Isolated Heart Muscle Research Articles

Force-velocity characteristics of isolated myocardium preparations from rats exposed to subchronic intoxication with lead and cadmium acting separately or in combination

This investigation continues our study of the effects of Pb-Cd poisoning on the heart, extending the enquiry from isometric to auxotonic contractions, thereby examining the effect on the ability of myocardial tissues to perform mechanical work. Different shifts were revealed in myocardial force-velocity relations following subchronic exposure of rats to lead acetate and cadmium chloride acting separately, in combination, or in combination with a bioprotective complex (BPC). The experiments were conducted on isolated preparations of trabecules and papillary muscles of the right ventricle in physiological loading conditions and on isolated heart muscle contractile proteins examined by the in vitro motility assay. The results of the latter correlate with the shifts in the ratio of cardiac myosin isoforms. The amount of work performed by the myocardium was calculated on the basis of the tension-shortening loop area and was found to be similar in the preparations from all experimental groups. This fact presumably reflects adaptive capacity of the myocardial function even when contractility is damaged due to the metallic intoxication of a moderate severity.Some characteristics of rat myocardium altered by the impact of lead-cadmium intoxication became fully or partly normalized if intoxication developed against background administration of a bioprotective complex (BPC). Together with previously reported results obtained in the isometric mode of contractility, all these results strengthen the scientific foundations of risk assessment and risk management projects in the occupational and environmental conditions characterized by human exposure to lead and/or cadmium.

The Effect of Non-Thermal Microwave-Treated Physiological Solution on Isolated Heart Muscle of Snail

The effects of non-thermal intensity of microwave (NT MW)- treated physiological solutions (PS) on muscle contractility, 45Ca uptake by muscle and muscle hydration of snail isolated heart in normal and K+-free PS were studied. The intra-cordial perfusion by NT MW-treated PS led to inhibition of heart beating with gradual decrease of the amplitudes of muscle contraction, the weakness of the Na+/K+ pump-induced transient inhibition of heart beating (TIHB), the inhibition of 45Ca2+ uptake by muscle, the increase of intracellular cGMP contents and muscle dehydration. The obtained data serve as additional evidence for hypothesis that cell bathing aqua solution is a primary target for non-thermal effect of MW on cells and organisms. Keywords: Non-thermal, MW, Heart muscle, Intracellular Ca2+, cGMP, Muscle hydration.

Urocortin 2 autocrine/paracrine and pharmacologic effects to activate AMP-activated protein kinase in the heart

Urocortin 2 (Ucn2), a peptide of the corticotropin-releasing factor (CRF) family, binds with high affinity to type 2 CRF receptors (CRFR2) on cardiomyocytes and confers protection against ischemia/reperfusion. The mechanisms by which the Ucn2-CRFR2 axis mitigates against ischemia/reperfusion injury remain incompletely delineated. Activation of AMP-activated protein kinase (AMPK) also limits cardiac damage during ischemia/reperfusion. AMPK is classically activated by alterations in cellular energetics; however, hormones, cytokines, and additional autocrine/paracrine factors also modulate its activity. We examined the effects of both the endogenous cardiac Ucn2 autocrine/paracrine pathway and Ucn2 treatment on AMPK regulation. Ucn2 treatment increased AMPK activation and downstream acetyl-CoA carboxylase phosphorylation and glucose uptake in isolated heart muscles. These actions were blocked by the CRFR2 antagonist anti-sauvagine-30 and by a PKCε translocation-inhibitor peptide (εV1-2). Hypoxia-induced AMPK activation was also blunted in heart muscles by preincubation with either anti-sauvagine-30, a neutralizing anti-Ucn2 antibody, or εV1-2. Treatment with Ucn2 in vivo augmented ischemic AMPK activation and reduced myocardial injury and cardiac contractile dysfunction after regional ischemia/reperfusion in mice. Ucn2 also directly activated AMPK in ex vivo-perfused mouse hearts and diminished injury and contractile dysfunction during ischemia/reperfusion. Thus, both Ucn2 treatment and the endogenous cardiac Ucn2 autocrine/paracrine pathway activate AMPK signaling pathway, via a PKCε-dependent mechanism, defining a Ucn2-CRFR2-PKCε-AMPK pathway that mitigates against ischemia/reperfusion injury.

