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To identify biological pathways associated with myocardial recovery from heart failure (HF), gene profiling and gene set enrichment analysis (GSEA) were examined in left ventricle of spontaneously hypertensive rats with HF (SHR-F) with no treatment, following treatment with the angiotensin converting enzyme inhibitor captopril, and treatment with captopril combined with the short chain fatty acid derivative phenylbutyrate. Failing hearts demonstrated depressed left ventricular ejection fraction, while ventricular volume and mass increased. Captopril treatment alone prevented further deterioration but did not improve myocardial function; relatively few transcripts were differentially expressed relative to untreated SHR-F. Gene sets identified by GSEA as downregulated with captopril treatment compared to SHR-F group included those related to hypoxia and reactive oxygen species, while upregulated gene sets included G protein signaling. Treatment with phenylbutyrate alone did not improve survival (no animals in this group survived the 30 day treatment period), while phenylbutyrate combined with captopril increased survival and significantly improved cardiac function in vivo and in vitro. Normalized microarray data identified 780 genes that demonstrated a combined treatment effect of which 258 genes were modified with HF. Fatty acid metabolism and ion transport were among the most significantly upregulated pathways in the combined treatment group compared to untreated SHR with HF, whereas those related to oxidative stress, growth, inflammation, protein degradation, and TGF-beta signaling were downregulated. These findings demonstrate improved myocardial function and regression of cardiac hypertrophy, and identify many HF related gene sets altered with phenylbutyrate and captopril treatment, but not captopril alone. These results characterize gene sets associated with recovery from HF, and suggest that phenylbutyrate may be a potentially effective adjunctive treatment, together with captopril, by synergistically modulating pathways that contribute to restoration of contractile function of the failing SHR heart., (Published by Elsevier Ltd.)

Gene expression, determined by micro-array analysis, and left ventricular (LV) remodeling associated with the transition to systolic heart failure (HF) were examined in the spontaneously hypertensive rat (SHR). By combining transcript and gene set enrichment analysis (GSEA) of the LV with assessment of function and structure in age-matched SHR with and without HF, we aimed to better understand the molecular events underlying the onset of hypertensive HF. Failing hearts demonstrated depressed LV ejection fraction, systolic blood pressure, and LV papillary muscle force while LV end-diastolic and systolic volume and ventricular mass increased. 1431 transcripts were differentially expressed between failing and non-failing animals. GSEA identified multiple enriched gene sets, including those involving inflammation, oxidative stress, cell degradation and cell death, as well as TGF-beta and insulin signaling pathways. Our findings support the concept that these pathways and mechanisms may contribute to deterioration of cardiac function and remodeling associated with hypertensive HF., (Published by Elsevier Inc.)

The objective of this study was to determine whether a simple, noninvasive method involving administration of isoproterenol could be used to produce myocardial injury and cardiac dysfunction in the mouse heart with a low incidence of mortality. Adult Swiss-Webster mice were injected with isoproterenol (100 mg/kg SC) once daily for 5 d. Myocardial histology and left ventricular (LV) function were assessed 10 to 14 d after the last isoproterenol injection in 14 surviving isoproterenol-treated mice and 15 saline-treated control mice. Left ventricular systolic and diastolic pressures were evaluated in vitro by means of isovolumically contracting, perfused Langendorff preparations. Isoproterenol induced marked endocardial injury, associated with hypertrophy of surviving myocytes, and an increase in myocardial fibrosis (collagen types I and III according to picrosirius red microscopy). The hearts from isoproterenol-treated mice demonstrated decreased LV compliance, as evidenced by an upward shift in the diastolic pressure-volume relationship, with normal LV systolic function. Isoproterenol administration provides a simple, noninvasive means to induce endocardial injury and diastolic dysfunction without significant impairment of systolic function. This model has a low incidence of mortality and may be useful to assess the effects of gene or stem cell therapy on cardiac dysfunction without the potential confounding effects of invasive procedures.

Thyroid hormone is known to affect myocardial glycogen stores and thereby possibly limit anaerobic performance of mammalian cardiac muscle. Thyroid hormone administration (3,5,3'-triiodo-L-thyroxine, 300 microg/kg/day, sc) for 10 days decreased left ventricle (LV) glycogen concentration relative to euthyroid animals (2.78+/-0.46 vs. 4.28+/-0.29 mg/g of LV (mean+/-SEM)) while increasing the percent of V(1) myosin isozyme, contractile activity and cardiac mass. In contrast, thyroidectomy increased myocardial glycogen stores (8.50+/-0.56 mg/g of LV) and shifted the myosin isozyme toward V(3), prolonged contractile activity and decreased LV mass. Thyroxine administration for 3, 7 and 10 days to thyroidectomized animals progressively decreased contractile duration and increased LV mass. Thyroxine administration for 3 or 7 days to thyroidectomized rats did not reduce glycogen stores (7.75+/-1.02 and 9.62+/-1.16 mg/g of LV, respectively), whereas myocardial glycogen declined to 3.30+/-0.58 mg/g of LV after 10 days of treatment. During hypoxia, cardiac muscle from thyroidectomized rats maintained greater active force and developed less contracture relative to euthyroid and, to a greater extent, than hyperthyroid rats. Removal of glucose from the bath decreased anaerobic performance and impaired recovery; however, myocardium from thyroidectomized rats remained more tolerant to hypoxia than the euthyroid group. Overall, the intrinsic LV glycogen content was positively correlated to anaerobic performance. These data demonstrate that the thyroid state profoundly affects myocardial growth, contractility and anaerobic performance of rat myocardium. Although energy demand may affect function during hypoxia, anaerobic substrate reserve (cardiac glycogen concentration) appears to be the primary factor determining tolerance to hypoxic stress., (2009 Wiley-Liss, Inc)

Background: The effects of long-term oral administration of L-arginine, a substrate for nitric oxide (NO) production, on left ventricular (LV) remodeling, myocardial function and the prevention of heart failure (HF) was compared to the angiotensin-converting enzyme (ACE) inhibitor captopril in a rat model of hypertensive HF (aged spontaneously hypertensive rat (SHR))., Methods: SHRs and age-matched normotensive Wistar-Kyoto (WKY) rats were assigned to either no treatment, treatment with L-arginine (7.5 g/l in drinking water) or captopril (1 g/l in drinking water) beginning at 14 months of age, a time when SHRs exhibit stable compensated hypertrophy with no hemodynamic impairment; animals were studied at 23 months of age or at the time of HF., Results: In untreated SHR, relative to WKY, there was significant LV hypertrophy, myocardial fibrosis, and isolated LV muscle performance and response to isoproterenol (ISO) were depressed; and, 7 of 10 SHRs developed HF. Captopril administration to six SHRs attenuated hypertrophy and prevented impaired inotropic responsiveness to ISO, contractile dysfunction, fibrosis, increased passive stiffness, and HF. In contrast, L-arginine administration to SHR increased LV hypertrophy and myocardial fibrosis while cardiac performance was depressed; and 7 of 9 SHRs developed HF. In WKY, L-arginine treatment but not captopril resulted in increased LV weight and the contractile response to ISO was blunted. Neither L-arginine nor captopril treatment of WKY changed fibrosis and HF did not occur., Conclusion: These data demonstrate that in contrast to captopril, long-term treatment with L-arginine exacerbates age-related cardiac hypertrophy, fibrosis, and did not prevent contractile dysfunction or the development of HF in aging SHR.

