Normal Tyrode Solution Research Articles

Acute hyperglycemia induces Cardiac Mitochondrial Depolarization

Mitochondrial ATP generation depends on the enormous potential across the inner mitochondrial membrane (ΔΨmito ~ −180 mV), among many other factors. Oxidative phosphorylation requires the large ΔΨmito and when the mitochondria are depolarized, oxidative phosphorylation becomes uncoupled and ATP production falls. Here we investigate the effect of hyperglycemia (30 mM versus the physiological glucose of 5 mM) on mitochondrial ΔΨmito under photonic stress in rat ventricular myocytes. Using tetra-methyl rhodamine methyl ester (TMRM) as a fluorescence monitor of ΔΨmito we examined how the time-dependent depolarization of mitochondria may be altered by hyperglycemia. Using a normal Tyrode's solution on isolated ventricular myocytes, changing the [glucose] has dramatic effect on the rate of ΔΨmito depolarization. After 3hours of hyperglycemic exposure, the rate of depolarization is reduced nearly 30% compared to controls. Antioxidant treatment protects cardiac myocytes from this hyperglycemic-induced effect while selectively inhibiting complex I causes an abrogation of the effect. These data elucidate the biophysical effects of acute hyperglycemia on cardiac myocytes.

Effect of Matricaria chamomilla extract on the electrically stimulation of ileum of guinea-pig in invitro model

Previous studies indicate that Matricaria chamomilla (MC), a folk remedy plant, is very useful in the treatment of spasmogenic bowel diseases but the way of its action is not clear. For this purpose, we studied the effect of MC extract on the electrically activation in Guinea-Pig ileum in an invitro model. We used guinea pig in this study. After of kill, we brouhght out a piece of ileum and fixed it in organ bath. After adaptation of the tissue, we used electrical stimulation and recorded the response by physiograph. Stimulation of guinea pig ileum at 0.1 Hz in normal Tyrode solution resulted in increase of electrical activity and twitch contractions of this muscle. Results indicated that addition of MC during 0.1 Hz stimulation diminished and enhanced this electrical activity in a dose dependent manner. High concentration of MC significantly increased electrical stimulation of ileum (P

Effects of GBE50 on physiological characteristics and intracellular free calcium of myocardium

To investigate effects of GBE50 on the physiological characteristics and intracellular free calcium of myocardium. The preparations of isolated atria of guinea pigs were used for recording the myocardial physiological characteristics and the technique of Fluo-3/AM was used for measuring the concentration of free calcium in myocytes. GBE50 (10, 20, 40, 80, 160 mg x L(-1)) prominently inhibited the contractive force and decreased spontaneous beat of right atrium in concentration-dependent manner. GBE50 (10, 20, 40, 80, 160 mg x L(-1)) inhibited the contractive force of left atrium, and markedly decreased the post rest potentiation of myocardial contractility in left atrium in concentration-dependent manner. GBE50 (10, 20, 40, 80, 160 mg x L(-1)) prolonged the function refractory period of isolated guinea pig left atrium. 50 mg x L(-1) GBE50 decreased [Ca2+]i of guinea pig ventricular myocytes in normal Tyrode's solution by (46.53 +/- 8.44)% in 12 minutes. GBE50 induces inhibitory effects of the spontaneous beat, contractility, post-rest potentiation and prolonged the function refractory period. GBE50 may decrease [Ca2+]i of guinea pig ventricular myocytes.

Calcium oscillations and T-wave lability precede ventricular arrhythmias in acquired long QT type 2

