Viscoelastic properties of pressure overload hypertrophied myocardium: effect of serine protease treatment

To determine whether and to what extent one component of the extracellular matrix, fibrillar collagen, contributes causally to abnormalities in viscoelasticity, collagen was acutely degraded by activation of endogenous matrix metalloproteinases (MMPs) with the serine protease plasmin. Papillary muscles were isolated from normal cats and cats with right ventricular pressure overload hypertrophy (POH) induced by pulmonary artery banding. Plasmin treatment caused MMP activation, collagen degradation, decreased the elastic stiffness constant, and decreased the viscosity constant in both normal and POH muscles. Thus, whereas many mechanisms may contribute to the abnormalities in myocardial viscoelasticity in the POH myocardium, changes in fibrillar collagen appear to play a predominant role.     

Novel regulatory mechanism of cardiomyocyte contractility involving ICAM-1 and the cytoskeleton

ICAM-1 mediates interaction of cardiomyocytes with the extracellular matrix and leukocytes and may play a role in altering contractility. To investigate this possibility, rat ventricular cardiomyocytes were activated using TNF-alpha, IL-1beta, or LPS, washed, cultured with quiescent rat polymorphonuclear leukocytes (PMNs) for 4 h, and electrically stimulated to determine fractional shortening. PMNs cultured with activated cardiomyocytes reduced control fractional shortening of 20.5 +/- 0.7% by -2.8 +/- 0.3% per adherent PMN (P < 0.001). Fixing PMNs with paraformaldehyde or glutaraldehyde did not prevent PMN-mediated decreases in cardiomyocyte fractional shortening. However, PMN adherence and decreased fractional shortening were prevented by anti-ICAM-1 and anti-CD18 antibodies. Reduced fractional shortening was reproduced in the absence of PMNs by ICAM-1 binding using cross-linking antibodies (reduced by 36 +/- 3% from control, P < 0.01). Immunofluorescent staining demonstrated increased cortical cytoskeleton-associated focal adhesion kinase expression after ICAM-1 cross-linking, suggesting involvement of the actin cytoskeleton. Indeed, disruption of F-actin filament assembly using cytochalasin D or latrunculin A did not prevent PMN adherence but prevented decreased fractional shortening. Inhibition of the cytoskeleton-associated Rho-kinase pathway with HA-1077 prevented ICAM-1-mediated decreases in cardiomyocyte contractility, further suggesting a central role of the actin cytoskeleton. Importantly, ICAM-1 cross-linking did not alter the total intracellular Ca2+ transient during cardiomyocyte contraction but greatly increased heterogeneity of intracellular Ca2+ release. Thus we have identified a novel regulatory mechanism of cardiomyocyte contractility involving the actin cytoskeleton as a central regulator of the normally highly coordinated pattern of sarcoplasmic Ca2+ release. Cardiomyocyte ICAM-1 binding, by PMNs or other ligands, induces decreased cardiomyocyte contractility via this pathway.     

Dissociation between cardiomyocyte function and remodeling with beta-adrenergic receptor blockade in isolated canine mitral regurgitation

The low-pressure volume overload of isolated mitral regurgitation (MR) is associated with increased adrenergic drive, left ventricular (LV) dilatation, and loss of interstitial collagen. We tested the hypothesis that beta1-adrenergic receptor blockade (beta1-RB) would attenuate LV remodeling after 4 mo of MR in the dog. beta1-RB did not attenuate collagen loss or the increase in LV mass in MR dogs. Using MRI and three-dimensional (3-D) analysis, there was a 70% increase in the LV end-diastolic (LVED) volume-to-LV mass ratio, a 23% decrease in LVED midwall circumferential curvature, and a >50% increase in LVED 3-D radius/wall thickness in MR dogs that was not attenuated by beta1-RB. However, beta1-RB caused a significant increase in LVED length from the base to apex compared with untreated MR dogs. This was associated with an increase in isolated cardiomyocyte length (171+/-5 microm, P<0.05) compared with normal (156+/-3 microm) and MR (165+/-4 microm) dogs. Isolated cardiomyocyte fractional shortening was significantly depressed in MR dogs compared with normal dogs (3.73+/-0.31 vs. 5.02+/-0.26%, P<0.05) and normalized with beta1-RB (4.73+/-0.48%). In addition, stimulation with the beta-adrenergic receptor agonist isoproterenol (25 nM) increased cardiomyocyte fractional shortening by 215% (P<0.05) in beta1-RB dogs compared with normal (56%) and MR (50%) dogs. In summary, beta1-RB improved LV cardiomyocyte function and beta-adrenergic receptor responsiveness despite further cell elongation. The failure to attenuate LV remodeling associated with MR could be due to a failure to improve ultrastructural changes in extracellular matrix organization.     

