Initial Positive Inotropic Effect Research Articles

Circulating microRNAs predispose to takotsubo syndrome following high-dose adrenaline exposure.

Aims Takotsubo syndrome (TTS) is an acute heart failure, typically triggered by high adrenaline during physical or emotional stress. It is distinguished from myocardial infarction (MI) by a characteristic pattern of ventricular basal hypercontractility with hypokinesis of apical segments, and in the absence of culprit coronary occlusion. We aimed to understand whether recently discovered circulating biomarkers miR-16 and miR-26a, which differentiate TTS from MI at presentation, were mechanistically involved in the pathophysiology of TTS.Methods and results miR-16 and miR-26a were co-overexpressed in rats with AAV and TTS induced with an adrenaline bolus. Untreated isolated rat cardiomyocytes were transfected with pre-/anti-miRs and functionally assessed. Ventricular basal hypercontraction and apical depression were accentuated in miR-transfected animals after induction of TTS. In vitro miR-16 and/or miR-26a overexpression in isolated apical (but not basal), cardiomyocytes produced strong depression of contraction, with loss of adrenaline sensitivity. They also enhanced the initial positive inotropic effect of adrenaline in basal cells. Decreased contractility after TTS-miRs was reproduced in non-failing human apical cardiomyocytes. Bioinformatic profiling of miR targets, followed by expression assays and functional experiments, identified reductions of CACNB1 (L-type calcium channel Cavβ subunit), RGS4 (regulator of G-protein signalling 4), and G-protein subunit Gβ (GNB1) as underlying these effects.ConclusionmiR-16 and miR-26a sensitize the heart to TTS-like changes produced by adrenaline. Since these miRs have been associated with anxiety and depression, they could provide a mechanism whereby priming of the heart by previous stress causes an increased likelihood of TTS in the future.

Possible reasons for the variability of the inotropic insulin effect in papillary muscles of ground squirrel myocardium

The effects of insulin (0.1-50 nM) on isometric twitch force (0.1 to 1.0 Hz; 30 +/- 1 degree C; 1.8 mM Ca(2+)) were studied in right ventricular papillary muscles from active ground squirrels of different seasons (summer, n = 14; autumn, n = 16 and winter interbout, n = 16) in control conditions and after one-hour pretreatment of PM with 2 mkM nifedipine (an L-type Ca(2+)-channel inhibitor) and 1.0 mM orthovanadate (a tyrosine phosphatase inhibitor). In active animals of different seasonal periods insulin causes both positive and negative inotropic effects. At low frequencies (0.1-0.5 Hz), insulin of low concentrations (0.1-1.0 nM) induces a transient (within the first 20 min after application) positive effect (about 15-25%). Application of high hormone concentration (10 nM) in a low range of stimulation frequencies causes a biphasic effect (a small initial positive inotropic effect followed by a marked negative one). At frequencies above 0.5-Hz stimulation, insulin of 10 nM concentration causes presumably a negative inotropic effect. It was proposed that ICaL is possibly involved in the insulin-induced negative inotropy in ground squirrels hearts. Alteration of protein phosphorylation in tyrosine residues is known to be a major link in the mechanism of insulin action. We performed a study on orthovanadate action (a known inhibitor of tyrosine phosphatase) on the inotropic insulin effect. In the group of summer animals the pretreatment of papillary muscles with orthovanadate (100 mkM) does not change the negative inotropic effect of insulin in a low range of stimulation frequencies but almost completely removes this effect at stimulation frequencies above 0.3 Hz (n = 4). Nifedipine (1-1.5 hr pretreatment), a blocker of L-type calcium channel, reduces the inhibitory effect of insulin in autumn and winter animals, and on the contrary intensifies it in summer animals. This fact indicates that different mechanisms must be involved in insulin actions in animals of summer and winter periods. The main findings of the present study are that insulin induces positive, negative or no inotropic effects in papillary muscles of ground squirrels myocardium. The character of the effects of insulin depends on the physiological state of animals; time and concentrations of the hormone applied; affected by conditions that alter cellular Ca(2+) loading and the ratio of protein-tyrosine kinases/phosphatases activity.