Green Tea Catechins Increase the Force of Contraction in Isolated Guinea Pig Atrial Muscle Preparations by Increasing the Amplitude of Intracellular Ca<sup>2+</sup> Concentration

It has been reported that green tea catechins enhance the force of contraction of isolated heart muscle preparations. However, it remains controversial whether or not the increase in force of contraction is related to an increase in the intracellular Ca(2+) concentration ([Ca(2+)]i). In this study, the relationship was investigated using a left atrial muscle preparation isolated from guinea pig heart. In the left atrial muscle preparations without fura-2/AM loading, neither EGC (epigallocatechin) nor EC (epicatechin) influenced the force of contraction, but EGCG (epigallocatechin gallate) and ECG (epicatechin gallate) increased the force of contraction in a dose-dependent manner. The ED(50) value of EGCG was significantly higher than that of ECG. In the atrial muscle preparations loaded with fura-2/AM, EGCG and ECG increased the amplitude of [Ca(2+)]i(peak [Ca(2+)]i minus diastolic [Ca(2+)]i) which is associated with the increase in force of contraction. Simple regression analysis between the degree of increase in the force of contraction and the increase in the amplitude of [Ca(2+)]i revealed a positive correlation in EGCG, ECG and CaCl(2). In addition, the slopes of the regression lines of EGCG and ECG were comparable with those of CaCl(2). It was suggested that atrial muscle preparations had a higher affinity for ECG than EGCG, and that the increase in the force of contraction by EGCG and ECG was closely related to the increase in the amplitude of [Ca(2+)]i.

Ca2+ Sensitization of Myocardial Force and Actomyosin ATPase by (D-Ala2, Met5) Enkephalinamide

The presence of proenkephalin mRNA and proenkephalin peptides in cardiac muscle cells suggests the local production of enkephalins in the myocardium. Yet, the effects of these peptides on the function of the contractile proteins are unknown. The effects of (D-Ala2, Met5) enkephalinamide (DALA) on the activity of the actin stimulated Ca, Mg-myosin ATPase in myofibrils and on the contractility and the activity of the related actomyosin ATPase of chemically skinned muscle fibres from pig myocardium were studied. In this article, it is shown that the myofibrillar actomyosin ATPase as well as the contractility and the actomyosin ATPase in skinned fibres are sensitized to Ca2+ ions by DALA. 10(-11) -10(-6) mol/l DALA decrease the effective concentration of Ca2+ stimulating the myofibrillar ATPase activity by 50% (EC50) from 4.0.10(-5) to 1.5.10(-5) mol/l (p < 0.05). The magnesium dependent myosin ATPase activity at low Ca2+ concentration (10(-9) mol/l) is increased. The EC50 values of Ca2+ for both force development and the related actomyosin ATPase activity of skinned fibres are decreased by DALA (10(-11) -10(-5) mol/l) from 2.5.10(-6) to 2.0.10(-6) mol/l (contractions; p < 0.01) and from 2.0.10(-6) to 1.3.10(-6) mol/l (ATPase activity; p < 0.01). The tension cost (ATPase/tension) of the fibres is unchanged by DALA. In conclusion, the results demonstrate a Ca2+ sensitization of the contractile proteins by low concentrations of DALA, indicating a direct regulatory involvement of enkephalins in the regulation of myocardial contractility. These results correspond with the positive inotropic effects of enkephalins in isolated heart muscle cells.

Synthesis and Biological Activity of Thienodiltiazem in Isolated Heart Muscle Preparations of Guinea Pigs

The new compound thienodiltiazem was synthesized and investigated regarding structure-activity relations and calcium antagonistic properties. Isometric contraction force was measured in guinea-pig papillary muscles and chronotropic activity was studied in right atria of guinea pigs. Compared to the parent drug diltiazem (CAS 42399-41-7) the thieno derivative had a more potent negative chronotropic effect on spontaneously beating right atria and a more potent inotropic effect on papillary muscles. The negative inotropic action was reversed by increasing the extracellular calcium concentration.