Background: Pressure overload in humans follows a chronic and progressive course, often resulting in eventual cardiac decompensation and death. Animal models of heart failure generally fail to mimic the temporal features observed in human disease often covering a major portion of the life span, and findings of short-term studies are of uncertain applicability. The purpose was to determine whether chronic pressure overload introduced gradually in young normotensive rats would lead predictably to heart failure and to characterize specific phenotype features that have been well documented in another model of heart failure., Methods: Rats underwent banding of the ascending aorta at 7 weeks of age such that the hemodynamic load increased gradually with ontogenic growth. Two groups of hypertrophied hearts from aortic-banded rats, with and without signs of heart failure, were compared with those of control rats at a mean age of 11 months., Results: Hearts of aorta-banded rats underwent a transition from stable compensated hypertrophy to heart failure that was characterized by augmented hypertrophy, depressed contractile function, elevated fibrosis, increased myocardial stiffness, and marked alterations in the expression of genes encoding contractile, regulatory, and extracellular matrix proteins., Conclusions: Gradual constriction of the rat aorta resulted in heart failure after a variable length of time (3 to 18 months). Despite differences in genotype, the ultimate phenotype associated with the transition to failure in the aorta-banded rat is nearly identical to that observed in the aged spontaneously hypertensive rat (SHR), with a few notable differences. The findings suggest that a common heart failure phenotype follows long-term pressure overload regardless of the underlying etiology.

Purpose: The objective of the study reported here was to investigate whether massage-like stroking of the thorax and cranial portion of the abdomen might relax unanesthetized rats sufficiently to permit in vivo echocardiography., Methods: Nine-month-old spontaneously hypertensive rats (SHR) were first conditioned to being held by hand for 10 to 15 min twice a day for seven to 10 days. During each session, the animal was placed in supine position, and the thorax and cranial abdominal area were gently stroked (approx. 5 cm/s, 12 to 14 times/min). After the conditioning period, echocardiography was initiated. We obtained serial transthoracic two-dimensional (2-D) and M-mode echocardiograms from nine-month-old SHR that were treated with isoproterenol (60 mg/kg of body weight, s.c., x 1, followed by 30 mg/kg/d x3), and from old (20 to 24 months old) SHR, studied when labored breathing, suggestive of heart failure, was evident (SHR-F). Measurements included end-diastolic volume (EDV) and end-systolic volume (ESV)., Results: In the isoproterenol-treated SHR, mean +/- SD echocardiographically derived EDV (2-D, 0.29 +/- 0.05; M-mode, 0.28 +/- 0.01 ml) was not significantly different from volume at necropsy (0.33 +/- 0.04 ml). Measurements of EDV and ESV by use of M-mode and 2-D echocardiography were significantly correlated (EDV R2 = 0.48, P = 0.05; ESV R2 = 0.39, P = 0.02). Echocardiography revealed pleuropericardial effusions (4/6), atrial thrombi (5/6), and left and right ventricular enlargement (6/6). The EDV and ESV were increased fivefold (P < 0.01) and threefold (P < 0.05), respectively, versus values for SHR not in heart failure (SHR-NF). Left ventricular ejection fraction of hearts from SHR-F was markedly decreased, compared with that in SHR-NF (44 +/- 7 versus 74 +/- 2%, respectively; P < 0.05). The presence or absence of left atrial thrombi and fluid in the thoracic cavity was confirmed at necropsy in SHR-F and SHR-NF., Conclusion: Thoracic massage permits use of echocardiography in unanesthetized rats, thereby providing a simple, non-invasive technique for assessment of cardiac structure and function in rats without the potentially adverse effects of anesthesia.

Background: Paired electrical stimulation and postextrasystolic potentiation (PESP) of contractility has been extensively studied in ventricular myocardium, but less is known about PESP of atrial contractility. Our aim was to determine whether PESP of atrial contractility could augment left ventricular (LV) preload and improve LV systolic performance., Methods and Results: A paired electrical stimulus closely following the pacing stimulus was applied to isolated atrial and ventricular myocardium from 4 dog hearts, and the interval dependent force potentiation was examined. In isolated atrial myocardium, paired pacing increased the active tension from a baseline of 1.36 +/- 0.23 to 2.60 +/- 0.57 g/mm(2); in ventricular myocardium active tension increased from 2.58 +/- 0.42 to 3.81 +/- 0.27 g/mm(2) (both P <.01). Then, LV pressure (micromanometer) and segment length (ultrasonic crystals) were measured in the intact hearts of 7 anesthetized dogs in which premature stimuli were applied to the atrium. In intact hearts, paired pacing of the atrium (coupling interval 200 ms) increased LV end-diastolic pressure from 3.8 +/- 1.0 to 6.4 +/- 1.0 mm Hg; systolic pressure increased from 105 +/- 6 to 112 +/- 7 mm Hg (both P <.05). LV pressure-length loop area (regional stroke work) increased 10.5 +/- 0.2%., Conclusions: Isolated atrial myocardium exhibits substantial PESP of contractility, which is similar to ventricular myocardium. In the intact heart, atrial PESP augments LV systolic performance by effecting an increase in LV preload. This technique may provide a means of improving cardiac performance in patients with heart failure.

Paired pacing has been shown to potentiate contractile function of cardiac muscle, and it has been suggested that this may enhance contractile function of diaphragmatic muscle. The primary goal of this study was to study the effect of paired pacing on potentiation of contractile function of diaphragmatic muscle compared to atrial and ventricular myocardium. Diaphragmatic muscle was isolated from mouse and rat, and atrial and ventricular myocardium from dogs. Potentiation was induced by isolated extrastimuli (equal in duration and intensity to the pacing stimulus) and by repetitive extrastimuli (i.e. paired pacing) at a paced rate of 12, 30 and 60 beats/min. Baseline studies were performed while preparations were isometrically contracting at L(max) in oxygenated Krebs-Henseleit solution at 28 degrees C. Maximal force generation in response to a premature stimulus was determined at each rate by scanning the coupling interval between paced beats. Under baseline conditions, diaphragmatic muscle contracted faster than atrial and ventricular muscle. In all tissues, maximum potentiation (increase in force above baseline) was approximately 100% of baseline force, and peak potentiation occurred at shorter coupling intervals with increasing rates of stimulation. Single and paired pacing of diaphragm potentiated the contraction during which the extrastimuli were introduced, while in cardiac muscle, extrastimuli potentiated the contraction following the extrastimulus. The maximum potentiated response occurred when the extrastimulus was introduced prior to the development of peak force in diaphragmatic muscle. In contrast, in atrial and ventricular muscle, a single or paired premature stimulus potentiated the subsequent beat when delivered late during relaxation. In cardiac muscle, maximal potentiation gradually occurred following several repetitive stimuli. Following cessation of single and paired pacing, the beat following the potentiated response immediately returned to baseline in diaphragmatic muscle, while a gradual decline was evident over several subsequent beats in cardiac muscle. Increasing the bath temperature from 28 to 37 degrees C resulted in a leftward shift in the peak potentiated force vs. coupling interval curve without a decline in the magnitude of potentiated force in diaphragmatic muscle. In diaphragm muscle, exposure to ryanodine markedly decreased baseline force and maximal potentiation. We conclude that closely timed extrastimuli applied to diaphragmatic muscle can potentiate developed force in a given contraction, while in cardiac tissue a delayed stimulus potentiates the subsequent beat. These differences in contractile responsiveness are not due to differences in loading conditions, but appear to reflect intrinsic differences in calcium handling.