Alternans of intracellular Ca(2+) (Ca(i)) underlies T-wave alternans, a predictor of cardiac arrhythmias. A related phenomenon, T-wave lability (TWL), precedes torsades de pointes (TdP) in patients and animal models with impaired repolarization. However, the role of Ca(i) in TWL remains unexplored. This study investigated the role of Ca(i) dynamics on TWL in a noncryoablated rabbit model of long QT syndrome type 2 (LQT2) using simultaneous measurements of Ca(i) transient (CaT), action potentials (APs), and electrocardiogram (ECG) during paced rhythms and focused on events that precede ventricular ectopy. APs and CaTs were mapped optically from paced Langendorff female rabbit hearts (n = 8) at 1.2-s cycle length, after atrioventricular node ablation. Hearts were perfused with normal Tyrode solution, then with dofetilide (0.5 μM), and reduced [K(+)] (2 mM) and [Mg(2+)] (0.5 mM) to elicit LQT2. Lability of ECG, voltage, and Ca(i) signals were evaluated during regular paced rhythm, before and after dofetilide perfusion. In LQT2, lability of Ca(i), voltage, and ECG signals increased during paced rhythm, before the appearance of early afterdepolarizations (EADs). LQT2 resulted in AP prolongation and multiple (1 to 3) additional Ca(i) upstrokes, whereas APs remained monophasic. When EADs appeared, Ca(i) rose before voltage upstrokes at the origins of propagating EADs. Interventions (i.e., ryanodine and thapsigargin, n = 3 or low [Ca](o) and nifedipine, n = 4) that suppressed Ca(i) oscillations also abolished EADs. In LQT2, Ca(i) oscillations (Ca(i)O) precede EADs by minutes, indicating that they result from spontaneous sarcoplasmic reticulum Ca(2+) release rather than spontaneous I(Ca,L) reactivation. Ca(i)O likely produce oscillations of Na/Ca exchange current, I(NCX). Depolarizing I(NCX) during the AP plateau contributes to the generation of EADs by reactivating Ca(2+) channels that have recovered from inactivation. TWL reflects CaTs and APs lability that occur before EADs and TdP.

Effects of Guanxinkang on expressions of ATP-sensitive potassium channel subunits Kir6.1, Kir6.2, SUR2A and SUR2B in ischemic myocytes of rats

To observe the effects of Guanxinkang injection, a compound traditional Chinese herbal medicine, on ATP-sensitive potassium (K(ATP)) channel subunits in ischemic myocardial cells of rats, and to explore the mechanism of Guanxinkang in protecting myocardial ischemic reperfusion injuries. Forty-eight Wistar rats were randomly divided into normal group, untreated group, glibenclamide group, pinacidil group, Guanxinkang group and Guanxinkang plus glibenclamide group. The ventricular myocytes were prepared from hearts of normal rats by enzymatic dissociation method. The ischemic ventricular myocytes underwent perfusion with normal Tyrode solution for 10 min, then stopping perfusion 30 min, and followed by 45 min of reperfusion. The glibenclamide, pinacidil and Guanxinkang were added into ventricular myocytes solution directly. Then the solutions were placed at 4 degrees centigrade. After 24-hour freezing at -80 degrees centigrade, mRNA and protein expressions of KATP subunits Kir6.1, Kir6.2, SUR2A and SUR2B were measured by real-time quantitative polymerase chain reaction and Western blotting respectively. In normal rat myocardial cells, there were SUR2A, Kir6.1, and Kir6.2 protein and gene expressions but no expression of SUR2B protein. In the untreated group, all subunit mRNA and protein expressions of KATP increased to some extent as compared with the normal group. Pinacidil, a potassium channel opener, significantly increased mRNA and protein expressions of KATP subunits, while the blocker glibenclamide had a reverse effect. Meanwhile, Guanxinkang injection significantly increased mRNA and protein expressions of K(ATP) subunits but with no significant difference as compared with pinacidil. Guanxinkang injection can obviously enhance the open of KATP channel and thus play a role in cardiovascular protection.

Inhibition of oxotremorine-induced desensitization of guinea-pig ileal longitudinal muscle in Ca2+-free conditions.