Cardiac dilatation and pump dysfunction without intrinsic myocardial systolic failure following chronic beta-adrenoreceptor activation

There is no direct evidence to indicate that pump dysfunction in a dilated chamber reflects the impact of chamber dilatation rather than the degree of intrinsic systolic failure resulting from myocardial damage. In the present study, we explored the relative roles of intrinsic myocardial systolic dysfunction and chamber dilatation as mediators of left ventricular (LV) pump dysfunction. Administration of isoproterenol, a beta-adrenoreceptor agonist, for 3 mo to rats (0.1 mg.kg(-1).day(-1)) resulted in LV pump dysfunction as evidenced by a reduced LV endocardial fractional shortening (echocardiography) and a decrease in the slope of the LV systolic pressure-volume relation (isolated heart preparations). Although chronic beta-adrenoreceptor activation induced cardiomyocyte damage (deoxynucleotidyl transferase-mediated dUTP nick-end labeling) as well as beta(1)- and beta(2)-adrenoreceptor inotropic downregulation (attenuated contractile responses to dobutamine and salbutamol), these changes failed to translate into alterations in intrinsic myocardial contractility. Indeed, LV midwall fractional shortening (echocardiography) and the slope of the LV systolic stress-strain relation (isolated heart preparations) were unchanged. A normal intrinsic myocardial systolic function, despite the presence of cardiomyocyte damage and beta-adrenoreceptor inotropic downregulation, was ascribed to marked increases in myocardial norepinephrine release, to upregulation of alpha-adrenoreceptor-mediated contractile effects as determined by phenylephrine responsiveness, and to compensatory LV hypertrophy. LV pump failure was attributed to LV dilatation, as evidenced by increased LV internal dimensions (echocardiography), and a right shift and increased volume intercept of the LV diastolic pressure-volume relation. In conclusion, chronic sympathetic stimulation, despite reducing beta-adrenoreceptor-mediated inotropic responses and promoting myocyte apoptosis, may nevertheless induce pump dysfunction primarily through LV dilatation, rather than intrinsic myocardial systolic failure.     

Alpha 1-adrenergic stimulation increases the Vmax of isolated myocardial papillary muscles.

alpha-Adrenergic agonists have been shown to increase the tension developed by myocardial muscle. However, their effects on the maximum velocity of unloaded muscle shortening (Vmax) have not been rigorously examined. In this study, the contractile effects of the alpha-adrenergic agonist phenylephrine were examined in the presence of propranolol in papillary muscles of two species, the dog and the rabbit. In rabbit papillary muscles studied at physiological calcium concentrations (1.25 mM), phenylephrine increased all indices of contractility (Vmax, tension, and maximum rate of tension developed (dT/dt)) starting at 10(-8) M. The percent increase in Vmax (121 +/- 8%) was less than that of tension (188 +/- 20%, p less than 0.05) and dT/dt (262 +/- 35%, p less than 0.01). These findings occurred at both 29 and 35 degrees C and were inhibited by adding 10(-5) M prazosin. Increasing extracellular calcium concentration from 1.25 to 15 mM caused changes in twitch configuration that were significantly different from those of phenylephrine. Calcium increased all indices of contractility more than did phenylephrine. This was particularly true for dT/dt (502 +/- 82 vs. 262 +/- 35% for phenylephrine, p less than 0.01). Nonetheless, the ratio of increase in tension to increase in Vmax under both experimental conditions was similar (the increase in Vmax was 64% of that of tension with phenylephrine and 69% with increased calcium). At 1.25 mM calcium, the increase in contractility caused by phenylephrine was much smaller in dog myocardium as compared with rabbit myocardium. Rather, the effects of phenylephrine on dog myocardium studied at 1.25 mM calcium resembled that of rabbit myocardium studied at 15 mM calcium.(ABSTRACT TRUNCATED AT 250 WORDS)     

