Positive Inotropic Effect Of Milrinone Research Articles

The Adenylate Cyclase Activator Forskolin Potentiates the Positive Inotropic Effect of the Phosphodiesterase Inhibitor Milrinone But Not of the Calcium Sensitizer Levosimendan nor of Its Hemodynamically Active Metabolites: An Apparent Conundrum.

OR-1855 and OR-1896 are 2 hemodynamically active metabolites of the inodilator levosimendan, with calcium sensitizing activity, but their mechanism of action is still not fully understood. It has been previously reported that the positive inotropic effect of levosimendan is not potentiated by the adenylate cyclase activator forskolin, whereas forskolin does potentiate the effects of the phosphodiesterase (PDE) inhibitor milrinone. To ascertain whether the active metabolites follow the same pattern of levosimendan, the positive inotropic effects of OR- 1855 and OR-1896 were studied in guinea-pig-isolated papillary muscle in the presence and absence of forskolin. OR-1855 and OR-1896 were also tested as inhibitors of PDE-III and PDE-IV. Our results show that 0.1 µM forskolin did not potentiate the positive inotropic effect of OR-1855 or OR-1896, as in the case of the parent compound levosimendan. As in previous studies, the positive inotropic effect of milrinone was markedly potentiated in the presence of forskolin. From these data, we propose an explanation for the divergent behavior of the calcium sensitizing drugs and PDE inhibitors.

Early administration of milrinone ameliorates lung and kidney injury during sepsis in juvenile rats.

A sepsis model was created, induced by cecal ligation and puncture (CLP), in juvenile rat groups. Milrinone (MIL), which is known to have a modulatory effect on pro-inflammatory cytokines, was administered to the designated rat groups in the early period before severe sepsis developed. The study was aimed at investigating the possible protective effects of milrinone on the lung and kidney tissues of rats in the late phase of sepsis. The rat pups were divided into seven groups with six animals in each group: (1) healthy rats who received no drug; (2) CLP-S12 (sacrificed at hour 12); (3) CLP-S24 (sacrificed at hour 24); (4) CLP-MIL1-S12 (administered with 0.5 mg/kg milrinone at hour 1 and sacrificed at hour 12); (5) CLP-MIL1-S24 (administered with 0.5 mg/kg milrinone at hour 1 and sacrificed at hour 24): (6) CLP-MIL12-S24 (administered with 0.5 mg/kg milrinone at hour 12 and sacrificed at hour 24), (7) and CLP-MIL1,12-S24 (administered with 0.5 mg/kg milrinone at hours 1 and 12 and sacrificed at hour 24). Significant differences were found between the early and late administration of milrinone in terms of both molecular and histopathological results. The results showed that the tissues were significantly preserved in the groups in which milrinone had been started in the early period compared to the sepsis control groups and the groups in which milrinone had been started in the late period. In addition to the positive inotropic effects of milrinone, its immunomodulatory properties that result in decreased cytokine storm can be beneficial during early period of sepsis.

Effects of milrinone on left ventricular cardiac function during cooling in an intact animal model

IntroductionDue to adverse effects of β-receptor agonists reported when applied during hypothermia, left ventricular (LV) cardiac effects of milrinone, a PDE3 inhibitor which mode of action is deprived the sarcolemmal β-receptor–G protein–PKA system, was tested during cooling to 15°C. MethodsSprague Dawley rats were instrumented to measure left ventricular (LV) pressure–volume changes using a Millar pressure–volume conductance catheter. Core temperature was reduced from 37 to 15°C (60min) using internal and external heat exchangers. Milrinone, or saline placebo, was given as continuous i.v. infusions for 30min at 37°C and during cooling. ResultsIn normothermic controls continuous milrinone infusion for 90min elevated cardiac output (CO) and stroke volume (SV) significantly. Significant differences in cardiac functional variables between the milrinone group and the saline control group during cooling to 15°C were found: Compared to saline treated animals throughout cooling from 33 to 15°CSV was significantly elevated in milrinone animals, the index of LV isovolumic relaxation, Tau, was significantly better preserved, and both HR and CO were significantly higher from 33 to 24°C. Likewise, during cooling between 33 and 28°C also LVdP/dtmax was significantly higher in the milrinone group. ConclusionsMilrinone preserved LV systolic and diastolic function at a significantly higher level than in saline controls during cooling to 15°C. In essential contrast to our previous results when using β-receptor agonists during hypothermia, the present experiment demonstrates the positive inotropic effects of milrinone on LV cardiac function during cooling to 15°C.

Milrinone inhibits contractility in skinned skeletal muscle fibers.

Direct action of the cardiotonic bipyridine milrinone on the cross bridges of single fibers of skinned rabbit skeletal muscle was investigated. At 10 degrees C and pH 7.0, milrinone reduced isometric tension in a logarithmically concentration-dependent manner, with a 55% reduction in force at 0.6 mM. Milrinone also reduced Ca2+ sensitivity of skinned fibers in terms of force production; the shift in the force-pCa curve indicated a change in the pCa value at 50% maximal force from 6.10 to 5.94. The unloaded velocity of shortening was reduced by 18% in the presence of 0.6 mM milrinone. Parts of the transient tension response to step change in length were altered by milrinone, so that the test and control transients could not be superimposed. The results indicate that milrinone interferes with the cross-bridge cycle and possibly detains cross bridges in low-force states. The results also suggest that the positive inotropic effect of milrinone on cardiac muscle is probably not due to the drug's direct action on the muscle cross bridges. The specific and reversible action of the bipyridine on muscle cross bridges makes it a potentially useful tool for probing the chemomechanical cross-bridge cycle.