The aim of this study was to investigate the differences between oxotremorine-induced and acetylcholine (ACh)-induced desensitization, particularly under Ca2+-free conditions, in guinea-pig ileal longitudinal muscle, and to elucidate the different mechanisms of desensitization that might exist between these two muscarinic agonists. Pretreatment of the tissue with 10(-7)-10(-5) M oxotremorine (desensitizing treatment) in normal Tyrode solution caused desensitization of the responses to ACh, as did the desensitizing treatment with ACh. However, Ca2+-free conditions significantly reduced oxotremorine-induced desensitization, contrary to the previous findings that Ca2+-free conditions enhanced ACh-induced desensitization. The desensitizing treatment with oxotremorine caused suppression of the responses to high K+ (tonic phase), as did the ACh treatment. Ca2+-free conditions removed this suppression, whereasthis condition enhanced ACh-induced suppression of the K+ response. A protein kinase C inhibitor, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (10(-4) M) had no effect on oxotremorine-induced desensitization of the ACh response. The results suggest that a voltage-gated Ca2+ channel was involved in oxotremorine-induced desensitization, as in ACh-induced desensitization, but that the process of inactivation of Ca2+ channels was different between oxotremorine and ACh, and that oxotremorine-induced desensitization was due not only to Ca2+ channel, but also to other unknown factors. Protein kinase C did not participate in oxotremorine-induced desensitization.

Abstract 5402: Two Photon Microscopy Measurements Of Sub-epicardial Sarcomere Length In Perfused Rat Hearts

Sarcomere length (SL) is a fundamental parameter underlying the Frank Starling relation in the heart, as it offers an absolute representation of myocardial stretch. Previous studies addressed the Frank Starling relation by measuring SL in isolated myocytes or muscle strips. Here, we report first data obtained using a novel technique to measure sub-epicardial SL in perfused hearts. Rat hearts were Langendorff perfused (normal Tyrode solution) at a constant pressure of 90mmHg, labeled with the fluorescent membrane marker di-4-ANEPPS, and then arrested with high-K + Tyrode for either 2-photon microscopy (n=4) or MRI (n=4). Image analysis software was developed to extract SL at the cell level from >1,400 2-photon images (Fig 1 ) and correct for cell angle. SL increased by 10±2 % between 30 and 80 min of perfusion (1.98±0.04 to 2.17±0.03 μm; p<0.05; Fig 1 ). Measurements of left ventricular myocardial volume (LVMV) were made in vivo and in perfused hearts using 3D MRI. LVMV increased by 24±7% from in vivo to 30 min of perfusion, and by 11±3 % between 30 and 90 min (539±35; 664±44; 737±49 mm 3 , respectively; p<0.05; Fig 1 ). We show that SL can be measured in isolated perfused hearts. The method allowed monitoring of changes in SL over time, and showed that SL and LVMV increase to a similar extent during 30–80 min perfusion with crystalloid solution, probably due to tissue oedema. This result, together with the increase in LVMV during the first 30 min, highlights the pronounced differences between in vivo , in situ , and in vitro model systems for studies of cardiac physiology and mechanics. Future research will compare changes in SL in healthy hearts and disease models involving contractile dysfunction. Figure 1: Left: 2-photon microscopy image of di-4-ANEPPS labeled myocardium. Right: SL and LVMV changes over time.

Chemical ablation of the Purkinje system causes early termination and activation rate slowing of long-duration ventricular fibrillation in dogs

Endocardial mapping has suggested that Purkinje fibers may play a role in the maintenance of long-duration ventricular fibrillation (LDVF). To determine the influence of Purkinje fibers on LDVF, we chemically ablated the Purkinje system with Lugol solution and recorded endocardial and transmural activation during LDVF. Dog hearts were isolated and perfused, and the ventricular endocardium was exposed and treated with Lugol solution (n = 6) or normal Tyrode solution as a control (n = 6). The left anterior papillary muscle endocardium was mapped with a 504-electrode (21 x 24) plaque with electrodes spaced 1 mm apart. Transmural activation was recorded with a six-electrode plunge needle on each side of the plaque. Ventricular fibrillation (VF) was induced, and perfusion was halted. LDVF spontaneously terminated sooner in Lugol-ablated hearts than in control hearts (4.9 +/- 1.5 vs. 9.2 +/- 3.2 min, P = 0.01). After termination of VF, both the control and Lugol hearts were typically excitable, but only short episodes of VF could be reinduced. Endocardial activation rates were similar during the first 2 min of LDVF for Lugol-ablated and control hearts but were significantly slower in Lugol hearts by 3 min. In control hearts, the endocardium activated more rapidly than the epicardium after 4 min of LDVF with wave fronts propagating most often from the endocardium to epicardium. No difference in transmural activation rate or wave front direction was observed in Lugol hearts. Ablation of the subendocardium hastens VF spontaneous termination and alters VF activation sequences, suggesting that Purkinje fibers are important in the maintenance of LDVF.

