Tricuspid valve complex- a study on its variations in North Indian population and clinical implications

Since cardiovascular diseases are emerging as major causes of morbidity and mortality in the modern era, emphasis is laid on understanding normal as well as variant cardiac anatomy. Moreover, the advancement in diagnostic and therapeutic cardio-invasive techniques have prompted the revision of our existing knowledge and understanding about fine details of atrio-ventricular, valvular and chordo-papillary complexes. This study is an endeavour to establish the morphology of the tricuspid valve and chordo-papillary complex of the right ventricle in north Indian population and to compare it with previously provided data by different researchers. The study was conducted using 52 formalin-fixed adult human hearts. The presence, number, shapes, length, number of additional heads of the papillary muscles were observed. The morphology of the tricuspid valve was also noted. The morphology and morphometry of the tricuspid valve and papillary muscles were defined. Awareness of such information, whether normal or variant, is considered a prerequisite for successful, uncomplicated cardiac surgeries and interventional radiology

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Intracellular Zn(2+) Increase in Cardiomyocytes Induces both Electrical and Mechanical Dysfunction in Heart via Endogenous Generation of Reactive Nitrogen Species.

Oxidants increase intracellular free Zn(2+) concentration ([Zn(2+)]i) in ventricular myocytes, which contributes to oxidant-induced alterations in excitation-contraction coupling (ECC). However, it is not clear whether increased [Zn(2+)]i in cardiomyocytes via increased reactive nitrogen species (RNS) has a role on heart function under pathological conditions, such as hyperglycemia. In this study, first we aimed to investigate the role of increased [Zn(2+)]i under in vitro condition in the development of both electrical and mechanical dysfunction of isolated papillary muscle strips from rat heart via exposed samples to a Zn(2+)-ionophore (Zn-pyrithione; 1 µM) for 20 min. Under simultaneous measurement of intracellular action potential and contractile activity in these preparations, Zn-pyrithione exposure caused marked prolongation in action potential repolarization phase and slowdown in both contraction and relaxation rates of twitch activity. Second, in order to demonstrate an association between increased [Zn(2+)]i and increased RNS, we monitored intracellular [Zn(2+)]i under an acute exposure of nitric oxide (NO) donor sodium nitroprusside, SNP, in freshly isolated quiescent cardiomyocytes loaded with FluoZin-3. Resting level of free Zn(2+) is significantly higher in cardiomyocytes under hyperglycemic condition compared to those of the controls, which seems to be associated with increased level of RNS production in hyperglycemic cardiomyocytes. Western blot analysis showed that Zn-pyrithione exposure induced a marked decrease in the activity of protein phosphatase 1 and 2A, member of macromolecular protein complex of cardiac ryanodine receptors, RyR2, besides significant increase in the phosphorylation level of extracellular signal-regulated kinase1/2 as a concentration-dependent manner. Overall, the present data demonstrated that there is a cross-relationship between increased RNS production and increased [Zn(2+)]i level in cardiomyocytes under pathological conditions such as hyperglycemia.

[Antiarrhythmic effect of TJ0711].