Conditional Overexpression of Neuronal Nitric Oxide Synthase Is Cardioprotective in Ischemia/Reperfusion

We previously demonstrated that conditional overexpression of neuronal nitric oxide synthase (nNOS) inhibited L-type Ca2+ channels and decreased myocardial contractility. However, nNOS has multiple targets within the cardiac myocyte. We now hypothesize that nNOS overexpression is cardioprotective after ischemia/reperfusion because of inhibition of mitochondrial function and a reduction in reactive oxygen species generation. Ischemia/reperfusion injury in wild-type mice resulted in nNOS accumulation in the mitochondria. Similarly, transgenic nNOS overexpression caused nNOS abundance in mitochondria. nNOS translocation into the mitochondria was dependent on heat shock protein 90. Ischemia/reperfusion experiments in isolated hearts showed a cardioprotective effect of nNOS overexpression. Infarct size in vivo was also significantly reduced. nNOS overexpression also caused a significant increase in mitochondrial nitrite levels accompanied by a decrease of cytochrome c oxidase activity. Accordingly, O(2) consumption in isolated heart muscle strips was decreased in nNOS-overexpressing nNOS(+)/αMHC-tTA(+) mice already under resting conditions. Additionally, we found that the reactive oxygen species concentration was significantly decreased in hearts of nNOS-overexpressing nNOS(+)/αMHC-tTA(+) mice compared with noninduced nNOS(+)/αMHC-tTA(+) animals. We demonstrated that conditional transgenic overexpression of nNOS resulted in myocardial protection after ischemia/reperfusion injury. Besides a reduction in reactive oxygen species generation, this might be caused by nitrite-mediated inhibition of mitochondrial function, which reduced myocardial oxygen consumption already under baseline conditions.

Ouabain attenuates cardiotoxicity induced by other cardiac steroids

All cardiac steroids have a similar structure, bind to and inhibit the ubiquitous transmembrane protein Na(+), K(+)-ATPase and increase the force of contraction of heart muscle. However, there are diverse biological responses to different cardiac steroids both at the cellular and at the molecular level. Moreover, we have recently shown that ouabain inhibits digoxin- and bufalin-induced changes in membrane traffic. The present study was designed to test the hypothesis that ouabain also has an inhibitory effect on cardiotoxicity induced by other cardiac steroids. The hypothesis was tested in isolated heart muscle preparations and in an in vivo model of cardiotoxicity in guinea pigs. Ouabain at a low dose attenuated the toxicity induced by bufalin and digoxin in heart muscle preparations. In addition, ouabain at the low dose (91 ng.kg(-1).h(-1)), but not at a higher dose (182 ng.kg(-1).h(-1)), delayed the development of digoxin-induced (500 microg.kg(-1).h(-1)) cardiotoxicity in anaesthetized guinea pigs, as manifested by delayed arrhythmia and terminal ventricular fibrillation, as well as a reduced heart rate. In addition, as observed with ouabain, the phosphoinositide 3-kinase inhibitor wortmannin (100 microg.kg(-1).h(-1)) delayed the digoxin-induced arrhythmia in anaesthetized guinea pigs. The present study demonstrates the inhibitory effect, probably through signal transduction pathways, of ouabain on digoxin- and bufalin-induced cardiotoxicity in guinea pigs. Further understanding of this phenomenon could be beneficial for increasing the therapeutic window for cardiac steroids in the treatment of chronic heart failure.

Hypoxia Induces Dilated Cardiomyopathy in the Chick Embryo: Mechanism, Intervention, and Long-Term Consequences

BackgroundIntrauterine growth restriction is associated with an increased future risk for developing cardiovascular diseases. Hypoxia in utero is a common clinical cause of fetal growth restriction. We have previously shown that chronic hypoxia alters cardiovascular development in chick embryos. The aim of this study was to further characterize cardiac disease in hypoxic chick embryos.MethodsChick embryos were exposed to hypoxia and cardiac structure was examined by histological methods one day prior to hatching (E20) and at adulthood. Cardiac function was assessed in vivo by echocardiography and ex vivo by contractility measurements in isolated heart muscle bundles and isolated cardiomyocytes. Chick embryos were exposed to vascular endothelial growth factor (VEGF) and its scavenger soluble VEGF receptor-1 (sFlt-1) to investigate the potential role of this hypoxia-regulated cytokine.Principal FindingsGrowth restricted hypoxic chick embryos showed cardiomyopathy as evidenced by left ventricular (LV) dilatation, reduced ventricular wall mass and increased apoptosis. Hypoxic hearts displayed pump dysfunction with decreased LV ejection fractions, accompanied by signs of diastolic dysfunction. Cardiomyopathy caused by hypoxia persisted into adulthood. Hypoxic embryonic hearts showed increases in VEGF expression. Systemic administration of rhVEGF165 to normoxic chick embryos resulted in LV dilatation and a dose-dependent loss of LV wall mass. Lowering VEGF levels in hypoxic embryonic chick hearts by systemic administration of sFlt-1 yielded an almost complete normalization of the phenotype.Conclusions/SignificanceOur data show that hypoxia causes a decreased cardiac performance and cardiomyopathy in chick embryos, involving a significant VEGF-mediated component. This cardiomyopathy persists into adulthood.

