients with normal myocardial fatty acid uptake, suggesting disproportionately decreased RV myocardial contractility for a given mPAP in patients with impaired myocardial fatty acid uptake. The patients with the impaired fatty acid uptake in the RV had a significantly higher death rate (log-rank test, p < 0.05). CONCLUSION: The results from this preliminary study suggest that myocardial fatty acid uptake is impaired in the failing hypertrophied RV due to PH.     

Reduced left ventricular hypertrophy in type 1 diabetic patients with end-stage renal failure. A comparison between groups investigated 1977-80 and 1991-93

BACKGROUND: During the last decade, control of hypertension, oedema, anaemia, uraemia, and blood glucose has improved in patients with diabetic nephropathy. We have investigated whether this has influenced cardiac function at the time of end-stage renal failure. STUDY DESIGN: Echocardiographic investigations were performed in 26 type 1 diabetic patients evaluated for kidney transplantation and the results compared with those obtained in healthy controls and in a similar group of patients investigated in 1977-1980. RESULTS: Blood pressure was 153 +/- 21/85 +/- 12 mmHg versus 174 +/- 17/91 +/- 9 (recent group versus early group). The left ventricular (LV) diameter index, a measure of volaemia, was increased in systole and diastole in the early but not in the recent group. Both groups had LV hypertrophy, but this was much less pronounced in the recent group; posterior wall thickness was 1.1 +/- 0.16 cm versus 1.3 +/- 0.26 cm (P = 0.0001) and LV mass index 132 +/- 43 g/m2 versus 166 +/- 44 g/m2 (P = 0.009). Blood pressure correlated significantly with indices of LV hypertrophy in the recent group. Systolic function was normal in both groups but diastolic function was disturbed in both and to the same extent, atrial systole contributing by 27 +/- 14% to ventricular filling. CONCLUSION: Better treatment of hypertension, fluid overload, and uraemia has led to less pronounced LV hypertrophy. The remaining correlation with blood pressure suggests that more could be gained by intensified antihypertensive treatment.     

Modulation of myocardial economy and efficiency in mammalian failing and non-failing myocardium by calcium channel activation and beta-adrenergic stimulation

OBJECTIVE: We investigated the energy-metabolic consequences of positive inotropic stimulation by the calcium channel activator, BAY K 8644, in comparison with isoprenaline, focussing both on the economy of force development and the efficiency of external work. METHODS: In the first instance, heat liberation was measured in isometrically contracting right ventricular papillary muscles from guinea pigs by means of antimony-bismuth thermopiles; in the second instance, external work and myocardial oxygen consumption were analyzed in isolated failing and non-failing working rat hearts. RESULTS: In the guinea pig muscle strip preparations BAY K 8644 (10(-5) M) and isoprenaline (10(-8 M) increased peak developed force from 13.7 +/- 2.7 to 37.6 +/- 14.9 mN/mm2 and from 13.6 +/- 5.2 to 38.8 +/- 3.3 mN/mm2, respectively (P < 0.01). Stress-time integral was increased from 10.3 +/- 3.0 to 34.7 +/- 19.2 mN.s/mm2 by BAY K 8644 and from 9.5 +/- 2.4 to 23.0 +/- 1.6 mN.s/mm2 by isoprenaline. Whereas a significant decrease in the ratio between stress-time integral and initial heat (integral of Pdt/IH) (i.e., economy contraction) was observed for isoprenaline (5.26 +/- 1.91 before and 3.11 +/- 0.72 N.m.s.J-1 after treatment (P < 0.01), BAY K 8644 did not significantly alter this index (5.26 +/- 2.39 before and 6.22 +/- 2.63 N.m.s.J-1 after treatment). Similar results were obtained for the ratio between stress-time integral and tension-dependent heat. Significantly more calcium ions were required for equieffective activation of the contractile proteins with isoprenaline as compared to BAY K 8644. In working preparations of sham-operated and infarcted rat hearts, the increase in myocardial oxygen consumption per minute (delta MVO2) for a given increase in external work per minute (delta P) was significantly higher with isoprenaline than with equipotent concentrations of BAY K 8644 or high calcium. CONCLUSIONS: Inotropic mycardial stimulation by BAY K 8644 is associated with higher economy and efficiency than stimulation by isoprenaline when analyzed both by heat measurements in isometric preparations and by myocardial oxygen consumption in working heart preparations.     