The spontaneously hypertensive rat (SHR) is an animal model of genetic hypertension which develops heart failure with aging, similar to man. The consistent pattern of a long period of stable hypertrophy followed by a transition to failure provides a useful model to study mechanisms of heart failure with aging and test treatments at differing phases of the disease process. The transition from compensated hypertrophy to failure is accompanied by changes in cardiac function which are associated with altered active and passive mechanical properties of myocardial tissue; these events define the physiologic basis for cardiac decompensation. In examining the mechanism for myocardial tissue dysfunction, studies have demonstrated a central role for neurohormonal activation, and specifically the renin-angiotensin-aldosterone system. Pharmacologic attenuation of this system at differing points in the course of the process suggests that prevention but not reversal of myocardial tissue dysfunction is possible. The roles of the extracellular matrix, apoptosis, intracellular calcium, beta-adrenergic stimulation, microtubules, and oxygen supply-demand relationships in ultimately mediating myocardial tissue dysfunction are reviewed. Studies suggest that while considerable progress has been made in understanding and treating the transition to failure, our current state of knowledge is limited in scope and we are not yet able to define specific mechanisms responsible for tissue dysfunction. It will be necessary to integrate information on the roles of newly discovered, and as yet undiscovered, genes and pathways to provide a clearer understanding of maladaptive remodeling seen with heart failure. Understanding the mechanism for tissue dysfunction is likely to result in more effective treatments for the prevention and reversal of heart failure with aging. It is anticipated that the SHR model will assist us in reaching these important goals.

Inotropic responsiveness to beta-adrenergic stimulation is generally found to be impaired in left ventricular (LV) hypertrophy and failure. To investigate the mechanisms by which angiotensin-converting enzyme inhibitor therapy may modulate inotropic responsiveness with long-term pressure overload, we studied the effects of captopril treatment on cardiac gene expression, LV muscle mechanical contraction, and intracellular calcium (Ca(2+)) transients from spontaneously hypertensive rats (SHR). LV papillary muscles from untreated SHR, age-matched normotensive Wistar-Kyoto rats (WKY), and SHR treated with captopril (CAP(Rx) started at 12, 18, and 21 months of age) were studied. All animals were studied at 24 months of age or when heart failure developed. In untreated SHR, alpha-myosin heavy chain (MHC) gene expression and protein were decreased, the Ca(2+) transient (with the bioluminescent indicator aequorin) was prolonged, and abundance of Na(+)/Ca(2+) exchanger mRNA levels increased in comparison to WKY. Active stress development at L(max) and the maximum rate of stress development were depressed and contractile duration prolonged in SHR relative to WKY. Isoproterenol administration further decreased active stress in untreated SHR despite an increase in intracellular Ca(2+) levels. In CAP(Rx) SHR, alpha-MHC gene expression and protein levels were increased, the Ca(2+) transient was not prolonged, Na(+)/Ca(2+) exchanger expression was downregulated, and papillary muscle function demonstrated increased active stress and maximum rate of stress development in response to isoproterenol. The increased abundance of alpha-MHC mRNA in conjunction with an increase in V(1) myosin isozyme suggests that captopril affects transcriptional regulation of cardiac gene expression. Restored LV inotropic responsiveness to beta-adrenergic stimulation in CAP(Rx) SHR appears to be coupled to normalization of Na(+)/Ca(2+) exchanger mRNA expression, upregulation of V(1) myosin isozyme levels, and increased speed of contraction.

Aging is associated with an increase in myocardial extracellular matrix components and contractile dysfunction. Transforming growth factor- beta(1)(TGF- beta(1)) has been shown to regulate expression of collagen genes and extracellular matrix component synthesis in the heart, and may contribute to the increase in myocardial fibrosis with aging. Therefore, we examined whether TGF- beta(1)heterozygous mutant mice would exhibit less age-associated myocardial fibrosis than normal mice. Twelve heterozygous TGF- beta(1)(+/-) deficient mice and 26 wild-type controls were examined to determine if there was a difference in development of myocardial fibrosis or mortality at 24 months of age due to the loss of one TGF- beta(1)allele. Animals which survived to 24 months of age were killed, and morphometric and functional studies were performed in isolated perfused hearts and in hearts from 6 month old control mice. Pressure-volume relations of the LV were assessed in the isovolumic (balloon in LV) Langendorff preparation. Eleven of 12 (92%) TGF- beta(1)deficient mice survived to 24 months of age in comparison to 66% (12/18) age-matched controls (P<0.05). Hearts from the 24 month old TGF- beta(1)deficient mice exhibited a decrease in myocardial fibrosis (4+/-1 v. 10+/-1% average LV fibrosis in TGF- beta(1)(+/-) and age-matched controls, respectively (P<0.05) and greater compliance (i.e.,lower LV end-diastolic pressure at a given balloon volume), decreased myocardial stiffness, and shorter contractile duration in comparison to 24-month-old wild-type controls. This suggests that modulation of collagen production and/or degradation by TGF- beta(1)may contribute to changes in myocardial structure and function with age. Thus, loss of one TGF- beta(1)allele appears to ameliorate age associated myocardial fibrosis and improve LV compliance, which may contribute to increased survival over the life span of these mice., (Copyright 2000 Academic Press.)

Genetically altered mice have become an increasingly important tool for the study of mechanisms of cardiac function, and therefore it is vital to characterize the basic contractile properties of the mouse heart. As a first approach to this goal, we first optimized perfusion conditions and characterized the effect of incremental left ventricular balloon inflation on end-diastolic, systolic and developed pressures in the isovolumically-contracting mouse heart. Under constant loading conditions, we determined developed pressure in response to changing perfusate calcium (1.25, 2.5, 3.75 and 5.0 mM) and perfusate temperature (30 and 37 degrees C). We then compared the intrinsic inotropic responsiveness to changes in extracellular calcium of left ventricular myocardium from mouse to that from the rat. In the baseline state (1.25 mM extracellular calcium; [Ca2+]o), both isometric contraction duration and normalized active force at the peak of the active force-length relationship (Lmax) were less in mouse than in rat myocardium. Under isotonic conditions, temporal parameters of shortening and the relative shortening were less in mouse vs rat myocardium. Increasing [Ca2+]o from 1.25 to 2.5 mM markedly increased active isometric force and rate of force development (+dF/dt) in the mouse. However, rat myocardium responded to a lesser extent. Under isotonic conditions, peak shortening and the rate of shortening also increased to a greater extent in mouse relative to rat myocardium. Increasing the bath calcium concentration to 5.0 mM increased isometric force and +dF/dt further in the rat but not the mouse, suggesting that two species operate at different points on the force vs [Ca2+]o relationship. We conclude that mouse myocardium exhibits increased sensitivity to changes in [Ca2+]o within the physiologic range in comparison to rat. These differences do not appear to be due to differences in loading conditions. The data suggest that differences in inotropic responsiveness to calcium may reflect intrinsic differences in myocardial calcium sensitivity between species.

To identify genes that are differentially expressed during the transition from compensated hypertrophy to failure, myocardial mRNA from spontaneously hypertensive rats (SHR) with heart failure (SHR-F) was compared with that from age-matched SHR with compensated hypertrophy (SHR-NF) and normotensive Wistar-Kyoto rats (WKY) by differential display reverse transcriptase-polymerase chain reaction. Characterization of a transcript differentially expressed in SHR-F yielded a cDNA with homology to the extracellular matrix protein osteopontin. Northern analysis showed low levels of osteopontin mRNA in left ventricular myocardium from WKY and SHR-NF but a markedly increased (approximately 10-fold) level in SHR-F. In myocardium from WKY and SHR-NF, in situ hybridization showed only scant osteopontin mRNA, primarily in arteriolar cells. In SHR-F, in situ hybridization revealed abundant expression of osteopontin mRNA, primarily in nonmyocytes in the interstitial and perivascular space. Similar findings for osteopontin protein were observed in the midwall region of myocardium from the SHR-F group. Consistent with the findings in SHR, osteopontin mRNA was minimally increased (approximately 1.9-fold) in left ventricular myocardium from nonfailing aortic-banded rats with pressure-overload hypertrophy but was markedly increased (approximately 8-fold) in banded rats with failure. Treatment with captopril starting before or after the onset of failure in the SHR reduced the increase in left ventricular osteopontin mRNA levels. Thus, osteopontin expression is markedly increased in the heart coincident with the development of heart failure. The source of osteopontin in SHR-F is primarily nonmyocytes, and its induction is inhibited by an angiotensin-converting enzyme inhibitor, suggesting a role for angiotensin II. Given the known biological activities of osteopontin, including cell adhesion and regulation of inducible nitric oxide synthase gene expression, these data suggest that it could play a role in the pathophysiology of heart failure.