ChemInform Abstract: Synthesis and Biological Activity of Thienodiltiazem in Isolated Heart Muscle Preparations of Guinea Pigs.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

AMP-Activated Protein Kinase Activates p38 Mitogen-Activated Protein Kinase by Increasing Recruitment of p38 MAPK to TAB1 in the Ischemic Heart

AMP-activated protein kinase (AMPK) promotes glucose transport, maintains ATP stores, and prevents injury and apoptosis during ischemia. AMPK has several direct molecular targets in the heart but also may interact with other stress-signaling pathways. This study examined the role of AMPK in the activation of the p38 mitogen-activated protein kinase (MAPK). In isolated heart muscles, the AMPK activator 5-aminoimidazole-4-carboxy-amide-1-beta-D-ribofuranoside (AICAR) increased p38 MAPK activation. In AMPK-deficient mouse hearts, expressing a kinase-dead (KD) alpha2 catalytic subunit, p38 MAPK activation was markedly reduced during low-flow ischemia (2.3- versus 7-fold in wild-type hearts, P<0.01) and was similarly reduced during severe no-flow ischemia in KD hearts (P<0.01 versus ischemic wild type). Knockout of the p38 MAPK upstream kinase, MAPK kinase 3 (MKK3), did not affect ischemic activation of either AMPK or p38 MAPK in transgenic mkk3(-/-) mouse hearts. Ischemia increased p38 MAPK recruitment to transforming growth factor-beta-activated protein kinase 1-binding protein 1 (TAB1), a scaffold protein that promotes p38 MAPK autophosphorylation. Moreover, TAB1 was associated with the alpha2 catalytic subunit of AMPK. p38 MAPK recruitment to TAB1/AMPK complexes required AMPK activation and was reduced in ischemic AMPK-deficient transgenic mouse hearts. The potential role of p38 MAPK in mediating the downstream action of AMPK to promote glucose transport was also assessed. The p38 MAPK inhibitor SB203580 partially inhibited both AICAR- and hypoxia-stimulated glucose uptake and GLUT4 translocation. Activation of p38 MAPK by anisomycin also increased glucose transport in heart muscles. Thus, AMPK has an important role in promoting p38 MAPK activation in the ischemic heart by inducing p38 MAPK autophosphorylation through interaction with the scaffold protein TAB1.

Role of the nitric oxide pathway in AMPK-mediated glucose uptake and GLUT4 translocation in heart muscle.

AMP-activated protein kinase (AMPK) is a serine-threonine kinase that regulates cellular metabolism and has an essential role in activating glucose transport during hypoxia and ischemia. The mechanisms responsible for AMPK stimulation of glucose transport are uncertain, but may involve interaction with other signaling pathways or direct effects on GLUT vesicular trafficking. One potential downstream mediator of AMPK signaling is the nitric oxide pathway. The aim of this study was to examine the extent to which AMPK mediates glucose transport through activation of the nitric oxide (NO)-signaling pathway in isolated heart muscles. Incubation with 1 mM 5-amino-4-imidazole-1-beta-carboxamide ribofuranoside (AICAR) activated AMPK (P < 0.01) and stimulated glucose uptake (P < 0.05) and translocation of the cardiomyocyte glucose transporter GLUT4 to the cell surface (P < 0.05). AICAR treatment increased phosphorylation of endothelial NO synthase (eNOS) approximately 1.8-fold (P < 0.05). eNOS, but not neuronal NOS, coimmunoprecipitated with both the alpha(2) and alpha(1) AMPK catalytic subunits in heart muscle. NO donors also increased glucose uptake and GLUT4 translocation (P < 0.05). Inhibition of NOS with N(omega)-nitro-l-arginine and N(omega)-methyl-l-arginine reduced AICAR-stimulated glucose uptake by 21 +/- 3% (P < 0.05) and 25 +/- 4% (P < 0.05), respectively. Inhibition of guanylate cyclase with ODQ and LY-83583 reduced AICAR-stimulated glucose uptake by 31 +/- 4% (P < 0.05) and 22 +/- 3% (P < 0.05), respectively, as well as GLUT4 translocation to the cell surface (P < 0.05). Taken together, these results indicate that activation of the NO-guanylate cyclase pathway contributes to, but is not the sole mediator of, AMPK stimulation of glucose uptake and GLUT4 translocation in heart muscle.