Cardiac and vascular responses of isolated rat tissues treated with diterpenes from Sinularia flexibilis (coelenterata: octocorallia)

The marine environment is a rich source of compounds with cardiovascular activity. This study characterizes the cardiac and vascular responses in isolated rat tissues of flexibilide, dihydroflexibilide and sinulariolide, three diterpenes isolated from the soft coral Sinularia flexibilis. On rat left ventricular papillary muscles, dihydroflexibilide and flexibilide showed similar potencies (-log EC50 = 4.69 +/- 0.05 and 4.66 +/- 0.06, respectively); the maximal response to dihydroflexibilide of 1.4 +/- 0.2 mN was 35 +/- 7% that of calcium chloride in the same muscles. All diterpenes relaxed rat thoracic aortic rings precontracted with KC1 (100 mM) with similar potencies (flexibilide, -log EC50 = 4.17 +/- 0.06). Flexibilide was further characterized and shown to increase force in isolated rat left atria by 0.8 +/- 0.5 mN at 1 x 10(-4) M, to increase rate of contraction in isolated rat right atria by 18 +/- 5 beta/min at 3 x 10(-5) M and to completely relax endothelium-denuded rat thoracic aortic rings (-log EC50 = 4.14 +/- 0.05). Toxicity as indicated by the occurrence of ectopic beats was not observed with the diterpenes at concentrations which produced complete relaxation of blood vessels, maximal positive inotropic activity and minor positive chronotropic responses. Thus, these compounds may be useful lead compounds in the search for improved treatment of cardiovascular disease, especially heart failure.     

Effects of angiotensin II on inotropy and intracellular Ca2+ handling in normal and hypertrophied rat myocardium

The direct inotropic effect of angiotensin II on the myocardium is still controversial and little information exists as to its potential modification by heart disorders. Therefore, this study performed simultaneous measurements of isometric force and intracellular Ca2+ concentrations ([Ca2+]i) in left ventricular papillary muscles from sham-operated and aortic-banded rats at 10 weeks post-surgery. Angiotensin II (10(-6) M) induced a reduction of peak systolic [Ca2+]i (0.56 +/- 0.03 to 0.48 +/- 0.04 microM; P<0.05) and a parallel but insignificant diminution of developed tension (10.5 +/- 1.3 to 9.6 +/- 0.8 mN/mm2) in normal papillary muscles from sham-operated animals. Hypertrophied papillary muscles from aortic-banded rats demonstrated a significant decline in both peak systolic [Ca2+]i (0.51 +/- 0.02 to 0.44 +/- 0.01 microM; P<0.05) and developed tension (8.4 +/- 1.1 to 6.8 +/- 1.7 mN/mm2; P<0.05) after addition of angiotensin II. The time courses of the mechanical contraction and the intracellular Ca2+ signal were prolonged by angiotension II in both groups. Isoproterenol dose-dependently increased developed tension and peak systolic [Ca2+]i in papillary muscles from sham-operated rats. In contrast, the positive inotropic response to isoproterenol was markedly reduced in hypertrophied muscles despite a seemingly unimpaired increase in peak systolic [Ca2+]i. Pretreatment with angiotensin II (10(-6) M) resulted in a significant attenuation of the systolic [Ca2+]i response to isoproterenol stimulation in both normal and hypertrophied papillary muscles. Neither the bradykinin B2 antagonist icatibent (10(-6) M) nor the nitric oxide (NO) inhibitor L-NMMA (10(-6) M) abolished the depressant effects of angiotension II. Thus, ANG II induces a parallel decline of the mechanical performance and Ca2+ availability in rat myocardium. These effects are more distinct in hypertrophied than in normal muscle and become accentuated during beta-adrenergic stimulation. The underlying mechanism is not associated with the NO pathway but might involve a negative functional coupling between the angiotensin and beta-adrenergic-receptor complex.     