Overexpression of Na+/Ca2+ exchanger gene attenuates postinfarction myocardial dysfunction

We monitored myocardial function in postinfarcted wild-type (WT) and transgenic (TG) mouse hearts with overexpression of the cardiac Na(+)/Ca(2+) exchanger. Five weeks after infarction, cardiac function was better maintained in TG than WT mice [left ventricular (LV) systolic pressure: WT, 41 +/- 2; TG, 58 +/- 3 mmHg; P < 0.05; maximum rising rate of LV pressure (+dP/dt(max)): WT, 3,750 +/- 346; TG, 5,075 +/- 334 mmHg/s; P < 0.05]. The isometric contractile response to beta-adrenergic stimulation was greater in papillary muscles from TG than WT mice (WT, 13.2 +/- 0.9; TG, 16.3 +/- 1.0 mN/mm(2) at 10(-4) M isoproterenol). The sarcoplasmic reticulum (SR) Ca(2+) content investigated by rapid cooling contractures in papillary muscles was greater in TG than WT mouse hearts. We conclude that myocardial function is better preserved in TG mice 5 wk after infarction, which results from enhanced SR Ca(2+) content via overexpression of the Na(+)/Ca(2+) exchanger.     

Effects of sevoflurane on the contractility of ferret ventricular myocardium.

Isotonic and isometric variables of contractility and relaxation of isolated ferret right ventricular papillary muscles were measured before and during exposure to incremental concentrations of sevoflurane (0-4.9% vol/vol) (30 degrees C) (n = 9). In a second group of muscles (n = 8), effects of sevoflurane were compared with those of low [Ca(2+)](o) (0.45-2.25 mM in steps of 0.45 mM). Sevoflurane caused a reversible concentration-dependent decrease in contractility (ED(50) of developed force 4.6+/-0.9% vol/vol). When compared with twitches of equal amplitude in low extracellular Ca(2+) concentration, sevoflurane accelerated both isometric and isotonic relaxation. The myocardial depressant effect of sevoflurane is less than that of isoflurane and results mainly from a decrease of intracellular Ca(2+) availability. The abbreviated isometric relaxation likely reflects a decrease in Ca(2+) sensitivity and the faster isotonic relaxation may reflect a mild stimulation of Ca(2+) uptake by the sarcoplasmic reticulum.     

Testosterone modulates cardiomyocyte Ca(2+) handling and contractile function

The extent to which sex differences in cardiac function may be attributed to the direct myocardial influence of testosterone is unclear. In this study the effects of gonadal testosterone withdrawal (GDX) and replacement (GDX+T) in rats, on cardiomyocyte shortening and intracellular Ca(2+) handling was investigated (0.5 Hz, 25 oC). At all extracellular [Ca(2+)] tested (0.5-2.0 mM), the Ca(2+) transient amplitude was significantly reduced (by approximately 50 %) in myocytes of GDX rats two weeks post-gonadectomy. The time course of Ca(2+) transient decay was significantly prolonged in GDX myocytes (tau, 455+/-80 ms) compared with intact (279+/-23 ms) and GDX+T (277+/-19 ms). Maximum shortening of GDX myocytes was markedly reduced (by more than 60 %) and relaxation significantly delayed (by more than 35 %) compared with intact and GDX+T groups. Thus testosterone replacement completely reversed the cardiomyocyte hypocontractility induced by gonadectomy. These results provide direct evidence for a role of testosterone in regulating functional Ca(2+) handling and contractility in the heart.     