A simplified transthoracic procedure using electrocauterization was used to induce myocardial injury in mice. After a single small incision through the skin and dissection of the underlying musculature, a modified electrocautery probe consisting of an insulated 20-gauge blunt needle with a polyethylene sleeve was inserted through the interior intercostal muscle at the fourth intercostal space and positioned on the anterior surface of the heart. The placement of the probe on the heart was indicated by mechanical motion of the exterior section of the needle. Electrocautery was applied to the distal exposed end of the probe. Of 10 mice that underwent this procedure, nine survived. After 10 days, myocardial damage was assessed by visual and histologic examination of the heart. In eight of nine surviving mice, transmural injury was induced in the left ventricle. The region of myocardial tissue damage on the surface of the left ventricle was 4.6 +/- 0.5 mm in diameter. This method provides a simple, noninvasive technique using a transthoracic electrocautery procedure to induce myocardial injury in the mouse heart with a low incidence of postoperative mortality.

Background: After a period of stable hypertrophy, male spontaneously hypertensive rats (SHR) develop heart failure between 18 to 24 months of age, with depression of active myocardial function and increased passive stiffness. We tested the hypothesis that chronic ACE inhibition by captopril would prevent and possibly reverse impairment of myocardial function., Methods and Results: Male SHR and normotensive Wistar-Kyoto rats (WKY) were assigned to no treatment or captopril treatment (2 g/L in drinking water) begun at ages 12, 18, and 21 months; animals were studied at 24 months of age, or earlier when evidence of heart failure was found in SHR (mean age, 19+/-2 months). In an additional group, captopril treatment was begun when SHR developed heart failure; surviving animals were studied at 24 months of age. In untreated SHR, relative to WKY, isometric stress development at Lmax, maximum rate of stress development, and shortening velocity were depressed, whereas passive stiffness was increased, in association with the development of myocardial fibrosis. In the SHR treated before cardiac dysfunction, captopril administration attenuated hypertrophy and prevented contractile dysfunction, fibrosis, and increased passive stiffness. Captopril treatment begun after cardiac function was impaired reduced left ventricular hypertrophy but did not restore intrinsic contractile function or reduce fibrosis or passive stiffness., Conclusions: In the male SHR, early treatment with captopril was associated with the most marked attenuation of dysfunction relative to the untreated SHR. Treatment initiated after the onset of heart failure improved clinical signs of heart failure and decreased left ventricular hypertrophy in surviving animals but did not reverse the fibrosis and contractile dysfunction associated with heart failure.

The spontaneously hypertensive rat (SHR) exhibits a transition from stable compensated left ventricular (LV) hypertrophy to heart failure (HF) at a mean age of 21 months that is characterized by a decrease in alpha-myosin heavy chain (alpha-MHC) gene expression and increases in the expression of the atrial natriuretic factor (ANF), pro-alpha1(III) collagen, and transforming growth factor beta1 (TGF-beta1) genes. We tested the hypotheses that angiotensin-converting enzyme inhibition (ACEI) in SHR would prevent and reverse HF-associated changes in gene expression when administered prior to and after the onset of HF, respectively. We also investigated the effect of ACEI on circulating and cardiac components of the renin-angiotensin system. ACEI (captopril 2 g/L in the drinking water) was initiated at 12, 18, and 21 months of age in SHR without HF and in SHR with HF. Results were compared with those of age-matched normotensive Wistar-Kyoto (WKY) rats, and to untreated SHR with and without evidence of HF. ACEI initiated prior to failure prevented the changes in alpha-MHC, ANF, pro-alpha1(III) collagen, and TGF-beta1 gene expression that are associated with the transition to HF. ACEI initiated after the onset of HF lowered levels of TGF-beta1 mRNA by 50% (P<.05) and elevated levels of alpha-MHC mRNA two- to threefold (P<.05). Circulating levels of renin and angiotensin I were elevated four- to sixfold by ACEI, but surprisingly, plasma levels of angiotensin II were not reduced. ACEI increased LV renin mRNA levels in WKY and SHR by two- to threefold but did not influence LV levels of angiotensinogen mRNA. The results suggest that the anti-HF benefits of ACEI in SHR may be mediated, at least in part, by effects on the expression of specific genes, including those encoding alpha-MHC, ANF, TGF-beta1, pro-alpha1(III) collagen, and renin-angiotensin system components.

Spontaneously hypertensive rats (SHR) commonly develop impairment of myocardial function between ages 18-24 months. Isolated muscle studies demonstrate depressed myocardial contractility and increased passive stiffness. Studies of the extracellular matrix in SHR with failure (SHR-F) demonstrate an increased expression of genes encoding extracellular matrix components (ECM), hydroxyproline concentration and fibrosis relative to age-matched non-failing animals. In the present study, tissue sections of hearts from SHR-F, non-failing SHR (SHR-NF) and non-hypertensive Wistar Kyoto rats (WKY) were hybridized with a cDNA probe for alpha1(I) collagen mRNA, which was found by Northern blot analysis to be elevated in SHR-F relative to hearts from control animals. In situ hybridization studies demonstrate increased perivascular and interstitial collagen alpha1(I) gene expression in myocardium from the SHR relative to WKY. In addition, failing hearts from the SHR demonstrate focal alpha1(I) collagen mRNA accumulation in the endocardium and at sites of degenerating single myocardial cells., (Copyright 1997 Academic Press Limited.)

Colchicine has been demonstrated to suppress the release of fibroblast growth factors, retard collagen formation and augment collagenase activity. Trials with colchicine in patients with hepatic fibrosis have suggested clinical benefit. The development of impaired myocardial function in the spontaneously hypertensive rat (SHR) is associated with a marked increase in myocardial fibrosis. The present study was carried out to test the hypothesis that chronic colchicine administration to the SHR would prevent the development of fibrosis and impaired myocardial performance. Colchicine (1 mg/l drinking water) was administered to male SHR and WKY rats from at age 13 months until 24 months or until evidence of heart failure was observed. Age-matched untreated SHR and colchicine treated and untreated WKY served as controls. At study, active and passive properties of isolated left ventricular muscle preparations were determined. Myocardial fibrosis was assessed by measuring hydroxyproline and histologic determination of interstitial cross-sectional area. Increases in LV hydroxyproline and interstitial area were found in untreated SHR relative to WKY; passive myocardial stiffness was increased and active muscle properties were depressed. In comparing colchicine treated vs untreated SHR, no differences in hydroxyproline, interstitial area or intrinsic myocardial function were found. In the WKY, colchicine increased myocardial interstitium and passive stiffness without changing hydroxyproline. Active myocardial function was not depressed. Thus, chronic colchicine administration neither attenuated the development of interstitial fibrosis nor prevented impaired myocardial function in the SHR. Colchicine treatment was associated with increased interstitium in WKY with increased passive myocardial stiffness.