To study the antiarrhythmic effect of the newly developed alpha/beta-blocker TJ0711, a variety of animal models of arrhythmia were induced by CaCl2, ouabain and ischemia/reperfusion. Glass microelectrode technique was used to observe action potentials of right ventricular papillary muscle of guinea pig. The onset time of arrhythmia induced by CaCl2 was significantly prolonged by TJ0711 at 0.75, 1.5 and 3 mg x kg(-1) doses. TJ0711 (1.5 and 3 mg x kg(-1)) can significantly shorten the ventricular tachycardia (VT) and ventricular fibrillation (VF) duration, the incidence of VF and mortality were significantly reduced. On ischemia-reperfusion-induced arrhythmic model, TJ0711 (0.25, 0.5, 1 and 2 mg x kg(-1)) can significantly reduce the ventricular premature contraction (PVC), VT, VF incidence, mortality, arrhythmia score with a dose-dependent manner. At the same time, rats serum lactate dehydrogenase (LDH) and creatine kinase (CK) activities decreased significantly by TJ0711 (1 and 2 mg x kg(-1)). Ouabain could cause arrhythmia in guinea pigs, when TJ0711 (0.375, 0.75, 1.5 and 3 mg x kg(-1)) was given, the doses of ouabain inducing a variety of arrhythmia PVC, VT, VF, cardiac arrest (CA) were significantly increased with a dose-dependent manner. In the TJ0711 0.1-30 micromol x L(-1) concentration range, guinea pig right ventricular papillary muscle action potential RP (rest potential), APA (action potential amplitude) and V(max) (maximum velocity of depolarization) were not significantly affected. APD20, APD50 and APD90 had a shortening trend but no statistical difference with the increase of TJ0711 concentration. TJ0711 has antiarrhythmic effect on the sympathetic nerve excitement and myocardial cell high calcium animal arrhythmia model. Myocardial action potential zero phase conduction velocity and resting membrane potential were not inhibited by TJ0711. APD20, APD50 and APD90 were shortened by TJ0711 at high concentration. Its antiarrhythmic action mechanism may be besides the action of blocking beta1 receptor, may also have a strong selective blocking action on alpha1 receptor and reducing intracellular calcium concentration.

SPRC protects hypoxia and re-oxygenation injury by improving rat cardiac contractile function and intracellular calcium handling.

S-Propargyl-L-cysteine (SPRC, also named as ZYZ-802) is a new compound synthesized in our lab. We investigated whether SPRC has exerted protective effects against cardiac hypoxia/re-oxygenation (H/R) and also explored its mechanisms. In our study, isolated ventricular myocytes were subject to a simulated hypoxia solution for 30 min to induce cell injury. Intracellular concentration of Ca(2+) ([Ca(2+)]i) was measured using specific dyes and detected by digital imaging apparatus. Apoptotic cells were evaluated by TUNEL assay. Intervention with SPRC (10 µM) 30 min before hypoxia, can significantly attenuate the apoptosis of isolated papillary muscles resulting from the H/R injury and protect morphology of the muscles. In isolated ventricular myocytes, SPRC considerably improved left ventricular functional recovery. SPRC also suppressed the increase of ([Ca(2+)]i) during hypoxia stage. By measuring the calcium transient of the cell we concluded that SPRC can preserve the RyR and SERCA activities and improve Ca(2+) handling during the H/R. Furthermore, the protective effect of SPRC can be partly blocked by CSE inhibitor PAG.

Effects of stretch and shortening on gene expression in intact myocardium.

Multiple cues have been suggested as the mechanical stimulus for the heart's hypertrophic response. Our work has previously suggested that the amount of cyclic shortening in cardiomyocytes controls myocyte shape and the amount of stretch controls myocyte size. To identify gene expression changes that occur in response to these mechanical perturbations, we used microarray analysis of papillary muscles cultured for 12 h at physiological or reduced levels of cyclic shortening and physiological or reduced mean stretch. Overall, genes related to extracellular matrix (ECM) were surprisingly prominent in our analysis. Connective tissue growth factor was among a small group of genes regulated by the amount of cyclic shortening regardless of the level of mean stretch, and many more ECM genes were regulated by shortening with reduced amounts of stretch. When we compared our results to gene expression data from an in vivo model of pressure overload (PO), which also decreases myocyte shortening, we found the genes that were commonly regulated in PO and our decreased shortening groups were most significantly enriched for ontology terms related to the ECM, followed by genes associated with mechanosensing and the cytoskeleton. The list of genes regulated in PO and our decreased shortening groups also includes genes known to change early in hypertrophy, such as myosin heavy chain 7, brain natriuretic peptide, and myosin binding protein C. We conclude that in intact myocardium, the amount of cyclic shortening may be an important regulator not only of myocyte genes classically associated with hypertrophy but also of ECM genes.

Decreased RyR2 refractoriness determines myocardial synchronization of aberrant Ca2+ release in a genetic model of arrhythmia.