Stimulus-evoked opioid inhibition in guinea-pig longitudinal muscle-myenteric plexus strip is modulated by NMDA receptors

Longitudinal muscle-myenteric plexus (LMMP) strips of the guinea-pig ileum were used to investigate the stimulus-evoked endogenous opioid inhibition and its modulation by ionotropic glutamate receptors. Regular cholinergic twitch responses evoked by a short 3-s-field stimulation in intervals of 80 s were found reduced after an interposed period of prolonged 40-s-field stimulation. In the presence of a peptidase-inhibitor-cocktail, the cholinergic twitch response following the period of prolonged stimulation was even further reduced as compared to normal Tyrode solution without peptidase inhibitors. In both instances, the impairment of the cholinergic twitch response was completely abolished by naloxone thus demonstrating its opioidergic nature. This endogenous inhibitory opioid effect was significantly mitigated by the NMDA-receptor antagonist MK-801, but not by the AMPA/kainate receptor antagonist CNQX. These results demonstrate by functional experiments that there is a significant opioid-mediated inhibition in guinea-pig LMMP preparations evoked by a prolonged electrical stimulation, and that an NMDA antagonist can mitigate the opioid inhibition.

FUNCTIONAL IMPAIRMENT OF CARDIAC TRANSIENT OUTWARD K+ CURRENT AS A RESULT OF ABNORMALLY ALTERED CELLULAR ENVIRONMENT

1. Physiological functions of cardiac cells require a normal cellular environment. Under pathological conditions, there is a loss of normal cellular environment due to metabolic perturbations and other abnormalities. To test the hypothesis that cellular environmental stresses can create an electrophysiological substrate for electrical disorders in the heart, we investigated the effects of hypoxia, acidosis and ischaemia on transient outward K+ current (I(to)) in single canine ventricular myocytes. 2. The I(to) was studied because it plays a critical role in initiating cardiac repolarization and, thereby, arrhythmias. It was found that I(to) was significantly depressed by some 30% under hypoxic conditions relative to that in a normal cellular environment with normal Tyrode's solution. 3. Acidosis created by lowering the pH of the external solution from 7.4 to 7.2 produced a substantial (approximately 35%) reduction of the I(to) amplitude. 4. A marked impairment of I(to) function was consistently observed in ischaemic hearts in the canine coronary artery ligation model, with an approximate 30% decrease in the size of I(to). 5. Importantly, the impairment of I(to) under these environmental stresses was largely reversible following restoration to normal conditions. 6. The results of the present study suggest that I(to) is susceptible to changes in the cellular environment and the functional impairment of I(to) under environmental stresses contributes to arrhythmias under relevant pathological conditions of the heart.

Nitroglycerin on contractile force in canine cardiac Purkinje fibers

The objective of this study was to investigate the effects of nitroglycerin (NG) on the contractile force in order to elucidate regulatory roles of cyclic guanosine monophosphate (cGMP) in the bundle of canine cardiac Purkinje fibers. NG suppressed the developed tension in canine Purkinje fibers immersed in normal Tyrode's solution. The contraction was also elicited in the depolarizing solution containing 30 mM K+ by adding isoproterenol (Iso) or tetraethylammonium (TEA). The suppression caused by NG was more marked in fibers in the depolarizing solution treated with TEA than in the depolarizing solution treated with Iso. 8-Bromo-cGMP and sodium nitroprusside had a slight inhibition on the contraction in the fibers. Methylene blue did not affect the decrease in contractile force induced by NG. The weaker inhibition caused by NG on the contraction in the presence of Iso might result from a possible increase in intracellular cAMP levels in the fibers.