Mechanisms of cardiodepression by an Na+–H+exchange inhibitor methyl-N-isobutyl amiloride (MIA) on the heart: lack of beneficial effects in ischemia–reperfusion injuryThis paper is one of a selection of papers published in this Special Issue, entitled Young Investigators' Forum.

Although Na+-H+ exchange (NHE) inhibitors such as methyl-N-isobutyl amiloride (MIA) are known to depress the cardiac function, the mechanisms of their negative inotropic effect are not completely understood. In this study, isolated rat hearts were perfused with MIA to study its action on cardiac performance, whereas isolated subcellular organelles such as sarcolemma, myofibrils, sarcoplasmic reticulum, and mitochondria were treated with MIA to determine its effect on their function. The effect of MIA on intracellular Ca2+ mobilization was examined in fura-2-AM-loaded cardiomyocytes. MIA was observed to depress cardiac function in a concentration-dependent manner in HCO3- -free buffer. On the other hand, MIA had an initial positive inotropic effect followed by a negative inotropic effect in HCO3-containing buffer. MIA increased the basal concentration of intracellular Ca2+ ([Ca2+]i) and augmented the KCl-mediated increase in [Ca2+]i. MIA did not show any direct effect on myofibrils, sarcolemma, and sarcoplasmic reticulum ATPase activities; however, this agent was found to decrease the intracellular pH, which reduced the myofibrils Ca2+-stimulated ATPase activity. MIA also increased Ca2+ uptake by mitochondria without having any direct effect on sarcoplasmic reticulum Ca2+ uptake. In addition, MIA did not protect the hearts subjected to mild Ca2+ paradox as well as ischemia-reperfusion-mediated injury. These results suggest that the increase in [Ca2+]i in cardiomyocytes may be responsible for the initial positive inotropic effect of MIA, but its negative inotropic action may be due to mitochondrial Ca2+ overloading as well as indirect depression of myofibrillar Ca2+ ATPase activity. Thus the accumulation of [H+]i as well as occurrence of intracellular and mitochondrial Ca2+ overload may explain the lack of beneficial effects of MIA in preventing the ischemia-reperfusion-induced myocardial injury.

Antagonistic Actions of Halothane and Sevoflurane on Spontaneous Ca2+Release in Rat Ventricular Myocytes

Halothane has been reported to sensitize Ca(2+) release from the sarcoplasmic reticulum (SR), which is thought to contribute to its initial positive inotropic effect. However, little is known about whether isoflurane or sevoflurane affect the SR Ca(2+) release process, which may contribute to the inotropic profile of these anesthetics. Mild Ca(2+) overload was induced in isolated rat ventricular myocytes by increase of extracellular Ca(2+) to 2 mM. The resultant Ca(2+) transients due to spontaneous Ca(2+) release from the SR were detected optically (fura-2). Cells were exposed to 0.6 mM anesthetic for a period of 4 min, and the frequency and amplitude of spontaneous Ca(2+) transients were measured. Halothane caused a temporary threefold increase in frequency and decreased the amplitude (to 54% of control) of spontaneous Ca(2+) transients. Removal of halothane inhibited spontaneous Ca release before it returned to control. In contrast, sevoflurane initially inhibited frequency of Ca(2+) release (to 10% of control), whereas its removal induced a burst of spontaneous Ca(2+) release. Isoflurane had no significant effect on either frequency or amplitude of spontaneous Ca(2+) release on application or removal. Sevoflurane was able to ameliorate the effects of halothane on the frequency and amplitude of spontaneous Ca(2+) release both on application and wash-off. Application of halothane and removal of sevoflurane sensitize the SR Ca(2+) release process (and vice versa on removal). Sevoflurane reversed the effects of halothane, suggesting they may act at the same subcellular target on the SR.