Plasmin potentiates synaptic N-methyl-D-aspartate receptor function in hippocampal neurons through activation of protease-activated receptor-1

Protease-activated receptor-1 (PAR1) is activated by a number of serine proteases, including plasmin. Both PAR1 and plasminogen, the precursor of plasmin, are expressed in the central nervous system. In this study we examined the effects of plasmin in astrocyte and neuronal cultures as well as in hippocampal slices. We find that plasmin evokes an increase in both phosphoinositide hydrolysis (EC(50) 64 nm) and Fura-2/AM fluorescence (195 +/- 6.7% above base line, EC(50) 65 nm) in cortical cultured murine astrocytes. Plasmin also activates extracellular signal-regulated kinase (ERK1/2) within cultured astrocytes. The plasmin-induced rise in intracellular Ca(2+) concentration ([Ca(2+)](i)) and the increase in phospho-ERK1/2 levels were diminished in PAR1(-/-) astrocytes and were blocked by 1 microm BMS-200261, a selective PAR1 antagonist. However, plasmin had no detectable effect on ERK1/2 or [Ca(2+)](i) signaling in primary cultured hippocampal neurons or in CA1 pyramidal cells in hippocampal slices. Plasmin (100-200 nm) application potentiated the N-methyl-D-aspartate (NMDA) receptor-dependent component of miniature excitatory postsynaptic currents recorded from CA1 pyramidal neurons but had no effect on alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate- or gamma-aminobutyric acid receptor-mediated synaptic currents. Plasmin also increased NMDA-induced whole cell receptor currents recorded from CA1 pyramidal cells (2.5 +/- 0.3-fold potentiation over control). This effect was blocked by BMS-200261 (1 microm; 1.02 +/- 0.09-fold potentiation over control). These data suggest that plasmin may serve as an endogenous PAR1 activator that can increase [Ca(2+)](i) in astrocytes and potentiate NMDA receptor synaptic currents in CA1 pyramidal neurons.     

Heart size-independent analysis of myocardial function in murine pressure overload hypertrophy

Pressure overload cardiac hypertrophy may be a compensatory mechanism to normalize systolic wall stress and preserve left ventricular (LV) function. To test this concept, we developed a novel in vivo method to measure myocardial stress (sigma)-strain (epsilon) relations in normal and hypertrophied mice. LV volume was measured using two pairs of miniature omnidirectional piezoelectric crystals implanted orthogonally in the endocardium and one crystal placed on the anterior free wall to measure instantaneous wall thickness. Highly linear sigma-epsilon relations were obtained in control (n = 7) and hypertrophied mice produced by 7 days of transverse aortic constriction (TAC; n = 13). Administration of dobutamine in control mice significantly increased the load-independent measure of LV contractility, systolic myocardial stiffness. In TAC mice, systolic myocardial stiffness was significantly greater than in control mice (3,156 +/- 1,433 vs. 1,435 +/- 467 g/cm(2), P < 0.01), indicating enhanced myocardial contractility with pressure overload. However, despite the increased systolic performance, both active (time constant of LV pressure decay) and passive (diastolic myocardial stiffness constant) diastolic properties were markedly abnormal in TAC mice compared with control mice. These data suggest that the development of cardiac hypertrophy is associated with a heightened contractile state, perhaps as an early compensatory response to pressure overload.     

Non-genomic effects of aldosterone on intracellular ion regulation and cell volume in rat ventricular myocytes

Aldosterone has non-genomic effects that express within minutes and modulate intracellular ion milieu and cellular function. However, it is still undefined whether aldosterone actually alters intracellular ion concentrations or cellular contractility. To clarify the non-genomic effects of aldosterone, we measured [Na+]i, Ca2+ transient (CaT), and cell volume in dye-loaded rat ventricular myocytes, and we also evaluated myocardial contractility. We found the following: (i) aldosterone increased [Na+]i at the concentrations of 100 nmol/L to 10 micromol/L; (ii) aldosterone (up to 10 micromol/L) did not alter CaT and cell shortening in isolated myocytes, developed tension in papillary muscles, or left ventricular developed pressure in Langendorff-perfused hearts; (iii) aldosterone (100 nmol/L) increased the cell volume from 47.5 +/- 3.6 pL to 49.8 +/- 3.7 pL (n=8, p<0.05); (iv) both the increases in [Na+]i and cell volume were blocked by a Na+-K+-2Cl- co-transporter (NKCCl) inhibitor, bumetanide, or by a Na+/H+ exchange (NHE) inhibitor, 5-(N-ethyl-N-isopropyl) amiloride; and (v) spironolactone by itself increased in [Na+]i and cell volume. In conclusion, aldosterone rapidly increased [Na+]i and cell volume via NKCC1 and NHE, whereas there were no changes in CaT or myocardial contractility. Hence the non-genomic effects of aldosterone may be related to cell swelling rather than the increase in contractility.     