Can Tissue Engineering Mend Broken Hearts?

Cardiac tissue engineering is an emerging field that may hold great promise for advancing the treatment of heart diseases. Cardiac tissue engineering is in its infancy, and the overall field of tissue engineering, which was formalized in the late 1980s at conferences and workshops sponsored by the National Science Foundation, is still new enough to warrant some description. By broad definition, tissue engineering involves the construction of tissue equivalents through the manipulation and combination of living cells and biomaterials. It is a multidisciplinary field combining diverse aspects of the life sciences, engineering, and clinical medicine. The overall goal of tissue engineering is to develop tissue equivalents for use in the repair, replacement, maintenance, or augmentation of tissues or organs. Although some aspects of cardiac tissue engineering research have been ongoing for generations, albeit without being known as such, directed efforts in the field are only beginning. The main justification for cardiac tissue engineering initiatives is straightforward: congenital and acquired heart diseases are substantial health problems, and there is a limited amount of donor tissue for use in surgical repairs. Heart defects are the most common congenital defect and are the leading cause of death in the first year of life.1,2⇓ Congenital heart defects may occur in as many as 14 of every 1000 live births,3 and approximately 25 000 surgical procedures are performed each year to correct them. Acquired heart diseases also have a profound effect on the population, and despite tremendous advances in medical and surgical treatments, it is estimated that each year 20 000 to 40 000 Americans could benefit from a heart transplant.4 Unfortunately, fewer than 2500 heart transplants are performed each year.4,5⇓ One of the principal reasons for the disparity between patient need and procedures performed is the lack of …

Involvement of opioid delta (delta)- and kappa (kappa)-receptors in ischemic preconditioning in a rat model of myocardial infarction.

There is controversy in the literature regarding the involvement of opioid delta (DOP, OP1)- and kappa (KOP,OP2)-receptors in ischemic preconditioning (IPC). Previous studies on this subject in our laboratories and elsewhere have been performed on either isolated heart muscles of experimental animals, or in open-heart surgery rats. To highlight this problem, we introduced an in vivo model of myocardial infarction in rats, which not only allowed electrocardiographic and enzymatic evaluation, but also morphometric assessment of myocardial infarction. In addition to these parameters, a direct receptor ligand study was undertaken, using [3H]-DPDPE, a specific opioid delta-receptor ligand. In our pharmacodynamic studies, we used the selective opioid delta-receptor agonist D-Ala2,D-Leu5 enkephalin (DADLE) and antagonist natrindole. For the evaluation of opioid kappa-receptors, the selective opioid agonist U-50488H and antagonist nor-BNI were employed. Ischemic preconditioning showed the best beneficial effect, compared with pharmacological stimulation of either opioid delta- or kappa-receptors. In normal rat myocytes, two types of opioid delta-receptors exist, namely, low-affinity and high-affinity opioid receptors. In acute myocardial infarction (30-min ischemia), the low-affinity type opioid receptors disappeared, most likely as a result of receptor downregulation due to an excessive release of enkephalins. There was no change in the density of the high-affinity opioid receptor type, but their affinity significantly increased (p < 0.05) by 58%. The radioligand receptor studies showed that opioid delta 1-receptor type was involved not only in triggering, but also in maintaining, the preconditioned state. On the basis of our pharmacodynamic studies, we suggest that both opioid delta 1- and kappa-receptors are involved in the phenomenon of IPC, but with different effects. After 30 min of left coronary artery occlusion, opioid delta-receptor agonist DADLE decreased the infarct size/area at risk from 59.80% in control, untreated, infarcted rats, to 20.40% in treated rats, without a significant effect (p > 0.05) on the occurrence of early cardiac arrhythmias. Opioid kappa-receptor agonist U-50488H produced an opposite effect on the myocardium. It decreased the infarct size/area at risk by 44%, decreased occurrence of early arrhythmias by 77% and also decreased ventricular ectopic beats by 80%. The opioid delta- and kappa-receptor agonists used in this study significantly reduced (p < 0.05) early (2 h) postinfarction mortality by 22% and 19%, respectively. Further studies are in progress to differentiate between the role of opioid kappa 1- and kappa 2-receptors and the molecular mechanisms of the effects of both opioid delta- and kappa-receptors.