Dual effects of mesenteric lymph isolated from rats with burn injury on contractile function in rat ventricular myocytes

Gut-derived factors in intestinal lymph have been shown to trigger myocardial contractile dysfunction. However, the underlying cellular mechanisms remain unclear. We examined the effects of physiologically relevant concentrations of mesenteric lymph collected from rats with 40% burn injury (burn lymph) on excitation-contraction coupling in rat ventricular myocytes. Burn lymph (0.1-5%), but not control mesenteric lymph from sham-burn animals, induced dual positive and negative inotropic effects depending on the concentrations used. At lower concentrations (<0.5%), burn lymph increased the amplitude of myocyte contraction (1.6 +/- 0.3-fold; n = 12). At higher concentrations (>0.5%), burn lymph initially enhanced myocyte contraction, which was followed by a block of contraction. These effects were partially reversible on washout. The initial positive inotropic effect was associated with a prolongation of action potential duration (measured at 90% repolarization, 2.5 +/- 0.6-fold; n = 10), leading to significant increases in the net Ca2+ influx (1.7 +/- 0.1-fold; n = 8). There were no significant changes in the resting membrane potential. The negative inotropic effect was accompanied by a decrease in the action potential plateau (overshoot decrease by 69 +/- 10%; n = 4) and membrane depolarization. Voltage-clamp experiments revealed that the positive inotropic effects of burn lymph were due to an inhibition of the transient outward K+ currents that prolong action potential duration, and the inhibitory effects were due to a concentration-dependent inhibition of Ca2+ currents that lead to a reduction of action potential plateau. These burn lymph-induced changes in cardiac myocyte Ca2+ handling can contribute to burn-induced contractile dysfunction and ultimately to heart failure.

Effects of Palmitoyl Carnitine on Perfused Heart and Papillary Muscle.

BACKGROUND: Palmitoyl carnitine accumulation during ischemia causes profound electrophysiological changes, resulting in arrhythmias. We studied the electrophysiological and contractile effects of palmitoyl carnitine. METHODS AND RESULTS: Extracellular recordings made by using the endocardial unipolar paced evoked response (PER) in isolated perfused rabbit hearts were compared with action potentials (AP) recorded from septal artery perfused rabbit papillary muscle. Left ventricular pressure was monitored in isolated hearts. In perfused hearts palmitoyl carnitine (30 µmol/L, 30 minutes) significantly (P <.001) increased the latency of activation (St-R interval) by 58% +/- 8% and reduced repolarization time (R-E interval) by 39% +/- 4%. PER duration (St-E interval), was reduced by 30% +/- 3%. Palmitoyl carnitine (30 µmol/L) significantly (P <.001) decreased resting membrane potential (19 +/- 2 mV) of AP, reduced peak amplitude (33.5 +/- 8 mV) and rate of rise of phase 0 (41 +/- 8 V/s). Significant reductions (P <.001) in the action potential duration 50% (129.4 +/- 28 ms) and 90% (139.8 +/- 32 ms) were also observed. An initial positive inotropic effect, which declined as irreversible contracture developed, was also observed. Verapamil (1 µmol/L), nifedipine (1 µmol/L), and caffeine (10 mmol/L) failed to abolish the positive inotropy. CONCLUSIONS: We suggest that palmitoyl carnitine disrupts intracellular calcium homeostasis leading to disturbances in electrical and contractile activity. Its accumulation during myocardial ischemia could contribute to calcium overloading and initiate lethal arrhythmias.

Tumor Necrosis Factor-α Induces a Biphasic Effect on Myocardial Contractility in Conscious Dogs