A study of microtubules in hypertrophied cardiomyocytes

Microtubules, components of the cytoskeleton, play an important role in the maintenance of cellular shape, in intracellular transport of organelles and membrane vesicles, and in signal transduction in the cell. Studies on the role of microtubules in hypertrophied myocardial cells have been carried out using an acute pressure overloading model in which stenosis was produced in the aorta. Clinically, however cardiac hypertrophy is most frequently caused by a gradual increase in blood pressure due to essential hypertension. We evaluated the role of microtubulas by observing their serial changes in cells from spontaneously hypertensive rats (SHR), which are used as a model of essential hypertension. Blood pressure, body weight, left ventricular weight, and the cell cross-sectional area were measured in SHR and Wistar-Kyoto (WKY) rats aged 4, 6, 10, and 16 weeks (5 rats each). Microtubules in cardiocytes were observed using confocal laser scanning microscopy by the immunofluorescence method against beta-tubulin. Microtubules in cardiocytes run primarily in a longitudinal direction, thinly through the intermyofibrillar spaces and densely around the nuclei. The numbers of microtubules were measured separately in the perinuclear region and the nonperinuclear region, and their total was calculated. The cross-sectional area of a whole cell and nucleus was measured at the level of the nucleus, and microtubule density was calculated by the number of nonperinuclear microtubules in the cytosolic area. Five myocardial cells were randomly selected in each rat, and the mean density (/micron2) was observed. For comparison, similar analysis was done in an acute pressure loading model. This study showed that: 1) In the acute pressure overload model, hypertrophy was immediately observed, and microtubules were increased in 16% of cardiocytes. In SHR, an increase in blood pressure and hypertrophy of hearts were observed at the age of 6 weeks and after. The density of microtubules at the age of 4, 6, 10, and 16 weeks was 0.69 +/- 0.03 (/micron2), 0.73 +/- 0.05 (/micron2), 0.64 +/- 0.02 (/micron2), respectively, in WKY rats, and 0.54 +/- 0.03 (/micron2), 0.54 +/- 0.02 (/micron2), and 0.51 +/- 0.02 (/micron2), respectively, in the SHR, showing a relatively stability in the latter. This suggested that a gradual increase of blood pressure could not be a stimulus that changes the density and distribution of microtubules. 2) The immunofluorescence method against beta-tubulin using confocal laser scanning microscopy seems to be good method for quantitative analysis of microtubules in cardiocytes.     

Effect of enalapril treatment on the heart of normal rats

Previous experiments showed that enalapril (EN) treatment as well as enalaprilic acid, when added to the perfusion bath, diminish the inotropic response of the papillary muscles to isoproterenol (ISO). The main objective of this study was to evaluate, in normal rats, the effect of EN on basal contractility and inotropic response to ISO on the whole perfused ventricles (Langendorff preparation). Blood pressure (BP), increase in body weight (IBW), ventricular weight/body weight ratio (R) and concentration of ventricular proteins and DNA were also analyzed. Five groups were studied: EN10: 5 mg/kg/day, 10 days; EN21(L): 5mg/kg/day, 21 days; EN21(H): 15 mg/kg/day, 21 days. C10 and C21 were untreated controls. Cardiac contractility was evaluated by the maximal developed pressure, maximal rate of rise of pressure and maximal velocity of relaxation; no changes were found due to EN treatments either on basal conditions or on ISO stimulation. Significant differences (p<0.05 vs C21) were: lower BP and R in EN21(L) and EN21(H), slower IBW in EN21(H), decreased ventricular DNA in EN21(H). In conclusion, daily treatment for ten or twenty one days with enalapril does not change either basal cardiac contractile performance or inotropic response to ISO in the Langendorff preparation. Longterm treatment with EN seems to modify nuclear processes involved in cardiomyocite DNA content.     

Oxygen consumption for constant work is minimal at lowest working contractility in normal dog hearts

We tested whether minimal myocardial oxygen consumption (MVO2) for a given external work would exist in the middle of a normal contractility range as previously predicted theoretically. The left ventricle of the excised cross-circulated dog heart preparation was connected to a volume servo pump. Myocardial contractility in terms of ventricular end-systolic elastance (Emax) was gradually increased from control 8.9 +/- 3.4 (mean +/- SD) to 30.0 mmHg/(ml/100 g) by epinephrine and decreased to 1.8 mmHg/(ml/100 g) by propranolol while heart rate, end-systolic pressure and stroke work were kept constant. MVO2 was determined as the product of total coronary flow and coronary arteriovenous oxygen content difference in each contractile state. We plotted MVO2 values against E(max) values in each heart. The MVO2-E(max) relation for a constant cardiac work showed that MVO2 was minimal at the low end of the covered E(max) range. We conclude that minimal MVO2 for a given cardiac work is generally obtained at the lowest working contractility in normal dog hearts. This conclusion might pose some problems in the previous theoretical prediction as to the contractility that achieves the minimal MVO2 in a given external work.     

EMD 53998 acts as Ca(2+)-sensitizer and phosphodiesterase III-inhibitor in human myocardium