Knockdown of lncRNA AK139328 alleviates myocardial ischaemia/reperfusion injury in diabetic mice via modulating miR‐204‐3p and inhibiting autophagy

This study was aimed at investigating the effects of lncRNA AK139328 on myocardial ischaemia/reperfusion injury (MIRI) in diabetic mice. Ischaemia/reperfusion (I/R) model was constructed in normal mice (NM) and diabetic mice (DM). Microarray analysis was utilized to identify lncRNA AK139328 overexpressed in DM after myocardial ischaemia/reperfusion (MI/R). RT‐qPCR assay was utilized to investigate the expressions of lncRNA AK139328 and miR‐204‐3p in cardiomyocyte and tissues. Left ventricular end diastolic diameter (LVEDD), left ventricular end systolic diameter (LVESD), left ventricular ejection fraction (LVEF) and fractioning shortening (FS) were obtained by transthoracic echocardiography. Haematoxylin‐eosin (HE) staining and Masson staining were utilized to detect the damage of myocardial tissues degradation of myocardial fibres and integrity of myocardial collagen fibres. Evans Blue/TTC staining was used to determine the myocardial infarct size. TUNEL staining was utilized to investigate cardiomyocyte apoptosis. The targeted relationship between lncRNA AK139328 and miR‐204‐3p was confirmed by dual‐luciferase reporter gene assay. MTT assay was used for analysis of cardiomyocyte proliferation. Western blot was utilized to investigate the expression of alpha smooth muscle actin (α‐SMA), Atg7, Atg5, LC3‐II/LC3‐I and p62 marking autophagy. Knockdown of lncRNA AK139328 relieved myocardial ischaemia/reperfusion injury in DM and inhibited cardiomyocyte autophagy as well as apoptosis of DM. LncRNA AK139328 modulated miR‐204‐3p directly. MiR‐204‐3p and knockdown of lncRNA AK139328 relieved hypoxia/reoxygenation injury via inhibiting cardiomyocyte autophagy. Silencing lncRNA AK139328 significantly increased miR‐204‐3p expression and inhibited cardiomyocyte autophagy, thereby attenuating MIRI in DM.     

Mechanical and energetic effects of chronic chagasic patients' antibodies on rat myocardium

Chagasic (Ch) and nonchagasic (NCh) IgG fraction (20 microg/ml) effects on cardiac performance of adult Wistar rat ventricles were studied with a novel approach applying a microcalorimetric technique. Resting heat (Hr) was significantly decreased by Ch antibodies (DeltaHrCh = 4.8 +/- 0.9 mW/g). Although the Hr decrease can be associated with diminished activity of the Na+/K+ pump, the magnitude of the effect (25% of control Hr) indicates that additional processes may also be affected. Ch antibodies induced an initial increase in developed pressure (P), which was associated with a decreased contractile economy. However, after 30 min of Ch antibody perfusion, P reached a significantly lower level (DeltaPCh = 3.8 +/- 1.2 mN/mm2) without changes in active heat per beat (Ha'). Consequently, Ha'/P ratio increased, indicating that the energetic cost per unit of P was higher. In contrast, P and Ha' were both significantly and reversibly decreased by NCh antibodies (DeltaPNCh = 4.4 +/- 1.2 mN/mm2; DeltaHa'NCh = 9.7 +/- 2.2 mJ/g), but Ha'/P remained unaffected. According to these data, normal hearts exposed to Ch antibodies present a biphasic mechanical response: 1) an initial period of increased contractility (and decreased global muscle economy) consistent with antibodies with beta1-adrenergic activity, such as those used in the present study, and 2) a decrease in P at 30 min of Ch antibody perfusion, which suggests that another Ca(2+)-related mechanism is compromised. These data contribute to redefine the role of antibody-mediated responses in the pathophysiology of chronic chagasic cardiomyopathy as agents of myocardial failure.     