The effects of treppe on left ventricular function in the isolated mouse heart perfused with physiological buffer or with erythrocyte-rich buffer were compared. Left ventricular systolic and diastolic pressures were measured in the isovolumically contracting (balloon in the left ventricle) mouse hearts. Hearts were isolated from 12 adult Swiss-Webster mice and perfused at constant pressure (approximately 85 mmHg) via the aorta. Perfusate consisted of non-recirculating oxygenated Krebs-Henseleit (KH) solution without or with washed cow red blood cells at a hematocrit of 20% (KH-RBC20). The measured ionized calcium concentration of the perfusates were adjusted to 2.2 mmol/l and the temperature held constant at 37 degrees C. Left ventricular systolic pressure, its derivative and diastolic pressures were recorded via a pressure transducer attached to a small latex balloon which was placed in the left ventricle through a left atrial incision. The balloon volume was adjusted to achieve an end-diastolic pressure of 4-8 mmHg. Left ventricular (LV) developed pressure averaged 111 +/- 4 (mean +/- S.E.M.) with KH alone and 108 +/- 4 mmHg with KH-RBC20 while the coronary flows were 3.1 +/- 0.18 and 0.95 +/- 0.15 ml/min respectively. In both KH solution alone and KH-RBC20, developed pressure remained relatively stable from 3 to 5 Hz while +/- dp/dt increased approximately 10% above values observed at 3 Hz. During KH perfusion with increasing stimulation rates, left ventricular pressure and +/- dP/dt, to a lesser extent, decreased while end-diastolic pressure markedly increased at stimulation rates higher than 5 Hz. However, KH-RBC20 perfusion prevented the marked increase in diastolic pressure with increasing stimulation rates (from 5 to 10 Hz). No significant difference in left ventricular developed pressure or +/dP/dt response to treppe were in evidence between groups. These results demonstrate that diastolic function of the isovolumically contracting mouse heart is sensitive to treppe and different techniques of perfusion. Buffer perfusion alone may limit accurate measurement of left ventricular diastolic properties and exacerbate changes in diastolic function, particularly under conditions of increased oxygen demand. The erythrocyte perfused mouse heart provides an in vitro model for determining cardiac function which is physiologically superior to buffer perfusion, and may be useful to investigators to assess gene influence on left ventricular function in genetically altered mice.

Inotropic responsiveness to beta-adrenergic stimulation is generally found to be depressed in cardiac hypertrophy and failure. To investigate whether inotropic responsiveness is associated with alterations in beta-adrenergic receptors in spontaneously hypertensive rats (SHR), we studied left ventricular myocardial contractile responses to isoproterenol and beta-adrenergic receptor density and affinity in age-matched rats (18 to 24 months), including SHR without heart failure, SHR with evidence of heart failure, and normotensive control Wistar-Kyoto rats (WKY). In the baseline state, papillary muscles from failing SHR demonstrated decreased isometric tension development and a reduction in maximal rate of tension development relative to normotensive WKY and compensated SHR. Compared with WKY, beta-adrenergic receptor density of the left ventricle was unchanged in nonfailing SHR and increased in failing SHR (P < .05 versus WKY and nonfailing SHR), and beta-adrenergic receptor affinity did not differ among groups. In the right ventricle, beta-adrenergic receptor density was decreased in failing SHR relative to WKY and nonfailing SHR, and beta-adrenergic receptor affinity was not different among groups. Muscle preparations did not exhibit a positive inotropic response to 10(-8) to 10(-5) mol/L isoproterenol or 6.3 mumol/L forskolin in either failing or nonfailing SHR, whereas a positive inotropic response to both drugs was observed in the normotensive WKY. The lusitropic response to isoproterenol and forskolin was intact and similar in both SHR groups and WKY. The findings suggest that in the SHR model of heart failure, impaired intrinsic left ventricular myocardial function and depressed inotropic responsiveness to beta-adrenergic stimulation are not associated with downregulation of the beta-adrenergic receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Objective: The aim was to test the hypothesis that reperfusion induced arrhythmias are associated with major alterations in intracellular calcium ([Ca2+]i) regulation., Methods: Intracellular calcium, epicardial electrical potentials, and isovolumetric left ventricular pressure were simultaneously recorded in isolated perfused intact rat hearts during ischaemia (10 min) and reperfusion. [Ca2+]i was measured using the bioluminescent calcium indicator aequorin., Results: Neither ventricular tachycardia nor ventricular fibrillation occurred during ischaemia. However, during the first minute of reperfusion ventricular tachycardia or fibrillation were frequently observed. Cellular calcium was altered by varying the perfusate calcium ([Ca2+]o; 0.5, 1.0, and 3.0 mmol.litre-1). 0% (0/6), 50% (5/10), 91% (10/11), respectively, of hearts showed ventricular tachycardia, ventricular fibrillation, or both upon reperfusion (P < 0.001, 0.5 v 3.0 mmol.litre-1). At all [Ca2+]o values examined, early ischaemia was associated with a rapid decrease in developed pressure and transient increase in the peak calcium transient followed by a gradual decline and subsequent increase in diastolic calcium during late ischaemia. The initiation of ventricular tachycardia/fibrillation upon reperfusion was immediately preceded by large increases in the amplitude of the calcium transient. These increases in systolic calcium were not seen in hearts in which ventricular arrhythmias did not occur., Conclusions: The association between reperfusion induced abrupt increases in peak calcium and the occurrence of ventricular tachycardia or fibrillation suggests that intracellular calcium transients may have a significant role in initiating these ventricular arrhythmias.

Background: Fibrosis is commonly found in association with cardiac hypertrophy and failure, but the relation of the connective tissue response to the development of impaired cardiac function remains unclear. We examined passive myocardial stiffness, active contractile function, and fibrosis in the spontaneously hypertensive rat (SHR), a model of chronic pressure overload in which impaired cardiac function follows a long period of stable hypertrophy., Methods and Results: We studied the passive and active mechanical properties of left ventricular (LV) papillary muscles isolated from normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR) at the ages of 12 months and 20 to 23 months. Seven of 15 SHR between 20 and 23 months of age had findings consistent with heart failure (SHR-F). In comparison to preparations from WKY rats and nonfailing SHR (SHR-NF), papillary muscles from the SHR-F group demonstrated increased passive stiffness (central segment exponential stiffness constant, kcs: SHR-F 95.6 +/- 19.8, SHR-NF 42.1 +/- 9.7, WKY rats 39.5 +/- 9.5 (mean +/- SD); SHR-F P < .01 versus SHR-NF, WKY rats). The increase in stiffness was associated with an increase in LV collagen concentration (SHR-F 8.71 +/- 3.14, SHR-NF 5.83 +/- 1.20, WKY rats 4.78 +/- 0.70 mg hydroxyproline/g dry LV wt; SHR-F P < .01 versus SHR-NF, WKY rats); an increase in interstitial fibrosis, as determined histologically (SHR-F 13.5 +/- 8.0%, SHR-NF 4.9 +/- 2.1%, WKY rats 3.6 +/- 0.8%; SHR-F P < .01 versus SHR-NF, WKY rats); and impaired tension development (SHR-F 3.18 +/- 1.27, SHR-NF 4.41 +/- 1.04, WKY rats 4.64 +/- 0.85 kdyne/mm2; SHR-F P < .05 versus SHR-NF; P < .01 versus WKY rats)., Conclusions: The development of heart failure in the aging SHR is associated with marked myocardial fibrosis, increased passive stiffness, and impaired contractile function relative to age-matched nonfailing SHR and nonhypertensive control animals. These data suggest that fibrosis or events underlying the connective tissue response are important in the transition from compensated hypertrophy to failure in the SHR.

Studies of hemodynamics and intrinsic left ventricular myocardial function are carried out to investigate the transition from stable hypertrophy to cardiac decompensation in the aging (18-24 months) spontaneously hypertensive rat (SHR). Echocardiographic data in awake animals demonstrate increased end-diastolic and end-systolic volumes and depressed ejection fractions in left ventricles from SHR with failure (SHR-F) as compared to age matched hypertensive (SHR-NF) and non-hypertensive control animals (WKY). Cardiac catheterization data in anesthetized animals demonstrate depression of both systolic pressure and +dP/dt, and elevated end-diastolic pressure in the SHR-F relative to the two control groups. Since loading conditions and altered demand states may contribute to altered ventricular function, studies of isolated perfused hearts were carried out which demonstrate impaired systolic stress development in the SHR-F group under conditions in which loading conditions are controlled; in addition, it is observed that increasing perfusion pressure by 30 mm Hg has little effect on function. Depression of systolic function and increases in passive stiffness of isolated muscle preparations from the SHR-F indicate impairment of systolic and diastolic function at the tissue level. While all of the preparations studied have potential shortcomings, an integration of findings from these complementary approaches supports the conclusion that heart failure develops in the aging SHR. Furthermore, these data suggest that impaired function is due to changes in the intrinsic properties of the myocardium and that the connective tissue response may play an important role. These studies, in conjunction with the findings of others who have studied the aging SHR, provide support for the use of the aging SHR as a model of the transition from compensated hypertrophy to failure.