Tumor necrosis factor-alpha (TNF-alpha) likely plays a role in the pathophysiology of myocardial depression observed in septic shock. To evaluate the hemodynamic effects of TNF-alpha in vivo while eliminating the influence of altered sympathetic tone, eight conscious chronically instrumented dogs were studied after pretreatment with propranolol (2 mg/kg) and atropine (2mg). Using three sets of piezoelectric crystals to measure left ventricular (LV) volume and LV manometers to measure pressure, we determined load-independent parameters of LV systolic performance before, during, and after infusion of recombinant human TNF-alpha (rhTNF-alpha, 40 micrograms/kg for 1 hour). Plasma was analyze for epinephrine and norepinephrine. Between 1 and 7 hours of exposure, rhTNF-alpha induced significant increases in circulating catecholamines. Norepinephrine rose from 268.6 +/- 47.2 to 426.2 +/- 87.0 pg/mL (P < .05) at 1 hour and peaked at 921.2 +/- 156.8 pg/mL (P < .001) at 4 hours after initiating rhTNF-alpha treatment. Similarly, epinephrine increased from 130.2 +/- 30.9 to 884.5 +/- 210.2 pg/mL (P < .05) at 1 hour and peaked at 3195.3 +/- 476 pg/mL (P < .001) at 4 hours. Before the surge of circulating catecholamines and despite complete beta adrenergic blockade, rhTNF-alpha induced a 7% to 40% increase in LV contractile performance during the 60-minute infusion. After this initial positive inotropic effect, rhTNF-alpha treatment led to precipitous systolic dysfunction between 2 and 7 hours of exposure; this myocardial depressant effect persisted at 25 hours. LV systolic performance declined to 19% to 35% of baseline values, depending on the specific contractile parameter evaluated. We conclude that rhTNF-alpha affects LV systolic function in a time-dependent biphasic manner. Increases in circulating catecholamines after rhTNF-alpha infusion cannot account for the early improvement in LV systolic performance.

Role of Na(+)-H+ exchange in mediating effects of endothelin-1 on normal and ischemic/reperfused hearts.

Endothelin (ET) has been shown to be elevated under conditions of cardiac pathology and to produce diverse cardiac effects, including coronary constriction and a positive inotropic influence. We characterized the concentration- and time-dependent effects of the most potent of the ET isoforms, ET-1 (0.4, 2, and 4 nmol/L), on myocardial contractility and coronary resistance and assessed its effects on the ischemic and reperfused heart. Because ET-1 has been shown to activate the Na(+)-H+ exchanger in cardiac myocytes, we determined the contribution of the antiport by examining the effects of ET-1 in the presence of the Na(+)-H+ exchange inhibitor methylisobutyl amiloride (MIA). At all three concentrations, ET-1 produced an initial positive inotropic effect that was reversed with continued perfusion, the degree of the reversal being dependent on ET-1 concentration. With 0.4 nmol/L, contractility returned to pre-ET-1 values, whereas after 75 minutes of perfusion with 4 nmol/L ET-1, contractility was depressed by 75%. At all concentrations, ET-1 produced a coronary-constricting effect, whereas an elevation in resting tension was observed only with 4 nmol/L ET-1. MIA significantly prevented the positive inotropic effect of ET-1 but had no effect on loss in function or elevation in resting tension produced by 4 nmol/L ET-1. Furthermore, MIA partially, but not significantly, attenuated the constricting effects of all ET-1 concentrations. In the ischemic heart, 0.4 nmol/L ET-1 appeared to delay the loss in contractility produced by cessation of flow, although the effect was not significant. Higher concentrations of ET-1 were without effect on ischemia-induced contractile depression, although their presence produced a marked elevation in resting tension during ischemia that was attenuated by MIA. Recovery in contractility was reduced by all concentrations of ET-1, although the effects of the lowest concentration were associated primarily with defective relaxation. The depressant effects of ET-1 either in normal or ischemic/reperfused hearts were irreversible. The inhibitory effects of ET-1 on contractile recovery were associated with diminished tissue glycogen and elevated lactate levels. High-energy phosphates after reperfusion were depressed in hearts treated with 4 nmol/L ET-1. The attenuation in contractile recovery and alterations in metabolite content were prevented by MIA. These results provide evidence that ET-1 produces complex effects on heart function that are likely mediated via different mechanisms and demonstrate its ability to aggravate ischemic and reperfusion injury through a mechanism possibly involving Na(+)-H+ exchange activation.

Effects of lysophosphatidylcholine on electrophysiological properties and excitation-contraction coupling in isolated guinea pig ventricular myocytes.