Dysregulated intracellular Ca(2+) signaling is implicated in a variety of cardiac arrhythmias, including catecholaminergic polymorphic ventricular tachycardia. Spontaneous diastolic Ca(2+) release (DCR) can induce arrhythmogenic plasma membrane depolarizations, although the mechanism responsible for DCR synchronization among adjacent myocytes required for ectopic activity remains unclear. We investigated the synchronization mechanism(s) of DCR underlying untimely action potentials and diastolic contractions (DCs) in a catecholaminergic polymorphic ventricular tachycardia mouse model with a mutation in cardiac calsequestrin. We used a combination of different approaches including single ryanodine receptor channel recording, optical imaging (Ca(2+) and membrane potential), and contractile force measurements in ventricular myocytes and intact cardiac muscles. We demonstrate that DCR occurs in a temporally and spatially uniform manner in both myocytes and intact myocardial tissue isolated from cardiac calsequestrin mutation mice. Such synchronized DCR events give rise to triggered electrical activity that results in synchronous DCs in the myocardium. Importantly, we establish that synchronization of DCR is a result of a combination of abbreviated ryanodine receptor channel refractoriness and the preceding synchronous stimulated Ca(2+) release/reuptake dynamics. Our study reveals how aberrant DCR events can become synchronized in the intact myocardium, leading to triggered activity and the resultant DCs in the settings of a cardiac rhythm disorder.

Modulation of ventricular transient outward K⁺ current by acidosis and its effects on excitation-contraction coupling.

The contribution of transient outward current (Ito) to changes in ventricular action potential (AP) repolarization induced by acidosis is unresolved, as is the indirect effect of these changes on calcium handling. To address this issue we measured intracellular pH (pHi), Ito, L-type calcium current (ICa,L), and calcium transients (CaTs) in rabbit ventricular myocytes. Intracellular acidosis [pHi 6.75 with extracellular pH (pHo) 7.4] reduced Ito by ~50% in myocytes with both high (epicardial) and low (papillary muscle) Ito densities, with little effect on steady-state inactivation and activation. Of the two candidate α-subunits underlying Ito, human (h)Kv4.3 and hKv1.4, only hKv4.3 current was reduced by intracellular acidosis. Extracellular acidosis (pHo 6.5) shifted Ito inactivation toward less negative potentials but had negligible effect on peak current at +60 mV when initiated from -80 mV. The effects of low pHi-induced inhibition of Ito on AP repolarization were much greater in epicardial than papillary muscle myocytes and included slowing of phase 1, attenuation of the notch, and elevation of the plateau. Low pHi increased AP duration in both cell types, with the greatest lengthening occurring in epicardial myocytes. The changes in epicardial AP repolarization induced by intracellular acidosis reduced peak ICa,L, increased net calcium influx via ICa,L, and increased CaT amplitude. In summary, in contrast to low pHo, intracellular acidosis has a marked inhibitory effect on ventricular Ito, perhaps mediated by Kv4.3. By altering the trajectory of the AP repolarization, low pHi has a significant indirect effect on calcium handling, especially evident in epicardial cells.

Thrombopoietin modulates cardiac contractility in vitro and contributes to myocardial depressing activity of septic shock serum.

Thrombopoietin (TPO) is a humoral growth factor that has been shown to increase platelet activation in response to several agonists. Patients with sepsis have increased circulating TPO levels, which may enhance platelet activation, potentially participating to the pathogenesis of multi-organ failure. Aim of this study was to investigate whether TPO affects myocardial contractility and participates to depress cardiac function during sepsis. We showed the expression of the TPO receptor c-Mpl on myocardial cells and tissue by RT-PCR, immunofluorescence and western blotting. We then evaluated the effect of TPO on the contractile function of rat papillary muscle and isolated heart. TPO did not change myocardial contractility in basal conditions, but, when followed by epinephrine (EPI) stimulation, it blunted the enhancement of contractile force induced by EPI both in papillary muscle and isolated heart. An inhibitor of TPO prevented TPO effect on cardiac inotropy. Treatment of papillary muscle with pharmacological inhibitors of phosphatidylinositol 3-kinase, NO synthase, and guanilyl cyclase abolished TPO effect, indicating NO as the final mediator. We finally studied the role of TPO in the negative inotropic effect exerted by human septic shock (HSS) serum and TPO cooperation with TNF-alpha and IL-1beta. Pre-treatment with the TPO inhibitor prevented the decrease in contractile force induced by HSS serum. Moreover, TPO significantly amplified the negative inotropic effect induced by TNF-alpha and IL-1beta in papillary muscle. In conclusion, TPO negatively modulates cardiac inotropy in vitro and contributes to the myocardial depressing activity of septic shock serum.