The effect of EMD 53998 (EMD) (0.1-100 mumol/l), chemically a racemic thiadiazinone derivative, suggested to be a potent Ca(2+)-sensitizer, was studied in human failing and nonfailing left ventricular myocardium. For comparison, the effects of the pyridazinone derivative pimobendan (0.1-300 mumol/l), isoprenaline (Iso) (0.001-3 mumol/l) as well as CaCl2 (1.8-15 mmol/l Ca2+) were investigated. The positive inotropic responses were examined in electrically driven (1 Hz, 37 degrees C) human left ventricular papillary muscle strips from terminally failing hearts (NYHAIV, n = 24) and nonfailing donor hearts (NF, n = 9). The effect of EMD on the Ca(2+)-sensitivity of skinned fiber preparations from the very same human failing hearts were studied as well. EMD and pimobendan increased force of contraction (FOC) in a concentration-dependent manner. As judged from the EC50-values, EMD increased FOC more potently than pimobendan. EMD was significantly more effective than pimobendan to increase FOC in papillary muscle strips from NYHA IV (EMD: +2.5 +/- 0.1 mN; pimobendan: +0.8 +/- 0.2 mN) as well as from nonfailing hearts (EMD: +3.1 +/- 0.5 mN; pimobendan: +1.2 +/- 0.2 mN). Only in terminally failing myocardium, EMD increased FOC as effectively as Iso. After inotropic stimulation with EMD, pimobendan, or Iso, carbachol (1000 mumol/l) reduced FOC in left ventricular papillary muscle strips, indicating a cAMP-dependent mode of action. In skinned fiber experiments, EMD increased Ca(2+)-sensitivity significantly more (p < 0.01) than pimobendan. In conclusion: EMD increases FOC in human myocardium via sensitizing of the contractile proteins towards Ca2+ and by inhibition of phosphodiesterase III-isoenzymes. EMD is a potent calcium sensitizing agent in human myocardium. Thiadiazinone derivatives could be one step in the evolution to more potent and selective calcium-sensitizers.     

Reduced cardiac contractile responsiveness to isoproterenol in obese rabbits

Although obesity is characterized by increased sympathetic nervous system activity, there is often a paradoxical reduction in cardiovascular end-organ response to sympathetic stimulation. Mechanisms involved in reduced sympathetic responsiveness in obesity have not been well characterized. Therefore, we determined cardiac contractile responsiveness to beta-stimulation in the obese rabbit model using both isolated heart (IH) and isolated papillary muscle (IPM) preparations. Female New Zealand White rabbits were fed control (IH: n=9; IPM: n=6) or 10% fat diets (IH: n=9; IPM: n=7) for 12 weeks. Contractile responsiveness in the IH was determined using a modified Langendorff preparation to evaluate the dose-response relationship between isoproterenol and 1) peak developed pressure/g of left ventricular wet weight and 2) maximal rate of pressure development (+dP/dt/P). Contractile responsiveness in the IPM was determined using right ventricular papillary muscles to evaluate the dose-response relationship between isoproterenol and (1) peak developed tension (T)/mm2 cross-sectional area (CSA) and (2) maximal rate of tension development (dT/dt/CSA). In the IH, baseline and maximum developed pressure/g were reduced in obese rabbits by 37% and 31%, respectively (P< or =.05). In the IPM, baseline and maximum T/CSA responses were reduced in obese rabbits by 59% and 33%, respectively (P< or =.05). Potency of isoproterenol as reflected by the EC50 did not differ between lean and obese animals in either preparation. These results demonstrate that left ventricular contractility in obesity is reduced at baseline and in response to stimulation with isoproterenol and suggest that decreased responsiveness to beta-stimulation may be a factor in the obesity-related systolic dysfunction.     

Cellular uncoupling during ischemia in hypertrophied and failing rabbit ventricular myocardium: effects of preconditioning

BACKGROUND: Patients with heart failure show a very high incidence of arrhythmias and sudden death that is often preceded by ischemia; however, data on electrophysiological changes during ischemia in failing myocardium are sparse. We studied electrical uncoupling during ischemia in normal and failing myocardium. METHODS AND RESULTS: Tissue resistance, intracellular Ca2+ concentration (Indo-1 fluorescence ratio), and mechanical activity were simultaneously determined in arterially perfused right ventricular papillary muscles from 11 normal and 15 failing rabbits. Heart failure was induced by combined volume and pressure overload. Before sustained ischemia, muscles were subjected to control perfusion (non-PC) or ischemic preconditioning (PC). The onset of uncoupling during ischemia was equal in non-PC normal (13.6+/-0.9 minutes of ischemia) and non-PC failing hearts (13.3+/-0.7 minutes of ischemia). PC postponed uncoupling in normal hearts by 10 minutes. In failing hearts, however, PC caused a large variability in the onset of uncoupling during ischemia (mean, 12.2+/-2.1; range, 5 to 22 minutes of ischemia). The duration of uncoupling process was prolonged in failing hearts (12.9+/-0.9 minutes) compared with normal hearts (7.8+/-0.4 minutes). The degree of heart failure and relative heart weight of the failing hearts significantly correlated with the earlier uncoupling after PC and the duration of uncoupling. In every experiment, the start of Ca2+ rise and contracture preceded uncoupling during ischemia. CONCLUSIONS: The duration of the process of ischemia-induced electrical uncoupling in failing hearts is prolonged compared with that in normal hearts. Ischemic PC has detrimental effects in severely failing papillary muscles because it advances the moment of irreversible ischemic damage.