Lysophosphoglyceride accumulation in ischemic myocardium has been implicated as a cause of arrhythmias. We examined the effects of lysophosphatidylcholine (LPC) in isolated guinea pig ventricular myocytes. In paced myocytes loaded with the Ca2+ indicator Indo-1-AM and studied at room temperature, 20 microM LPC caused an initial positive inotropic effect followed by spontaneous automaticity, a decline in active cell shortening, and progressive diastolic shortening (contracture) leading to cell death. These changes were accompanied by a progressive increase in cytosolic [Ca2+]i. In patch-clamped myocytes dialyzed internally with high EGTA concentrations, LPC caused membrane depolarization, shortening of the action potential duration, and abnormal automaticity as seen in multicellular preparations. Voltage clamp experiments revealed the appearance of a nonselective leak conductance without significant changes in the delayed rectifier K+ current, inward rectifier K+ current, L-type Ca2+ current, and T-type Ca2+ current. Pretreatment with 20 mM caffeine and [Ca2+]o-free solution did not prevent the leak current. In patch clamped myocytes loaded with 0.1 mM Fura-2 salt, the [Ca2+]i transient induced by either voltage clamps or brief caffeine exposure remained normal until the nonselective leak current developed. The Na(+)-Ca2+ exchange current elicited during caffeine-induced [Ca2+]i transients also did not appear to be altered by LPC. Qualitatively similar results were obtained in myocytes studied at 35 degrees C. The membrane detergent saponin (0.005% wt/wt) mimicked all of the effects of LPC. We conclude that under these experimental conditions the effects of LPC are most compatible with a detergent action causing membrane leakiness with resultant depolarization, [Ca2+]i overload, and contracture.

Effects of 5-lipoxygenase inhibition on cardiac anaphylaxis in isolated guinea pig hearts.

Synthesis of the cardioactive peptidoleukotrienes depends on the activity of 5-lipoxygenase. It is unclear whether inhibition of endogenous leukotriene biosynthesis can alter vasoconstriction and negative inotropic effects associated with inflammatory states in the heart. In an attempt to study this problem, two 5-lipoxygenase inhibitors, the novel compound RG 6866, N-methyl-4-benzyloxyphenyl acetohydroxamic acid, and AA 861 have been examined in Langendorff-perfused guinea pig hearts undergoing cardiac anaphylaxis and compared to indomethacin and LY 171883, a cyclooxygenase inhibitor and a leukotriene antagonist, respectively. In paced hearts perfused at constant pressure, RG 6866 had no apparent direct effects, infused intracardially at 20 micrograms/min, about 3 mumol/l. LY 171883, but not RG 6866 or indomethacin, antagonized coronary vasoconstriction by exogenous leukotriene D4. When hearts were challenged with antigen (ovalbumin 1 mg i.c.) in the presence of antihistamines, a significant reduction in coronary flow occurred within 30 s which was maximal at 2-3 min, -48 +/- 3%, and persisted for at least 10 min. An initial positive intropic effect was followed by a 25% fall in developed pressure. Both 5-lipoxygenase inhibitors blocked effects of antigen challenge on coronary flow: RG 6866, -4 +/- 3%, AA 861, -11 +/- 3%; reduction at 2 min. The late negative inotropic effect was also blocked. The combined 5-lipoxygenase inhibitor/leukotriene antagonist, RG 5901, and the leukotriene antagonist, LY 171883, were also effective, but not indometacin. These results provide further evidence for the contribution of leukotrienes to cardiac anaphylaxis. More importantly, these findings suggest that 5-lipoxygenase inhibitors can be beneficial in situations where endogenous leukotrienes produce coronary vasoconstriction and myocardial depression.

Pharmacologic Basis of Responses to Midazolam in the Isolated, Cross-Perfused, Canine Right Atrium

The effects of midazolam on atrial rate and contractile force in the isolated canine atrium perfused with donor blood were investigated. When midazolam in a dose range of 100-1000 micrograms was injected directly into the sinus node artery of the isolated atrium, biphasic (negative followed by positive) chronotropic and triphasic (positive, negative followed by positive) inotropic effects were induced. After propranolol or imipramine, the positive chronotropic and the secondary positive inotropic effects were significantly suppressed, but the initial positive inotropic effect was not affected. Tetrodotoxin, atropine, or R05-4864, a selective ligand for peripheral benzodiazepine binding sites, did not modify midazolam-induced effects. When midazolam in a dose of 0.1-1 mg/kg was administered intravenously to the donor dog, monophasic negative chronotropic and inotropic effects in the isolated atrium were observed but were not as prominent. We conclude that midazolam has direct cardiac inhibitory properties including catecholamine release due to a tyramine-like action.