Subthreshold nitric oxide synthase inhibition improves synergistic effects of subthreshold MMP‐2/MLCK‐mediated cardiomyocyte protection from hypoxic injury

Injury of myocardium during ischaemia/reperfusion (I/R) is a complex and multifactorial process involving uncontrolled protein phosphorylation, nitration/nitrosylation by increased production of nitric oxide and accelerated contractile protein degradation by matrix metalloproteinase‐2 (MMP‐2). It has been shown that simultaneous inhibition of MMP‐2 with doxycycline (Doxy) and myosin light chain kinase (MLCK) with ML‐7 at subthreshold concentrations protects the heart from contractile dysfunction triggered by I/R in a synergistic manner. In this study, we showed that additional co‐administration of nitric oxide synthase (NOS) inhibitor (1400W or L‐NAME) in subthreshold concentrations improves this synergistic protection in the model of hypoxia–reoxygenation (H‐R)‐induced contractile dysfunction of cardiomyocytes. Isolated cardiomyocytes were subjected to 3 min. of hypoxia and 20 min. of reoxygenation in the presence or absence of the inhibitor cocktails. Contractility of cardiomyocytes was expressed as myocyte peak shortening. Inhibition of MMP‐2 by Doxy (25–100 μM), MLCK by ML‐7 (0.5–5 μM) and NOS by L‐NAME (25–100 μM) or 1400W (25–100 μM) protected myocyte contractility after H‐R in a concentration‐dependent manner. Inhibition of these activities resulted in full recovery of cardiomyocyte contractility after H‐R at the level of highest single‐drug concentration. The combination of subthreshold concentrations of NOS, MMP‐2 and MLCK inhibitors fully protected cardiomyocyte contractility and MLC1 from degradation by MMP‐2. The observed protection with addition of L‐NAME or 1400W was better than previously reported combination of ML‐7 and Doxy. The results of this study suggest that addition of NOS inhibitor to the mixture of inhibitors is better strategy for protecting cardiomyocyte contractility.     

Single cell mechanics of rat cardiomyocytes under isometric, unloaded, and physiologically loaded conditions

One of the most salient characteristics of the heart is its ability to adjust work output to external load. To examine whether a single cardiomyocyte preparation retains this property, we measured the contractile function of a single rat cardiomyocyte under a wide range of loading conditions using a force-length measurement system implemented with adaptive control. A pair of carbon fibers was used to clamp the cardiomyocyte, attached to each end under a microscope. One fiber was stiff, serving as a mechanical anchor, while the bending motion of the compliant fiber was monitored for force-length measurement. Furthermore, by controlling the position of the compliant fiber using a piezoelectric translator based on adaptive control, we could change load dynamically during contractions. Under unloaded conditions, maximal shortening velocity was 106 +/- 8.9 microm/s (n = 13 cells), and, under isometric conditions, peak developed force reached 5,720 nN (41.6 +/- 5.6 mN/mm(2); n = 17 cells). When we simulated physiological working conditions consisting of an isometric contraction, followed by shortening and relaxation, the average work output was 828 +/- 123 J/m(3) (n = 20 cells). The top left corners of tension-length loops obtained under all of these conditions approximate a line, analogous to the end-systolic pressure-volume relation of the ventricle. All of the functional characteristics described were analogous to those established by studies using papillary muscle or trabeculae preparations. In conclusion, the present results confirmed the fact that each myocyte forms the functional basis for ventricular function and that single cell mechanics can be a link between subcellular events and ventricular mechanics.     