FK506 does not affect cardiac contractility and adrenergic response in vitro

FK506 (tacrolimus) is a new immunosuppressant being used in cardiac allograft transplantation. While cyclosporine A has been shown to exert an acute negative inotropic effect on isolated heart muscle preparations, little is known of the inotropic influence of FK506. The Ca 2+ release channel of human skeletal muscle and cardiac muscle is associated with FK506 binding proteins (FKBP), FKBP12 and FKBP12.6, respectively. FKBPs can be dissociated by treatment with FK506. As a consequence of FK506 exposure, isolated skeletal muscle and cardiac muscle ryanodine receptors show altered gating characteristics. Therefore, we analyzed the direct inotropic effect of FK506 exposure to isolated, intact heart muscle preparations from the human and rabbits. Experiments were performed on isolated, electrically stimulated right atrial auricular muscle strips obtained from human myocardium during elective open heart surgery and on intact right ventricular trabeculae from rabbit hearts. The human preparations were exposed to concentrations of 8×10 −9, 8×10 −8 and 8×10 −6 M FK506 followed by a cumulative dose–response curve with isoprenaline as a non-selective β-adrenoceptor agonist. Our data suggest that FK506 does not exert any positive or negative inotropic effect in either human or rabbit myocardium.

CGP 41251, a new potential anticancer drug, improves contractility of rat isolated cardiac muscle subjected to hypoxia.

The aim of the present work was to examine the effects of 4'-N-benzoyl staurosporine (CGP 41251), a protein kinase C inhibitor with broad antiproliferative activity in many cell lines, on the rat isolated heart contractility under normoxic and hypoxic conditions. Additionally, we examined the effects of CGP 41251, WB-4101 (alpha1a -adrenoceptor antagonist), chloroethylclonidine (CEC) (alpha1b-adrenoceptor antagonist) and selective damage of endocardial endothelium by Triton X-100 on the protection against hypoxia induced by preconditioning of rat heart tissue. Experiments were performed on rat isolated left ventricular papillary muscle. The following parameters were measured: force of contraction (Fc), velocity of contraction (+dF/dt) and velocity of relaxation (-dF/dt). The temperature of the bath solution was 37 degrees C +/- 0.5 degrees C, and rate of electrical stimulation was 0.5 Hz. At concentrations less than 1 microM CGP 41251 did not cause any changes in contractility of rat heart. At 1 and 3 microM, significant positive inotropic action was observed. Treatment of rat papillary muscle by CGP 41251 at 3 microM reduced decreasing of contractility by simulated hypoxia and reperfusion. Moreover, protective effects of preconditioning was not affected by addition of CGP 41251 neither at 1 nor at 3 microM. Pretreatment with CEC at 3 microM, and selective damage of endocardial endothelium induced by fast (1-s) immersion of papillary muscle in 0.5% Triton X-100, but not pretreatment with WB-4101, abolished the protective effects of preconditioning. The results imply that CGP 41251 improves contractility of heart muscle under normoxic and hypoxic conditions, and does not alter hypoxic preconditioning in rat isolated cardiac tissue. Moreover, it was shown that alpha1b-adrenoceptors and endocardial endothelium are involved in triggering of preconditioning in rat isolated heart muscle.

Intracellular, nonreceptor-mediated signaling by adenosine: induction and prevention of neuronal apoptosis.

Inhibitory effect of adenosine on the isolated heart muscle and vascular system were first described in 1929. Since then, numerous reviews have been published on the diverse actions of this nucleoside on a wide variety of cell types. Essentially all effects of adenosine in neurons and non-neuronal cells are mediated by activation of nucleoside membrane receptors coupled to specific intracellular second messenger pathways. This brief review describes two novel actions of adenosine in peripheral sympathetic neurons, which are not mediated by adenosine receptors. First is described how adenosine and related nucleosides are able to induce apoptosis during the initial stages of neuronal growth and development in vitro and in vivo. Second is discussed how adenosine is able to prevent or delay apoptosis in more mature sympathetic neurons subjected to nerve growth factor deprivation in culture. Both the induction and prevention of apoptosis are independent of receptor activation, and totally dependent on the intracellular accumulation and subsequent phosphorylation of adenosine. The physiological significance and mechanisms by which adenosine can induce apoptosis in one situation, and rescue from apoptosis in another, are described in this article.