Myocyte Enhancer Factors 2A and 2C Induce Dilated Cardiomyopathy in Transgenic Mice

Cardiac hypertrophy and dilation are mediated by neuroendocrine factors and/or mitogens as well as through internal stretch- and stress-sensitive signaling pathways, which in turn transduce alterations in cardiac gene expression through specific signaling pathways. The transcription factor family known as myocyte enhancer factor 2 (MEF2) has been implicated as a signal-responsive mediator of the cardiac transcriptional program. For example, known hypertrophic signaling pathways that utilize calcineurin, calmodulin-dependent protein kinase, and MAPKs can each affect MEF2 activity. Here we demonstrate that MEF2 transcription factors induced dilated cardiomyopathy and lengthening of myocytes. Specifically, multiple transgenic mouse lines with cardiac-specific overexpression of MEF2A or MEF2C presented with cardiomyopathy at base line or were predisposed to more fulminant disease following pressure overload stimulation. The cardiomyopathic response associated with MEF2A and MEF2C was not further altered by activated calcineurin, suggesting that MEF2 functions independently of calcineurin in this response. In cultured cardiomyocytes, MEF2A, MEF2C, and MEF2-VP16 overexpression induced sarcomeric disorganization and focal elongation. Mechanistically, MEF2A and MEF2C each programmed similar profiles of altered gene expression in the heart that included extracellular matrix remodeling, ion handling, and metabolic genes. Indeed, adenoviral transfection of cultured cardiomyocytes with MEF2A or of myocytes from the hearts of MEF2A transgenic adult mice showed reduced transient outward K(+) currents, consistent with the alterations in gene expression observed in transgenic mice and partially suggesting a proximal mechanism underlying MEF2-dependent cardiomyopathy.

Overexpression of neuronal calcium sensor‐1 alters contractile function in adult rat ventricular myocytes

Neuronal-calcium sensor-1 (NCS-1), a small calcium binding protein, has been shown to interact with ion channels (including Ca channels) in neuronal cells. We have previously reported that NCS-1 was also present in rat cardiomyocytes. The present study examined whether NCS-1 overexpression in ventricular myocytes alters the contractile function. Isolated adult rat ventricular myocytes were infected with an adenovirus construct expressing NCS-1 (MOI: 100). At 48 hours after infection, myocytes were rinsed with control culture medium, and cell shortening was elicited in normal Tyrode's solution with field stimulation. The kinetics of steady-state cell shortening and the post-rest potentiation were monitored, and compared with those of non-infected time-control myocytes. Our results show that the rise time (10–90% of time-to-peak) and half-width (50% of duration) of cell shortening in NCS-1-infected cells were significantly reduced (31.1±0.9 and 57.3±2.3 ms vs. 37.5±1.5 and 68.7±2.8 ms in control cells, respectively, n=23 from 5 hearts). However, the decay time (10–90% of relaxation) was not significantly altered in the NCS-1-infected cells. Post-rest potentiation in NCS-1 cells was significantly increased after a 60-sec rest period (P=0.037, n=18), but not after a 30-sec rest (P=0.076, n=22). Taken together, our data suggest that NCS-1 alters the contractile function by facilitating contraction, with a slight alteration of sarcoplasmic reticular function in adult rat ventricular myocytes.

Sodium-Calcium Exchange Initiated by the Ca2+Transient: An Arrhythmia Trigger Within Pulmonary Veins