The homologous rat chromogranin A1-64 (rCGA1-64) modulates myocardial and coronary function in rat heart to counteract adrenergic stimulation indirectly via endothelium-derived nitric oxide.

Chromogranin A (CGA), produced by human and rat myocardium, generates several biologically active peptides processed at specific proteolytic cleavage sites. A highly conserved cleavage N-terminal site is the bond 64-65 that reproduces the native rat CGA sequence (rCGA1-64), corresponding to human N-terminal CGA-derived vasostatin-1. rCGA1-64 cardiotropic activity has been explored in rat cardiac preparations. In Langendorff perfused rat heart, rCGA1-64 (from 33 nM) induced negative inotropism and lusitropism as well as coronary dilation, counteracting isoproterenol (Iso) - and endothelin-1 (ET-1) -induced positive inotropic effects and ET-1-dependent coronary constriction. rCGA1-64 also depressed basal and Iso-induced contractility on rat papillary muscles, without affecting calcium transients on isolated ventricular cells. Structure-function analysis using three modified peptides on both rat heart and papillary muscles revealed the disulfide bridge requirement for the cardiotropic action. A decline in Iso intrinsic activity in the presence of the peptides indicates a noncompetitive antagonistic action. Experiments on rat isolated cardiomyocytes and bovine aortic endothelial cells indicate that the negative inotropism observed in rat papillary muscle is probably due to an endothelial phosphatidylinositol 3-kinase-dependent nitric oxide release, rather than to a direct action on cardiomyocytes. Taken together, our data strongly suggest that in the rat heart the homologous rCGA1-64 fragment exerts an autocrine/paracrine modulation of myocardial and coronary performance acting as stabilizer against intense excitatory stimuli.

Both Purkinje cells and left ventricular posteroseptal reentry contribute to the maintenance of ventricular fibrillation in open-chest dogs and swine: effects of catheter ablation and the ventricular cut-and-sew operation.

BACKGROUND: Radiofrequency catheter ablation (RFCA) targeting the left ventricular posteroseptum (LVPS) and posterior papillary muscle (PPM) terminates or prevents ventricular fibrillation (VF) in rabbit and dog hearts. However, whether the mechanism of VF maintenance is reentry or focal Purkinje firing is unclear. METHODS AND RESULTS: In the present study the effects of RFCA (endocardial ablation of PPM+LVPS in 7 dogs and 7 swine), left ventricular anterolateral wall ablation (LVAL in 7 dogs), and the cut-and-sew operation (CSO: along the left ventricular posterior wall (LVPW) beside PPM in 7 swine) on VF inducibility were compared. (1) VF inducibility was decreased from 100+/-0% to 21.9+/-31.2% (p<0.0001) by PPM+LVPS endocardial ablation, but not by LVAL ablation in dogs. (2) LVPW CSO reduced VF inducibility (100+/-0% to 43.6+/-9.5%, p<0.0001) in swine. (3) In contrast to the canine Purkinje network, which is mostly localized to the subendocardium, the swine Purkinje network extends to the subepicardial layer with a higher density (p<0.001). CONCLUSION: Both PPM+LVPS ablation (Purkinje destruction) in dogs and LVPW CSO (blocking reentry) in swine reduce VF inducibility, suggesting that in both species focal firing from the Purkinje network and reentry around the PPM contributes to the maintenance of VF.