Suppression of positive inotropic and toxic effects of cardiac glycosides by amiloride

Effects of amiloride on the inotropic and toxic actions of cardiac glycosides were examined using left atrial muscle isolated from guinea pig heart. Preincubation of atrial muscle with amiloride significantly decreased the maximum positive inotropic effect of dihydrodigoxin but failed to reduce that of isoproterenol. Amiloride prevented the contracture and significantly reduced the incidence of arrhythmias induced by 2 μM digoxin. Similar experiments examining 5 μM digoxin-induced arrhythmias showed that amiloride increased both the time required to produce arrhythmias and the fractional occupancy of sarcolemmal Na,K-ATPase by digoxin at the onset of arrhythmias. The antagonism of cardiac glycoside actions was best observed during the decline in developed tension elicited by amiloride subsequent to its initial positive inotropic effect. Amiloride had no effect on binding site concentration for ATP-dependent [ 3H]ouabain binding but decreased affinity of the binding sites for ouabain in membrane preparations obtained from guinea pig heart. Furthermore, amiloride inhibited Na,K-ATPase activity and increased the IC 50 value for ouabain inhibition of the enzyme. These results indicate that amiloride antagonizes the positive inotropic and toxic effects of cardiac glycosides. Possible mechanisms for the antagonism include inhibition of sarcolemmal Na +/Ca 2+ or Na +/H + exchange.

Contractile failure and recovery in rat hearts perfused with Na+- or K+-free medium

Ca2+ or K+ concentrations during Na+-free perfusion and Ca2+ or Na+ concentrations during K+-free perfusion were varied and the effects on the contractile force were recorded during failure as well as recovery of the hearts. The initial positive inotropic effect of zero Na+ was directly related to the Ca2+ or K+ concentration of the medium. Time time of failure was not affected by changes in Ca2+ concentrations of the Na+-free medium whereas lowering of K+ concentration prolonged the time of failure. The initial negative inotropic effect of perfusion with zero K+ was increased by low Ca2+ and was changed to a positive effect by high Ca2+; the time of failure was inversely related to the Ca2+ concentration. Lowering of Na+ in zero K+ medium caused fibrillation of the heart. The recovery upon reperfusion with control medium of Na+-deprived hearts was improved by a reduction in Ca2+ concentration during Na+-free perfusion and was adversely affected by increased Ca2+. Increases or decreases in K+ during Na+-free perfusion reduced recovery. The length of the fibrillation period was affected in a characteristic manner by changes in Ca2+ or K+ concentrations. The recovery of the K+-deprived hearts was also influenced by changes in Ca2+ or Na+ concentrations during K+-free perfusion. In addition to altering the electrical properties of the myocardium, the early effect of Na+ or K+ lack appears to be on the Ca2+ movements across the sarcolemma and this is followed by the intracellular Ca2+ overload. The study emphasizes the importance of a proper balance between the Na+, K+, and Ca2+ concentrations for normal functioning of the heart.

The Nature of the Initial Positive Inotropic Effect of K Depletion in Bullfrog Atrial Muscle

In frog atrium, K depletion exerts initial, and late positive inotropic effects. The initial effect on the membrane potential, current and tension components was studied using a double gap method under voltage clamped and unclamped conditions. In both cases, the presence of the initial effect was demonstrated. The effect was clearer in preparations in which a positive inotropic effect to toxic doses of ouabain (10(-6) M) had already been attained. This suggests that the initial phase is independent from Na-pump inhibition. Voltage clamp study revealed that the slow inward calcium current (ICa) was enhanced transiently after K depletion concomitant with an augmentation of ICa-dependent phasic tension. ICa-independent tonic tension was not altered at this initial phase. For the enhancement of ICa, two mechanisms appeared to be involved, one an enhancement of gCa and the other a faster and/or longer opening of the slow channel. These results indicated that an enhancement of ICa together with the well-known prolongation of action potential were the cause of the initial positive inotropic effect of K depletion.