We studied functional and intracellular calcium responses to treppe and extracellular calcium in spontaneously hypertensive rat (SHR) hearts during the transition from compensated pressure overload to failure. Intracellular calcium was measured using aequorin, a bioluminescent Ca2+ indicator. Experiments were performed with intact, isovolumically contracting, buffer-perfused hearts from three rat groups: (1) aging SHR with evidence of heart failure (SHR-F), (2) age-matched SHR with no evidence of heart failure (SHR-NF), and (3) age-matched normotensive Wistar-Kyoto (WKY) rats. In each experiment, left ventricular pressure and intracellular calcium transients were simultaneously recorded. Hearts were studied at 30 degrees C and paced at a rate of 1.6 Hz while being perfused with oxygenated Krebs-Henseleit solution (95% O2/5% CO2) at 100 mm Hg. At the baseline state, peak systolic pressure was greatest in the SHR-NF group and lowest in the SHR-F group. Peak and resting [Ca2+]i were not significantly different among groups; however, the calcium transient was prolonged in the SHR-NF and SHR-F groups. With increasing perfusate [Ca2+]o from 0.5 to 3.0 mmol/L, the relative increases in peak [Ca2+]i and peak systolic pressure were similar among groups. When stimulation rate was increased from 1.6 to 2.0, 2.4, 2.8, and 3.2 Hz, peak [Ca2+]i, peak systolic pressure, and +/- dP/dt fell in SHR-F hearts. Peak systolic pressure decreased in the SHR-NF group at rates above 2.4 Hz but did not decline in the WKY group. Peak [Ca2+]i increased in the WKY and SHR-NF groups with increasing heart rates. Peak systolic pressure did not fall significantly in the WKY group at any heart rate. Elevation of diastolic [Ca2+]i and/or calcium transient and pressure alternans were present in 8 of 13 SHR-F hearts at the highest stimulation rate, findings that were absent in both the WKY and SHR-NF hearts. We conclude the following: (1) Under baseline conditions, depressed contractile function of failing myocardium cannot be explained by decreased peak [Ca2+]i, (2) relative increases in [Ca2+]i and inotropy with increasing [Ca2+]o are proportional among groups; and (3) although peak systolic [Ca2+]i and inotropy are maintained with increasing stimulation rate in the WKY and SHR-NF groups, peak systolic [Ca2+]i and pressure decrease in parallel in the SHR-F heart with increasing stimulation rate, suggesting that impaired calcium cycling may contribute to compromised pump function in the SHR-F heart.

To examine the mechanisms by which thyroid hormone modulates the inotropic state of rat myocardium, we studied the effects of thyroid state on isolated rat left ventricular papillary muscle function and intracellular calcium transients in the baseline state and in response to calcium and isoproterenol. Marked differences in contractile state of papillary muscles from hypothyroid and thyroid hormone-treated rats seen under baseline conditions (1.0 mM bath calcium, 30 degrees C, stimulation rate 12/min) do not appear to be due to differences in intracellular calcium concentration ([Ca2+]i) or to changes in myofilament calcium sensitivity but correlate with shifts in myosin isozyme distribution. In response to superimposed inotropic interventions (calcium, 0.625-5.0 mM, or isoproterenol, 10(-8)-10(-6) M), myocardial thyroid state modulates peak [Ca2+]i and inotropy, both of which are increased in thyroid hormone-treated relative to hypothyroid myocardium. The change in inotropy appears to be proportional to peak [Ca2+]i, whether mediated directly by calcium or as a result of beta-adrenergic stimulation. Thus, whereas baseline differences between hypothyroid and thyroid hormone-treated myocardium appear to be due to differences in myosin isozymes and presumed changes in adenosinetriphosphatase activity and cross-bridge cycling, superimposed inotropic responses appear to be mediated by changes in [Ca2+]i.

The failing heart is characterized by impaired cardiac muscle function and increased interstitial fibrosis. Our purpose was to determine whether the functional impairment of the failing heart is associated with changes in levels of mRNA encoding proteins that modulate parameters of contraction and relaxation and whether the increased fibrosis observed in the failing heart is related to elevated expression of genes encoding extracellular matrix components. We studied hearts of 18- to 24-month-old spontaneously hypertensive rats with signs and symptoms of heart failure (SHR-F) or without evidence of failure (SHR-NF) and of age-matched normotensive Wistar-Kyoto (WKY) rats. Compared with WKY rats, SHR-NF exhibited left ventricular (LV) hypertrophy (2.2-fold) and right ventricular (RV) hypertrophy (1.5-fold), whereas SHR-F were characterized by comparable LV hypertrophy (2.1-fold) and augmented RV hypertrophy (2.4-fold; all P < .01). Total RNA was isolated from ventricles and subjected to Northern blot analysis. In SHR-F hearts, the level of alpha-myosin heavy chain mRNA was decreased in both ventricles to 1/3 and 1/5 of the SHR-NF and WKY values, respectively (both P < .01). Levels of beta-myosin heavy chain, alpha-cardiac actin, and myosin light chain-2 mRNAs were not significantly altered in hearts of SHR-NF or SHR-F. Levels of alpha-skeletal actin were twofold greater in SHR-NF hearts compared with WKY hearts and were intermediate in SHR-F hearts. Levels of atrial natriuretic factor (ANF) mRNA were elevated threefold in the LV of SHR-NF (P < .05) but were not significantly increased in the RV of SHR-NF compared with WKY rats. During the transition to failure (SHR-F versus SHR-NF), ANF mRNA levels increased an additional 1.6-fold in the LV and were elevated 4.7-fold in the RV (both P < .05). Levels of sarcoplasmic reticulum Ca(2+)-ATPase (SRCA) mRNA were maintained in the LV of hypertensive and failing hearts at levels not significantly different from WKY values. In contrast, the level of RV SRCA mRNA was 24% less in SHR-NF compared with WKY rats, and during the transition to failure, this difference was not significantly exacerbated (29% less than the WKY value). The levels of fibronectin and pro-alpha 1(I) and pro-alpha 1(III) collagen mRNAs were not significantly elevated in either ventricle of the SHR-NF group but were fourfold to fivefold higher in both ventricles of SHR-F (all P < .05).(ABSTRACT TRUNCATED AT 400 WORDS)

Objective: The aim was to compare the effects of simulated ischaemia-reperfusion with those of hypoxia-reoxygenation in isolated muscle preparations from the ferret right ventricle., Methods: Ischaemia was simulated using fluorocarbon immersion plus hypoxia. Intracellular calcium transients were determined from aequorin luminescence during isometric contractions., Results: Hypoxia in fluorocarbon and physiological saline solution resulted in a similar reversible depression of the peak of the calcium transient. Peak active tension, however, was more depressed in fluorocarbon than in physiological salt solution. The dissociation between peak light and peak active tension was most pronounced on reoxygenation, with near complete recovery of peak light, while there was little recovery of tension in fluorocarbon. When fluorocarbon was then replaced by physiological salt solution, peak active tension recovered promptly. A prolongation of the decay of the calcium transient was seen in both fluorocarbon and physiological salt solution during hypoxia, which shortened promptly on reoxygenation. The time to the peak of the calcium transient was most prolonged during hypoxia in fluorocarbon and there was gradual recovery on reoxygenation., Conclusions: While some changes in the calcium transient during simulated ischaemia are rapidly reversible with reoxygenation (in fluorocarbon), suggesting an effect of hypoxia, others are incompletely reversed or only reversed with physiological salt solution, suggesting an effect of metabolite accumulation. The pronounced dissociation between peak light and peak active tension during reoxygenation in fluorocarbon is promptly reversed by changing to physiological salt solution, suggesting that metabolite retention in the postischaemic period may contribute to depressed myocardial function after reperfusion.