A framework for modeling of mechano-electrical feedback mechanisms of cardiac myocytes and tissues.

The term cardiac mechano-electrical feedback precis the various phenomena related to modulation of electrophysiology by mechanical deformation of cells and tissues of the heart. The significance of mechano-electrical feedback and the underlying mechanisms are still poorly understood despite intense experimental research. We introduce and discuss a framework for computational modeling and simulation of mechano-electrical feedback at ion channel, cell and tissue level. The framework consists of modules to reconstruct electrical currents through mechano-sensitive ion channels, their effect on myocytes' electrophysiology, strain-modulation of tissue conductivities and electrical conduction in myocyte clusters and myocardium. We applied the framework to study the effect of strain on conduction velocity in papillary muscle. The simulations reconstructed strain-conduction velocity relationships as reported in experimental studies. Furthermore, the computational studies indicated that increased stimulus frequency aggravated the reduction of conduction velocity for larger strains. Mathematical modeling of mechano-electrical feedback will help to integrate experimental data and predict behavior at system level. Computational simulations might give otherwise unavailable insights, particularly with respect to clinical relevance of mechano-electrical feedback.

Blebbistatin specifically inhibits actin-myosin interaction in mouse cardiac muscle.

Blebbistatin is a powerful inhibitor of actin-myosin interaction in isolated contractile proteins. To examine whether blebbistatin acts in a similar manner in the organized contractile system of striated muscle, the effects of blebbistatin on contraction of cardiac tissue from mouse were studied. The contraction of paced intact papillary muscle preparations and shortening of isolated cardiomyocytes were inhibited by blebbistatin with inhibitory constants in the micromolar range (1.3-2.8 muM). The inhibition constants are similar to those previously reported for isolated cardiac myosin subfragments showing that blebbistatin action is similar in filamentous myosin of the cardiac contractile apparatus and isolated proteins. The inhibition was not associated with alterations in action potential duration or decreased influx through L-type Ca(2+) channels. Experiments on permeabilized cardiac muscle preparations showed that the inhibition was not due to alterations in Ca(2+) sensitivity of the contractile filaments. The maximal shortening velocity was not affected by 1 muM blebbistatin. In conclusion, we show that blebbistatin is an inhibitor of the actin-myosin interaction in the organized contractile system of cardiac muscle and that its action is not due to effects on the Ca(2+) influx and activation systems.

Quantification of shock-induced microscopic virtual electrodes assessed by subcellular resolution optical potential mapping in guinea pig papillary muscle.

INTRODUCTION: The primary objective of this study was the quantitative description of shock-induced, locally occurring virtual electrodes in natural cardiac tissue. METHODS AND RESULTS: Multiscale optical potential mapping using 10x, 20x, and 40x magnifying objectives, achieving resolutions of 0.13, 0.065, and 0.033 mm, was performed when applying uniform shocks (+/-10 V/cm, 5 ms) during diastole and action potential plateau. A procedure was developed to identify local potential deviations as depolarizing or hyperpolarizing peaks and to quantify their occurrence and characteristic amplitudes, lateral extents, and dynamics. At shock onset, peaks of either polarity developed significantly faster (tau = 0.92 +/- 0.65 ms, N = 64) than the average bulk polarization (tau = 2.25 +/- 0.96 ms, P < 0.001) and appeared locally fixed, changing their polarity at shock reversal. The mean peak magnitude (21.2 +/- 12 mV) and the amplitude distribution were essentially independent from the magnification. The peak density continuously increased with decreasing peak extent (taken at 70% of the amplitude), reaching a maximum of approximately 3 peaks/mm2 in the range of approximately 30-65 microm. There was no correlation between peak amplitude and size throughout. Potentially exciting peaks were found with a density of 0.04-0.2 peaks/mm2 corresponding to estimated 1-5 peaks/mm3. CONCLUSIONS: Our results suggest that microscopic inhomogeneities form a substantial substrate for far-field excitation in natural cardiac tissue. Here, we effectively bridged the gap between the extensively studied myocyte cultures and larger heart preparations.