Increased coronary perfusion augments cardiac contractility in the rat through stretch-activated ion channels

The role of stretch-activated ion channels (SACs) in coronary perfusion-induced increase in cardiac contractility was investigated in isolated isometrically contracting perfused papillary muscles from Wistar rats. A brief increase in perfusion pressure (3-4 s, perfusion pulse, n = 7), 10 repetitive perfusion pulses (n = 4), or a sustained increase in perfusion pressure (150-200 s, perfusion step, n = 7) increase developed force by 2.7 +/- 1.1, 7.7 +/- 2.2, and 8.3 +/- 2.5 mN/mm(2) (means +/- SE, P < 0.05), respectively. The increase in developed force after a perfusion pulse is transient, whereas developed force during a perfusion step remains increased by 5.1 +/- 2.5 mN/mm(2) (P < 0.05) in the steady state. Inhibition of SACs by addition of gadolinium (10 micromol/l) or streptomycin (40 and 100 micromol/l) blunts the perfusion-induced increase in developed force. Incubation with 100 micromol/l N(omega)-nitro-L-arginine [nitric oxide (NO) synthase inhibition], 10 micromol/l sodium nitroprusside (NO donation) and 0.1 micromol/l verapamil (L-type Ca(2+) channel blockade) are without effect on the perfusion-induced increase of developed force. We conclude that brief, repetitive, or sustained increases in coronary perfusion augment cardiac contractility through activation of stretch-activated ion channels, whereas endothelial NO release and L-type Ca(2+) channels are not involved.     

Alterations in myocardial collagen content affect rat papillary muscle function

We investigated the influence of myocardial collagen volume fraction (CVF, %) and hydroxyproline concentration (microg/mg) on rat papillary muscle function. Collagen excess was obtained in 10 rats with unilateral renal ischemia for 5 wk followed by 3-wk treatment with ramipril (20 mg. kg(-1). day(-1)) (RHTR rats; CVF = 3.83 +/- 0. 80, hydroxyproline = 3.79 +/- 0.50). Collagen degradation was induced by double infusion of oxidized glutathione (GSSG rats; CVF = 2.45 +/- 0.52, hydroxyproline = 2.85 +/- 0.18). Nine untreated rats were used as controls (CFV = 3.04 +/- 0.58, hydroxyproline = 3.21 +/- 0.30). Active stiffness (AS; g. cm(-2). %L(max)(-1)) and myocyte cross-sectional area (MA; micrometer(2)) were increased in the GSSG rats compared with controls [AS 5.86 vs. 3.96 (P < 0.05); MA 363 +/- 59 vs. 305 +/- 28 (P < 0.05)]. In GSSG and RHTR groups the passive tension-length curves were shifted downwards, indicating decreased passive stiffness, and upwards, indicating increased passive stiffness, respectively. Decreased collagen content induced by GSSG is related to myocyte hypertrophy, decreased passive stiffness, and increased AS, and increased collagen concentration causes myocardial diastolic dysfunction with no effect on systolic function.     

Inhibition of eicosanoid-mediated coronary constriction during myocardial ischemia

Thromboxane A2 and cysteinyl leukotrienes are highly effective microvessel constrictors in normally perfused myocardium. Their release during acute coronary thrombosis might augment myocardial underperfusion. The constrictor action of these substances could be modified substantially, however, by concomitant myocardial ischemia. We compared the effects of the two eicosanoid constrictors in normally perfused and ischemic myocardium of 24 open-chest, pentobarbital-anesthetized pigs. Left anterior descending coronary flow was measured after intracoronary bolus injections of the stable thromboxane A2 analog U46619 (1-10 micrograms) or leukotriene D4 (LTD4, 1-10 micrograms). Each dose was given before and during myocardial ischemia induced by a snare adjusted to produce 63 +/- 2% decrease in coronary flow for 10 min. Marked dose-independent inhibition of eicosanoid-induced coronary flow decrease occurred during ischemia. With 10 micrograms U46619, coronary flow decrease in the unoccluded state (25 +/- 2 from 55 +/- 4 ml/min pretreatment baseline) was virtually eliminated during snare occlusion (1 +/- 1 from 21 +/- 3 ml/min pretreatment baseline, P less than 0.001). Similar results occurred with LTD4. Distal coronary pressure during ischemia indicated a lack of microvessel responsiveness to the eicosanoids rather than a buffering of resistance change by the snare. U46619 and LTD4 did induce transient, small reductions in regional shortening fraction during ischemia. Our data suggest that eicosanoid-induced constriction of myocardial resistance vessels is not a likely complication of acute coronary thrombosis. However, eicosanoids could depress residual contractility in moderately ischemic regions.     