The hypothesis that an increased or prolonged Ca2+ transient during an abbreviated action potential can give rise to early afterdepolarizations (EADs) and triggered arrhythmia by enhanced forward sodium-calcium (Na-Ca) exchange was examined. Because pulmonary veins have the shortest action potential of any cardiac tissue, we examined this hypothesis in canine pulmonary vein sleeves during interventions further shortening the action potential and increasing the calcium transient. Extracellular bipolar electrode, intracellular microelectrode, and isometric force (a surrogate marker for the Ca2+ transient) recordings were obtained from superfused canine pulmonary veins. An elevation and prolongation of the terminal phase of repolarization (EADs) were observed during interventions increasing contractile force; isoproterenol or norepinephrine (3.2 x 10(-11) to 3.2 x 10(-7)M), hypothermia, and pacing (post-extrasystolic potentiation, post-pacing pause). The EAD formation was prevented by ryanodine (10 microM) or reversed by transiently increasing [Ca2+](o) from 1.35 to 5 mM (inhibition of forward Na-Ca exchange). Pacing-induced EADs were enhanced by re-introduction of normal Tyrode solution (Na+ = 130 mM) after substitution of 30 mM NaCl with 30 mM LiCl (stimulation of forward Na-Ca exchange). With norepinephrine or isoproterenol (3.2 x 10(-8)M) + acetylcholine (10(-7)M) (to enhance the Ca2+ transient and further shorten the abbreviated action potential, respectively), tachycardia-pause initiated arrhythmia (1,132 +/- 153 beats/min) lasting >1 s was observed. Rapid firing was prevented by either suppression of the Ca2+ transient (ryanodine) or transiently increasing [Ca2+](o). The data show EAD formation in superfused canine pulmonary veins, enhanced by an increased Ca2+ transient and increased Na-Ca exchange current. With subsequent shortening of the action potential with acetylcholine, tachycardia-pause triggers rapid firing within the PV sleeve.

Founder of cardiac cellular electrophysiology: Honouring Silvio Weidmann, 7 April 1921– 11 July 2005

Founder of cardiac cellular electrophysiology: Honouring Silvio Weidmann, 7 April 1921– 11 July 2005

The Different Effects of Phenothiazines on Cardiac Action Potential Duration

The Different Effects of Phenothiazines on Cardiac Action Potential Duration

Effects of halothane, isoflurane, sevoflurane and desflurane on contraction of ventricular myocytes from streptozotocin-induced diabetic rats.

Various clinically used volatile general anaesthetics (e.g. sevoflurane, halothane, isoflurane and desflurane) have been shown to have significant negative inotropic effects on normal ventricular muscle. However, little is known about their effects in ventricular tissue from diabetic animals. Streptozotocin (STZ)-induced diabetes is known to induce changes in the amplitude and time course of shortening and one report suggests that the inotropic effects of anaesthetics are ameliorated in papillary muscles from diabetic animals. The aim of these studies was to investigate this further in electrically stimulated (1 Hz) ventricular myocytes. Cells were superfused with either normal Tyrode (NT) solution or NT containing anaesthetic (1 mM) for a period of 2 min (at 30-32 degrees C). Myocytes from STZ rats were shown to have a significantly longer time to peak shortening (p > 0.001, n = 50) and the amplitude of shortening tended to be greater but this was not significant (p = 0.13, n = 50). Halothane, isoflurane, desflurane and sevoflurane significantly (p < 0.05) reduced the magnitude of shortening of control cells by 72.5 +/- 3.2%, 46.5 +/- 9.7%, 28.9 +/- 4.3% and 22.8 +/- 5.6%, respectively (n > 11 per group) but their steady-state negative inotropic effect was found to be no different in cells from STZ-treated rats (73.0 +/- 4.8%, 40.7 +/- 4.7%, 25.0 +/- 5.2% and 19.8 +/- 5.2%, respectively, n > 10 per group). Therefore, we conclude that the inotropic effects of volatile anaesthetics were not altered by STZ treatment.

Effects of acidosis on ventricular myocyte shortening and intracellular Ca2+ in streptozotocin-induced diabetic rats.

We have investigated the effects of acute acidosis on ventricular myocyte shortening and intracellular Ca2+ in streptozotocin (STZ)-induced diabetic rat. Shortening and intracellular Ca2+ were measured in electrically stimulated myocytes superfused with either normal Tyrode solution pH adjusted to either 7.4 (control solution) or 6.4 (acid solution). Experiments were performed at 35-36 degrees C. At 8-12 weeks after treatment, the rats that received STZ had lower body and heart weights compared to controls, and blood glucose was characteristically increased. Contractile defects in myocytes from diabetic rat were characterized by prolonged time to peak shortening. Superfusion of myocytes from control and diabetic rats with acid solution caused a significant reduction in the amplitude of shortening; however, the magnitude of the response was not altered by STZ treatment. Acid solution also caused significant and quantitatively similar reductions in the amplitude of Ca2+ transients in myocytes from control and diabetic rats. Effects of acute acidosis on amplitude of myocyte contraction and Ca2+ transient were not significantly altered by STZ treatment. Altered myofilament sensitivity to Ca2+ and altered mechanisms of sarcoplasmic reticulum Ca2+ transport might partly underlie the acidosis-evoked reduction in amplitude of shortening in myocytes from control and STZ-induced diabetic rat.