1. To determine whether diltiazem protects the hypoxic myocardium by reducing contractile work, we have compared the effects of diltiazem and quiescence on left ventricular (LV) papillary muscle subjected to hypoxia. Papillary muscles were obtained from male Charles River CD rats weighing 150-250 g. 2. Four groups of muscles were studied: control (N = 6), non-stimulation (N = 10), diltiazem 10(-4) M (N = 6) and diltiazem 10(-4) M plus non-stimulation (N = 10). 3. Isolated rat LV papillary muscles were studied in Krebs-Henseleit solution with a calcium concentration of 2.52 mM at 28 degrees C while contracting isometrically at a stimulation rate of 0.2 Hz. Resting tension and active isometric tension were measured. 4. Both diltiazem and quiescence significantly attenuated contracture tension during hypoxia (0.91 +/- 0.10 vs 2.26 +/- 0.49 g/mm2 for diltiazem vs control, and 0.55 +/- 0.18 vs 2.26 +/- 0.49 g/mm2 for quiescence vs control). Recovery of active tension was improved in the diltiazem groups during reoxygenation (4.16 +/- 0.42 vs 3.75 +/- 0.51, 3.53 +/- 0.15 vs 2.90 +/- 0.13, 5.84 +/- 0.33 vs 6.48 +/- 0.29 and 5.98 +/- 0.90 vs 7.67 +/- 0.68 g/mm2 for diltiazem, diltiazem non-stimulation, non-stimulation and control groups). 5. The results suggest that the protective effect of diltiazem during hypoxia was due to the reduction in energy demand of the myocardium.

Left ventricular isovolumic stress development and metabolic parameters were studied in 18-24-month-old spontaneously hypertensive rats (SHRs) and age-matched Wistar-Kyoto (WKY) rat controls using the isolated, isovolumic (balloon in left ventricle) buffer-perfused rat heart preparation. After WKY rats and all SHRs were compared, SHRs were divided into two groups: those animals with (SHR-F) and without (SHR-NF) evidence of heart failure. Hearts were perfused at 100 mm Hg using a constant pressure system at a temperature of 37 degrees C. In the baseline state, peak systolic pressure was greatest in the SHR-NF group and lowest in the SHR-F group. Peak midwall stress was greatest in the WKY group and, again, lowest in the SHR-F group. Oxygen consumption was lowest in the SHR-F group. When the oxygen cost of stress development was estimated by normalizing myocardial oxygen consumption by peak developed midwall stress, values were lowest in the WKY, greater in the SHR-NF, and greatest in the SHR-F group. Lactate production did not occur in the baseline state in any of the groups. Functional and metabolic responses to graded hypoxia, induced by changing the gas mixture of the perfusate from 95% to 50%, 25%, and 0% oxygen at perfusion pressures of 100 and 130 mm Hg, were studied. Increasing perfusion pressure generally resulted in small increases in peak systolic pressure and myocardial oxygen consumption but did not substantially reverse the contractile or metabolic deficit present in the SHR-F group.(ABSTRACT TRUNCATED AT 250 WORDS)

To investigate the mechanism of impaired myocardial function after long-term pressure overload, we studied cardiac muscle mechanical contraction and intracellular calcium transients using the bioluminescent indicator aequorin. Left ventricular papillary muscle preparations were examined from three groups of rats: 1) aging spontaneously hypertensive rats (SHR) with clinical and pathological evidence suggesting heart failure (SHR-F group), 2) age-matched SHRs with no evidence of heart failure (SHR-NF group), and 3) age-matched normotensive Wistar-Kyoto rats (WKY group). Isometric force development was depressed in both SHR groups relative to the WKY group. Resting [Ca2+]i was lower in the SHR-F group, and the time to peak [Ca2+]i was prolonged in this group. The relative increases in peak [Ca2+]i with the inotropic interventions of increased [Ca2+]o and the addition of isoproterenol were similar among groups. Although inotropy increased in all groups with increased [Ca2+]o, after isoproterenol, inotropy increased only in the WKY group. Thus, in SHR myocardium, [Ca2+]i increased after isoproterenol, but inotropy failed to increase. Myosin isozymes were shifted toward the V3 isoform in both SHR groups; the V3 isoform was virtually 100% in papillary muscles from the SHR-F group. These changes may reflect events directly contributing to the development of heart failure or represent adaptive changes to chronic pressure overload and heart failure.

We have observed that many spontaneously hypertensive rats (SHR) between the ages of 18 and 24 mo develop findings suggestive of heart failure, including pleural and pericardial effusions, left atrial thrombi, and right ventricular hypertrophy. Isolated left ventricular papillary muscle function was studied in these animals (SHR-F), in age-matched SHRs without evidence of heart failure (SHR-NF), and in nonhypertensive controls (WKY). Preparations from SHR-F showed depression of active tension development (3.58 +/- 1.75 g/mm2, means +/- SD) compared with both SHR-NF (7.17 +/- 0.94) and WKY (6.17 +/- 1.00) (P less than 0.01). Shortening velocity was also depressed in SHR-F (0.95 +/- 0.38 lengths/s) compared with SHR-NF (1.60 +/- 0.30; P less than 0.05) and WKY groups (2.15 %/- 0.48; P less than 0.01). Depression of muscle function was not found before 18 mo of age. Thus the aging SHR is a model in which one can observe the transition from chronic stable left ventricular hypertrophy to overt heart failure. Furthermore, left ventricular papillary muscles from SHRs with heart failure show evidence of significant contractile dysfunction, suggesting that impairment of intrinsic myocardial function underlies the development of heart failure.

We studied calcium responsiveness of skinned muscle preparations from the right and left ventricles of rats with cardiac hypertrophy and cardiac hypertrophy plus failure. To test the hypothesis that differences in contractile function are due to changes in myofilament calcium responsiveness, we compared preparations from spontaneously hypertensive rats with cardiac failure, spontaneously hypertensive rats without cardiac failure, and age-matched normotensive Wistar-Kyoto control rats 18-24 months of age. Rats with failure had pleural/pericardial effusions, left atrial thrombi, and right and left ventricular hypertrophy. Muscles were skinned by saponin (250 micrograms/ml) and activated with a series of calcium buffers. Data were plotted as pCa (-log[Ca2+]) versus isometric force and then fit to a modified Hill equation. Values for 50% maximal activation (calcium sensitivity), maximal calcium-activated force, and the slope of the calcium-force relation were compared. Our data indicate that with the development of hypertrophy, calcium sensitivity of left ventricular muscles remains unaffected, but maximal calcium-activated force is increased. In contrast, maximal calcium-activated force declines toward control levels with the development of left ventricular failure, despite the continued presence of significant hypertrophy. In the normotensive rats, the left ventricle is more sensitive to calcium than the right ventricle (pCa50 = 6.0 +/- 0.05 versus 5.7 +/- 0.09; p less than 0.05); however, both the calcium sensitivity and maximal calcium-activated force of the right ventricle increase with the development of compensatory hypertrophy secondary to left ventricular failure. These changes that occur in rats with cardiac hypertrophy and failure may represent important physiological adaptive mechanisms.