Cerivastatin and hypercholesterolemia reduce apoptosis of cardiomyocytes in guinea pig papillary muscle subjected to hypoxia/reoxygenation.

The aim of this study was to assess how cerivastatin influences contractility and degree of myocardial damage in papillary muscle subjected to hypoxia-reoxygenation in hypercholesterolemic guinea pigs. Study group consisted of guinea pigs, fed standard, hypercholesterolemic or hypercholesterolemic diets with low dose of cerivastatin. During experimental hypoxia-reoxygenation, the contractility was measured. Apoptosis of cardiomioctes was assessed with the use of TUNEL technique. Total cholesterol in standard, hypercholesterolemic and cerivastatin-treated group was 35 +/- 8 mg/dl, 131.6 +/- 30.4 mg/dl and 121.2 +/- 26.2 mg/dl, respectively, and was significantly higher in rats fed hypercholesterolemic and hypercholesterolemic + cerivastatin diets than in control group (p < 0.01). There were no significant differences between all analyzed groups in the post-ischemic cardiac function. Percentage of apoptotic cells after hypoxia-reoxygenation injury in groups fed standard, hypercholesterolemic and hypercholesterolemic + cerivastatin diets was 30 +/- 8%, 20 +/- 4% and 5 +/- 7%, respectively, and was significantly lower in groups that received hypercholesterolmic (p < 0.01) and hypercholesterolemic + cerivastatin (p < 0.001) diets in comparison with standard diet-fed group. In the group treated with cerivastatin, the percentage of apoptotic cells was additionally lower in comparison with hypercholesterolemic group (p < 0.01). Negative correlation between percentage of apoptotic cells and HDL level was found when all groups were considered jointly (r = -0.41, p < 0.05). Our study clearly shows that cerivastatin in hypercholesterolemic animals and hypercholesterolemia itself limit cardiomiocyte damage after hypoxia-reoxygenation.

Positive inotropic effect of levosimendan is correlated to its stereoselective Ca2+-sensitizing effect but not to stereoselective phosphodiesterase inhibition.

In order to clarify the mechanisms of the positive inotropic actions of levosimendan and its optical isomer, dextrosimendan, we compared their concentration-dependent effects in intact papillary muscles, permeabilized cardiomyocytes and in purified phosphodiesterase enzyme preparations of guinea-pig hearts. In papillary muscles twitch tension increased with EC50 values of 60 nM and 2.8 microM for levosimendan and dextrosimendan, respectively. Hence, the two enantiomers exhibited a 47 times potency difference in their positive inotropic effects in a preparation where theoretically Ca2+-sensitization and phosphodiesterase inhibition could both contribute to the positive inotropic effects. In guinea-pig cardiomyocytes, levosimendan and dextrosimendan increased isometric force production (at pCa 6.2) due to Ca2+-sensitization with EC50 values of 8.4 nM and 0.64 microM, respectively, with a similar relative potency difference of 76. A major difference appeared in their relative pharmacological potencies, however, when the inhibitory effects of the two enantiomers were assayed on phosphodiesterase III, purified from guinea pig left ventricle (i.e. the phosphodiesterase isoenzyme which is dominant in that tissue). Levosimendan was a 427 times more potent phosphodiesterase inhibitor than dextrosimendan, with IC50 values of 7.5 nM, and 3.2 microM, respectively. Taken together, our data support the hypothesis that levosimendan and dextrosimendan exert their positive inotropic effects via a stereoselective Ca2+-sensitizing mechanism and not via stereoselective inhibition of phosphodiesterase III in the myocardium.

[Cardiac remodeling and inflammation]. / Remodelación cardíaca e inflamación.