Brief rapid pacing depresses contractile function via Ca(2+)/PKC-dependent signaling in cat ventricular myocytes

The purpose of this study is to determine the effects of brief rapid pacing (RP; approximately 200-240 beats/min for approximately 5 min) on contractile function in ventricular myocytes. RP was followed by a sustained inhibition of peak systolic cell shortening (-44 +/- 4%) that was not due to changes in diastolic cell length, membrane voltage, or L-type Ca(2+) current (I(Ca,L)). During RP, baseline and peak intracellular Ca(2+) concentration ([Ca(2+)](i)) increased markedly. After RP, Ca(2+) transients were similar to control. The effects of RP on cell shortening were not prevented by 1 microM calpain inhibitor I, 25 microM L-N(5)-(1-iminoethyl)-orthinthine, or 100 microM N(G)-monomethyl-L-arginine. However, RP-induced inhibition of cell shortening was prevented by lowering extracellular [Ca(2+)] (0.5 mM) during RP or exposure to chelerythrine (2-4 microM), a protein kinase C (PKC) inhibitor, or LY379196 (30 nM), a selective inhibitor of PKC-beta. Exposure to phorbol ester (200 nM phorbol 12-myristate 13-acetate) inhibited cell shortening (-46 +/- 7%). Western blots indicated that cat myocytes express PKC-alpha, -delta, and -epsilon as well as PKC-beta. These findings suggest that brief RP of ventricular myocytes depresses contractility at the myofilament level via Ca(2+)/PKC-dependent signaling. These findings may provide insight into the mechanisms of contractile dysfunction that follow paroxysmal tachyarrhythmias.     

Significance of inducible nitric oxide synthase in acute myocarditis caused by Trypanosoma cruzi (Tulahuen strain)

Chagas' disease, caused by Trypanosoma cruzi, is associated with myocarditis and expression of myocardial cytokines and inducible nitric oxide synthase (NOS2). To assess the functional significance of NOS2 in murine Chagas' disease, we infected NOS2 knockout (NOS2(-/-)) and C57BL/6x129sv (wild type) mice with the Tulahuen strain of T. cruzi. Serial transthoracic echocardiography was performed to assess the progression of left and right ventricular dysfunction in infected mice. Uninfected wild type and NOS2(-/-) mice served as controls. At day 10 post-infection (p.i.), infected wild type mice had larger left ventricular end-diastolic diameter (2.52+/-0.14-vs-2.11+/-0.06 mm, P<0.02) and right ventricle (0.6+/-0.2-vs-0 visual grade, P<0.02) as compared with uninfected wild type mice. At day 19 p.i., compared with uninfected controls, infected wild type mice had larger left ventricular end-diastolic diameter (3.30+/-0.29-vs-2.11+/-0.07 mm), left ventricular end-systolic diameter (1.86+/-0.29-vs-0.88+/-0.05 mm), right ventricle (1.6+/-0.2-vs-0 visual grade), lower heart rate (413+/-27-vs-557+/-25 beats per min), left ventricular relative wall thickness (0.44+/-0.05-vs-0.64+/-0.03) and fractional shortening (45+/-4-vs-57+/-2%) [P<0.05 for all]. In contrast, no differences in left ventricular end-diastolic diameter or fractional shortening were noted among infected and uninfected NOS2(-/-) mice at day 19 p.i. Compared with uninfected controls, infected NOS2(-/-) mice had significantly lower heart rate (272+/-23-vs-512+/-31 beats per min, P<0.01) and larger right ventricle (0.6+/-0.2-vs-0, P<0.05 visual grade). The magnitude of right ventricular dilation in NOS2(-/-) mice was less than that observed in infected wild type mice. At necropsy, the heart weight was greater (129+/-16-vs-109+/-7 mg, P=0.02) and myocardial inflammation more severe in infected wild type compared with infected NOS2(-/-) mice. Myocardial interleukin (IL)-1beta, IL-6, tumour necrosis factor-alpha, and interferon-gamma were induced in all infected mice. These data indicate that nitric oxide derived from NOS2 plays an important role in the development and progression of ventricular dilation and systolic dysfunction in acute murine chagasic myocarditis caused by infection with the Tulahuen strain.