The effects of the radiographic contrast agents Renografin, Isopaque, and Hypaque on the mechanical performance of isometrically contracting rat papillary muscle was compared to a new contrast agent, Amipaque. It was found that exposure to Amipaque resulted in significantly less depression of contractile activity than any of the other agents. Changes in mechanical performance associated with exposure to Renografin, Isopaque, or Hypaque could not be attributed to either hypocalcemia or hypoxia. The decrease in developed tension and increase in resting tension was similar to that produced by Krebs-Henseleit containing hypertonic (1420 mmol) glucose or sucrose. Thus, the data support the concept that hyperosmolarity of the radiographic contrast agents may be primarily responsible for the adverse effects on mechanical performance of the myocardium associated with angiography.

Myocardial mechanics and high-energy phosphate content [ATP and creatine phosphate (CrP)] of isolated left ventricular papillary muscle preparations from male spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY) were compared at 6 and 18 mo of age. In comparison with oxygenated (95% O2-5% CO2) glucose-supplied (5.5 mM) papillary muscles from hearts of WKY rats, papillary muscles from hypertrophied hearts of the 18-mo-old SHR exhibited a prolonged time to peak tension, electromechanical delay time, and an increase in resting tension measured at the apex of the length-tension curve. Adenine nucleotide (ATP and ADP) contents of oxygenated papillary muscles were not significantly different between SHR and WKY strains at 6 or 18 mo of age, but CrP content of hearts from adult WKY and SHR were higher than for aged WKY and SHR rats. For up to 30 min of hypoxia (95% N2-5% CO2), muscles from the 18-mo-old SHR and WKY rats demonstrated improved tolerance to hypoxia compared with muscles from younger animals. However, at 60 min of hypoxia the 18-mo-old SHR demonstrated lower active tension and adenylate energy charge [(1/2 ADP + ATP)/(ATP + ADP + AMP)]. At higher glucose concentrations (22 mM), both 18-mo-old WKY and SHR demonstrated improved tolerance to hypoxia; moreover, the differences between strains were no longer evident. Following reoxygenation with 5.5 mM glucose, contracture tension and CrP content recovered to near prehypoxic control levels, whereas developed tension and ATP content remained moderately depressed for all groups.(ABSTRACT TRUNCATED AT 250 WORDS)

To study ventricular cellular volume regulation when cell membranes and ion pumps cannot prevent swelling, rat ventricular sections were incubated in modified Krebs-Henseleit solutions in which 1) potassium was substituted for sodium, ion for ion; or 2) sodium chloride was reduced to decrease osmolarity to 228, 171, or 114 mosM. Ventricular water, [3H]inulin and [3H]mannitol spaces, potassium, sodium, chloride, and protein contents, and resting transmembrane potentials were measured. Increases in ventricular cellular volume were less than 30% in potassium-substituted and extremely dilute media (114 mosM), in contrast to increases of over 100% in identically treated renal cortical slices. In potassium-substituted solution, the fluid gained by ventricular cells during incubation was hypertonic with respect to the bathing medium. In dilute solution (171 and 114 mosM), ventricular, cellular, and extracellular osmolarities equilibrated only after substantial losses of cellular ions had occurred. These findings support the existence of mechanical limitations to ventricular cellular swelling, which may be caused by a unique network of interstitial collagen present in ventricular myocardium.

We studied the relationship between the slow inward current, tissue ATP content, and the development of hypoxic myocardial contracture in the rat. Resting tension and active isometric tension were measured using isolated left ventricular papillary muscle preparations. Action potentials and membrane currents were studied using a single sucrose gap voltage clamp technique. The slow inward current (isi) was partially inhibited by verapamil and completely inhibited by cobalt but was not reduced by ryanodine. Active tension was reduced by all three drugs. Non-stimulation, verapamil, and ryanodine did not prevent contracture development with hypoxia, but contracture was markedly reduced by pre-treatment with cobalt. Despite contrasting effects on contracture, both non-stimulation and cobalt partially prevented ATP depletion with hypoxia, suggesting that contracture is not directly related to total muscle ATP content. Cobalt appears to block hypoxic contracture via a mechanism other than simple blockade of isi, inhibition of contraction, or preservation of tissue ATP content.

Isolated left ventricle papillary muscle mechanics and structure were studied in male spontaneously hypertensive (SHR) rats and two control groups of animals, the normotensive Wistar (NR) and the Wistar-Kyoto rat (WKY). Active tension and its first derivative (dT/dtmax) normalized for muscle cross-sectional area were increased in preparations from the SHR at all ages studied relative to control groups (P less than 0.01). However, when these parameters were normalized for myofibrillar cross-sectional area determined from electronmicroscopic point counting data, differences between groups were no longer significant. Force-velocity relations provided no evidence for a depression of shortening velocity at any load in the SHR at any age relative to the two control groups. The duration of mechanical activity, as determined by time-to-peak isometric tension and analysis of muscle force-velocity-time relations, was prolonged only in the 18 month old SHR (P less than 0.01). Thus, while changes in isolated muscle performance occur at a time when hemodynamic impairment is reported in the intact animal (male 18 month SHR), no evidence for depression of isolated muscle function is seen in the SHR at 6, 12 or 18 months of age.

The effect of acetylstrophanthidine (AS), a rapid-acting digitalis-like agent, on the ventricular fibrillation (VF) threshold was examined in normal and denervated chloralose-anesthetized dogs. In neurally intact dogs an intravenous bolus of AS (0.075 mg/kg) increased the VF threshold up to a maximum 50% (P less than 0.01) within 30 min after injection. The augmented VF threshold following intravenous administration of AS was not altered by vagotomy. Bilateral stellectomy in vagotomized dogs, as well as carotid sinus and aortic arch denervations, however, prevented the AS induced increase in VF threshold. In neurally intact dogs beta-adrenergic blockade with propranolol (0.25 mg/kg) precluded AS effects. These data suggest that the increase in the VF threshold resulting from AS administration in the normal canine ventricle is due to withdrawal of sympathetic tone mediated via the baroreceptor reflex. The direct effect of AS on the myocardium is to decrease the VF threshold.

The mechanical performance of isolated left ventricular muscle preparations from rats exposed to smoke from Kentucky Reference cigarettes was examined for possible chronic effects. Eight rats were subjected to smoke for periods of 10 min/hr for 5 hr/day for 180 days. Nineteen additional rats served as either sham-smoked controls or weight-matched, food-deprived controls. Rats exposed to tobacco smoke had a significant diminution in body and left ventricular weight compared to sham-smoked controls. When compared to food-deprived rats, no differences in weights were observed. Contraction mechanics were measured for each muscle at the peak of its length tension curve. No significant difference in cardiac muscle performance was found in rats exposed to tobacco smoke when compared to control animals with respect to contractile performance under oxygenated conditions, muscle performance during 60 min of hypoxia or subsequent reoxygenation, or sensitivity of mechanical performance to isoproterenol. Thus, chronic cigarette smoke exposure did not alter the intrinsic mechanical performance of isolated rat ventricular muscle.

We compared myocardial mechanics and myosin isozymes of right and left ventricular papillary muscles from adult (6 to 8 month old) male rats. Analysis of force velocity relations indicate that right ventricular papillary muscles contract more rapidly than left at light loads (2.68 +/- 0.13 vs 2.18 +/- 0.07 muscle lengths/s measured 75 ms following stimulation, at 0.5 g/mm2; P less than 0.01). Right ventricular papillary muscles had significantly more of the alpha heavy chain containing V1 myosin isozyme and less of the V3 containing beta heavy chain myosin isozyme than left ventricular preparations (P less than 0.05). Papillary muscle and ventricular free wall myosin isozyme distribution were not significantly different within their respective chambers. The presence of a relatively larger proportion of the alpha heavy chain containing myosin isozyme (V1) in right ventricle papillary muscles relative to left correlated with the more rapid velocities of shortening seen in right ventricular papillary muscles (r = 0.60; P less than 0.01).