The cardiac remodeling is a progressive response of the heart to acute and chronic insults regardless its etiology. This process is characterized by changes in the size, shape and function and is associated with a worse prognosis in patients with heart failure. The acute myocardial infarction is the most common cause of remodeling. In the first minutes after injury in the ischemic zone there is an important augment in the synthesis and release of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) interleukin-6 (IL-6), interleukin-1-beta (IL-1beta) and transforming growth factor 1-beta (TGF-1beta). This acute releasing of cytokines could regulate the survival or apoptosis of myocytes in infarcted zone and, their negative inotropic effects could represent an adaptative response to delimit the injury and to decrease myocardial energy demand. This significant upregulation of proinflammatory cytokines can extend to noninfarcted zone and triggers a second phase of elevated levels of cytokines that promote interstitial fibrosis and collagen deposition in the contralateral noninfarcted myocardium leading to a dysfunctional ventricle. This article will review the recent reports that support the idea of a cardioprotective role for this early inflammatory response and a deleterious role of the delayed response that mediate the fibrosis that is a typical feature of the remodeling process.

Restricting excessive cardiac action potential and QT prolongation: a vital role for IKs in human ventricular muscle.

BACKGROUND: Although pharmacological block of the slow, delayed rectifier potassium current (IKs) by chromanol 293B, L-735,821, or HMR-1556 produces little effect on action potential duration (APD) in isolated rabbit and dog ventricular myocytes, the effect of IKs block on normal human ventricular muscle APD is not known. Therefore, studies were conducted to elucidate the role of IKs in normal human ventricular muscle and in preparations in which both repolarization reserve was attenuated and sympathetic activation was increased by exogenous dofetilide and adrenaline. METHODS AND RESULTS: Preparations were obtained from undiseased organ donors. Action potentials were measured in ventricular trabeculae and papillary muscles using conventional microelectrode techniques; membrane currents were measured in ventricular myocytes using voltage-clamp techniques. Chromanol 293B (10 micromol/L), L-735,821 (100 nmol/L), and HMR-1556 (100 nmol/L and 1 micromol/L) produced a <12-ms change in APD while pacing at cycle lengths ranging from 300 to 5000 ms, whereas the IKr blockers sotalol and E-4031 markedly lengthened APD. In voltage-clamp experiments, L-735,821 and chromanol 293B each blocked IKs in the presence of E-4031 to block IKr. The E-4031-sensitive current (IKr) at the end of a 150-ms-long test pulse to 30 mV was 32.9+/-6.7 pA (n=8); the L-735,821-sensitive current (IKs) magnitude was 17.8+/-2.94 pA (n=10). During a longer 500-ms test pulse, IKr was not substantially changed (33.6+/-6.1 pA; n=8), and IKs was significantly increased (49.6+/-7.24 pA; n=10). On application of an "action potential-like" test pulse, IKr increased as voltage became more negative, whereas IKs remained small throughout all phases of the action potential-like test pulse. In experiments in which APD was first lengthened by 50 nmol/L dofetilide and sympathetic activation was increased by 1 micromol/L adrenaline, 1 micromol/L HMR-1556 significantly increased APD by 14.7+/-3.2% (P<0.05; n=3). CONCLUSIONS: Pharmacological IKs block in the absence of sympathetic stimulation plays little role in increasing normal human ventricular muscle APD. However, when human ventricular muscle repolarization reserve is attenuated, IKs plays an increasingly important role in limiting action potential prolongation.

Both beta-adrenergic receptor stimulation and cardiac tissue type have important roles in elucidating the functional effects of I(Ks) channel blockers in vitro.