Anti-muscarinic actions of mitoxantrone in isolated heart muscles of guinea pigs

A hypotheses that mitoxantrone is a competitive antagonist at muscarinic cholinergic receptors was examined in guinea-pig hearts. In isolated left atrial muscle preparations, electrically paced at 2 Hz, the muscarinic agonist, carbachol, caused a concentration-dependent decrease in developed tension. Mitoxantrone caused a parallel right-ward shift of the concentration–response curve for carbachol. Schild plots for the effect of mitoxantrone on the carbachol concentration–response relationship were linear with a slope of 0.88 which was not significantly different from the unity. The right-ward shift of the carbachol concentration–response relationship by mitoxantrone significantly reversed after an additional incubation with a mitoxantrone-free solution, although the reversal was incomplete after a 2-h incubation in the mitoxantrone-free solution. Mitoxantrone caused a concentration-dependent displacement of specific [ 3H]quinuclidinyl benzilate binding to membrane preparations obtained from ventricular muscles of guinea-pig hearts. These results indicate that mitoxantrone acts as a competitive antagonist for the muscarinic receptors.

Peroxynitrite irreversibly decreases diastolic and systolic function in cardiac muscle

Much of the damaging action of nitric oxide in heart may be due to its diffusion-limited reaction with superoxide to form peroxynitrite. Direct infusion of peroxynitrite into isolated perfused hearts fails to model the effects of in situ formation because the bulk of peroxynitrite decomposes before reaching the myocytes. To examine the direct effects of peroxynitrite on the contractile apparatus of the heart, we exposed intact and skinned rat papillary muscles to a steady state concentration of 4-μM peroxynitrite for 5 min, followed by a 30-min recovery period to monitor irreversible effects. In intact muscles developed force fell immediately to 26% of initial force, recovering to 43% by 30 min. Resting tension increased by 600% immediately, and was still elevated 500% by 30 min. Nitrotyrosine immunochemistry showed that peroxynitrite can induce tyrosine nitration at low concentrations and is capable of penetrating 200–380 μm into the papillary muscle after a 5-min infusion. Decomposed peroxynitrite had no effect on either intact or skinned muscle developed force or resting tension. Our results show that peroxynitrite directly damages both developed force and resting tension of isolated heart muscle, which can be extrapolated to systolic and diastolic injury in intact hearts.

Glimepiride (Hoe490) inhibits the rilmakalim induced decrease in intracellular free calcium and contraction of isolated heart muscle cells from guinea pigs to a lesser extent than glibenclamide

Glibenclamide is a potent inhibitor of the ATP-dependent potassium channel. Opening of the ATP-dependent potassium channel is regarded as a mechanism of ischemic preconditioning. This in vitro study examines the influence of glibenclamide and glimepiride, a new sulfonylurea, on the negative inotropic action of the potassium channel opener rilmakalim in isolated ventricular myocytes. Cardiac myocytes were isolated from adult guinea pig hearts by collagenase perfusion and incubated with rilmakalim (concentration range 0.1–12.0 μM), glibenclamide (concentration range 0.03–3.0 μM) plus rilmakalim (3.0 or 7.5 μM), and glimepiride (0.03–9.0 μM) plus rilmakalim (3.0 or 7.5 μM) and paced by electrical field stimulation. Contractility of the myocytes was evaluated by digital image analysis, intracellular free calcium was determined by means of fura-2 fluorescence measurements, and cell viability was assessed morphologically as well as by measurement of lactate dehydrogenase activity. Rilmakalim reduced the systolic intracellular free calcium and contractility of ventricular myocytes in a concentration dependent manner. This effect was antagonized by glibenclamide at lower concentrations (0.3 μM) than glimepiride (3.0 μM). The smaller antagonistic action of glimepiride on the negative inotropic effect of rilmakalim as compared with glibenclamide most likely reflects a less potent inhibition of ATP-dependent potassium channels by glimepiride.