O2 consumption of mechanically unloaded contractions of mouse left ventricular myocardial slices.

Left ventricular (LV) myocardial slices were isolated from murine hearts (300 microm thick) and were stimulated at 1 Hz without external load. Mean myocardial slice O(2) consumption (MVo(2)) per minute (mMVo(2)) without stimulation was 0.97 +/- 0.14 ml O(2).min(-1).100 g LV(-1) and mean mMVo(2) with stimulation increased to 1.80 +/- 0.17 ml O(2).min(-1).100 g LV(-1) in normal Tyrode solution. Mean DeltamVo(2) (the mMVo(2) with stimulation - the mMVo(2) without stimulation) was 0.83 +/- 0.12 ml O(2).min(-1).100 g LV(-1). There were no differences between mean mMVo(2) with and without stimulation in Ca(2+)-free solution. The increases in extracellular Ca(2+) concentrations up to 14.4 mM did not affect the mMVo(2) without stimulation but significantly increased the mMVo(2) with stimulation up to 140% of control. The DeltamMVo(2) significantly increased up to 190% of the control in a dose-dependent manner. In contrast, the shortening did not increase in a dose-dependent manner. Cyclopiazonic acid (CPA; 30 microM) significantly reduced the DeltamMVo(2) to 0.27 +/- 0.06 ml O(2).min(-1).100 g LV(-1) (35% of control). The combination of 5 mM 2,3-butanedione monoxime (BDM) and 30 microM CPA did not further decrease DeltamMVo(2). Although BDM (3-5 mM) decreased the DeltamMVo(2) by 28-30% of control in a dose-independent manner, 3-5 mM BDM decreased shortening in a dose-dependent manner. Our results indicate that the DeltamMVo(2) of mouse LV slices during shortening under mechanically unloaded conditions consists of energy expenditure for total Ca(2+) handling during excitation-contraction coupling, basal metabolism, but no residual cross-bridge cycling.

Direct myocardial depressant effect of naloxone: mechanical and electrophysiological actions in vitro.

Naloxone, an opioid antagonist, has a cardiovascular pressor effect and has been used in various types of shock states. The aim was to determine the non-opioid effect on direct cardiac muscle contractility and explore the mechanism using guinea pig right ventricular papillary muscles. With institutional approval, isometric contractile force was measured in modified normal and 26 mM K+ Tyrode solution at various stimulation rates. The effects of naloxone on sarcoplasmic recticulum function were evaluated by measuring rested-state contraction in low Na+ (25 mM) Tyrode solution and rapid cooling contracture in modified normal Tyrode solution. Normal and slow action potentials (APs) were measured using conventional microelectrode technique. Patch clamp study was performed to examine the direct effect on Ca2+ current in cardiac ventricular myocytes. Naloxone (50, 100, and 200 microM) caused a concentration-dependent depression of peak force and maximal rate of force development. Modest depression, approximately 20%, was shown in rested-state contraction in low Na+ Tyrode solution. Naloxone (100 microM) modestly depressed the rapid cooling contracture to 80 +/- 3% of baseline, accompanied by prolongation of time to peak contracture by approximately 37%. In 26 mM K+ Tyrode solution, naloxone (100 microM) markedly and selectively depressed the late force development. While naloxone (100 microM) did not alter the amplitude and dV/dt-max in normal and slow APs at 0.25 Hz, AP duration was prolonged significantly. In patch clamp experiment, naloxone (50 microM) depressed Ca2+ current by approximately 50%. The direct myocardial depressant effect of naloxone appeared to be in part caused by depression of Ca2+ influx through cardiac membrane. Sarcoplasmic reticulum function appeared to be modestly depressed.