INTRODUCTION: Conflicting results associated with the use of I(Ks) blockers on the action potential duration (APD) have raised a question as to whether the variable results arise from the use of different cardiac tissues, beta-adrenergic stimulation, or by the "selectivity" of the chosen I(Ks) blockers. METHODS: We used the highly selective I(Ks) blocker (-)-[3R, 4S] chromanol 293B [(-) chromanol] to mimic drug-induced long QT1 in isolated rabbit Purkinje fibers, papillary muscles, and ventricular trabeculae using the conventional microelectrode technique. RESULTS: I(Ks) block with (-) chromanol at 1 x 10(-5) M did not significantly change the APD at different stimulation rates in all three cardiac tissues. Isoproterenol (Iso:1 x 10(-7) M) shortened APD(90), and (-) chromanol (1 x 10(-5) M) largely prevented this shortening in isolated papillary muscles at 1 Hz [-3% with Iso combined (-) chromanol group versus -16% with iso group; p<0.05] and also at 2 Hz (+7% versus -25% with Iso group; p<0.05), but did not significantly prevent this shortening in isolated Purkinje fibers. In isolated trabeculae, (-) chromanol combined with Iso significantly prolonged the APD(90) by 15% at 1 Hz (versus -10% with Iso group; p<0.05) and by 5% at 2 Hz (versus -11% with Iso group; p<0.05). DISCUSSION: Our study shows that only during beta-adrenoceptor stimulation, pharmacological inhibition of the I(Ks) current plays an important role in the APD recorded from isolated ventricular trabeculae and papillary muscles, but not from Purkinje fibers. These results indicate that the APD prolonging effects of I(Ks)channel blockers during beta-adrenergic receptor stimulation can only be detected from isolated rabbit papillary muscles and ventricular trabeculae, but not Purkinje fibers.

Effect of sustained hypobaric hypoxia during maturation and aging on rat myocardium. I. Mechanical activity.

Long-lasting cardioprotection may be attained by chronic hypoxia. The basal parameters of contractile function and their response to hypoxia/reoxygenation were measured under isometric conditions, in papillary muscles isolated from left ventricle of rats that were submitted to 53.8 kPa in a hypobaric chamber from 7 wk of age and for their lifetime and of their siblings kept at 101.3 kPa. During acclimatization, hematocrit increased, body weight gain decreased, and heart weight increased with right ventricle hypertrophy. Papillary muscle cross-sectional area was similar in both control and hypoxic groups up to 45 wk of exposure. Developed tension (DT) was 34-64% higher in rats exposed to hypoxia for 10, 26, and 45 wk than in their age-matched controls, whereas resting tension was unchanged. Maximal rates of contraction and relaxation showed a similar pattern of changes as DT. Recovery of DT and maximal rates of contraction and relaxation after 60-min hypoxia and 30-min reoxygenation was also improved in adult hypoxic rats to values similar to those of young rats. Heart acclimatization was lost after 74 wk of exposure. Results are consistent with the development of cardioprotection during high-altitude acclimatization and provide an experimental model to study the mechanisms involved, which are addressed in the accompanying paper.

Effect of sustained hypobaric hypoxia during maturation and aging on rat myocardium. II. mtNOS activity.

Mitochondrial nitric oxide (NO) production was assayed in rats submitted to hypobaric hypoxia and in normoxic controls (53.8 and 101.3 kPa air pressure, respectively). Heart mitochondria from young normoxic animals produced 0.62 and 0.37 nmol NO.min(-1).mg protein(-1) in metabolic states 4 and 3, respectively. This production accounts for a release to the cytosol of 29 nmol NO.min(-1).g heart(-1) and for 55% of the NO generation. The mitochondrial NO synthase (mtNOS) activity measured in submitochondrial membranes at pH 7.4 was 0.69 nmol NO.min(-1).mg protein(-1). Rats exposed to hypobaric hypoxia for 2-18 mo showed 20-60% increased left ventricle mtNOS activity compared with their normoxic siblings. Left ventricle NADH-cytochrome-c reductase and cytochrome oxidase activities decreased by 36 and 12%, respectively, from 2 to 18 mo of age, but they were not affected by hypoxia. mtNOS upregulation in hypoxia was associated with a retardation of the decline in the mechanical activity of papillary muscle upon aging and an improved recovery after anoxia-reoxygenation. The correlation of left ventricle mtNOS activity with papillary muscle contractility (determined as developed tension, maximal rates of contraction and relaxation) showed an optimal mtNOS activity (0.69 nmol.min(-1).mg protein(-1)). Heart mtNOS activity is regulated by O(2) in the inspired air and seems to play a role in NO-mediated signaling and myocardial contractility.