The role of cAMP- and cGMP-dependent protein kinases in the cardiac actions of the new calcium sensitizer, levosimendan.

The role of phosphodiesterase III inhibition and calcium sensitization in the cardiac actions of levosimendan, (R)-[[4-(1,4,5, 6-tetrahydro-4-methyl-6-oxo-3-pyridazinyl)phenyl]hydrazono]propane dinitrile, was studied. Various heart preparations were used to investigate positive inotropy, chromotropy, coronary flow and calcium sensitivity of contractile proteins. The cAMP- and cGMP-dependent protein kinases (PKA and PKG) were inhibited by KT5720 and KT5823, respectively. Furthermore, the synthesis of cAMP was stimulated by forskolin and increased phosphorylation of troponin I was induced by isoprenaline. In Langendorff guinea-pig heart, levosimendan (0.01-1 microM) and milrinone (0.1-10 microM) increased the left ventricular systolic peak pressure almost to the same extent. In the presence of KT5720 (1 microM) milrinone was devoid of positive inotropic activity. In contrast, KT5720 did not antagonize the inotropic effect of levosimendan at < or = 0.03 microM (-up to the EC50 of levosimendan). The effects of levosimendan and milrinone on heart rate and coronary flow were not affected by KT5720. The PKG inhibitor, KT5823 (1 microM), on the other hand, potentiated the levosimendan-induced increase in coronary flow while it had no effect on the increase induced by milrinone. The mechanical parameters were not affected by KT5823. In the papillary muscle, the positive inotropic effect of milrinone but not that of levosimendan was potentiated by forskolin (0.1 microM). In contrast to milrinone, the positive inotropy by levosimendan was decreased by isoprenaline pretreatment (0.1 microM; 3 min). In line with this, the calcium-sensitizing effect of levosimendan was decreased in skinned fibers prepared from isoprenaline-treated hearts. Our results indicate that the cardiac effects of levosimendan at its therapeutically relevant concentrations were not mediated through PKA or PKG and its positive inotropy is therefore most probably due to the previously reported troponin-C-mediated calcium sensitization of contractile proteins.

Mechanical and electrophysiological effects of milrinone on the force‐frequency relationship in mammalian myocardium

Isometric force, action potential and current-voltage relation were studied in guinea-pig and ferret papillary muscles. Milrinone (1 microM) increased peak twitch force by 40 +/- 4%, reduced time to peak tension (TPT) by 12.1 +/- 3% (n = 6, P < 0.01) and reduced time to half relaxation by 17.3 +/- 4.1% (n = 6, P < 0.01). The effect of milrinone was potentiated by rolipram, a RI-PDE inhibitor which in itself had no inotropic effect. After the addition of rolipram peak isometric force was increased by 104 +/- 8% (n = 6, P < 0.001), TPT was further reduced whereas time to half relaxation was slightly increased after the addition of rolipram. Action potential duration at 75% repolarization was decreased by 11 +/- 5 ms (n = 6, P < 0.05). Milrinone also potentiated the second inward current (Isi) by 21 +/- 3.2% (n = 6, P < 0.01). Peak twitch force in response to a test stimulus after an interval, i.e. mechanical restitution was increased at all intervals. The onset of restitution was faster and time to full restitution also shortened. Maximum postextrasystolic potentiation was greater in the presence of milrinone, whereas relative potentiation was smaller in presence of milrinone (46 +/- 7%) than in control (74 +/- 7%). The recirculation fraction of activator calcium was enhanced by milrinone from 0.35 +/- 0.04 to 0.48 +/- 0.07. The results support the view that the positive inotropic effect of milrinone is due to a greater inflow of calcium during the action potential and a more efficient intracellular calcium handling.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of positive inotropic effects of milrinone, dobutamine and ouabain.

This study compared the positive inotropic actions of milrinone in isolated rabbit myocardium with that of the conventional positive inotropic drug, dobutamine, and a cardiac glycoside, ouabain. Maximal increase in developed tension (g/mm2) was significantly (p < 0.05) greater with ouabain (from 2.0 to 4.4; 132 +/- 11%) than with dobutamine (from 3.9 to 6.0; 77.8 +/- 22.4%) or milrinone (from 2.4 to 3.6; 54.0 +/- 12.0%). Maximal augmentation of the rate of tension development (g/s/mm2), however, was similar with ouabain (from 14.3 to 39.7; 187 +/- 20%) and dobutamine (from 25.9 to 64.4; 174 +/- 35%), and both were significantly (p < 0.05) greater than with milrinone (from 18.2 to 30; 73.1 +/- 14.7%). In combination with dobutamine, however, the dose-response curve of milrinone was shifted to the left, and its ED10 was significantly (p < 0.001) reduced by 100-fold to 1.6 x 10(-7) M. Thus, milrinone is significantly less potent than dobutamine or ouabain in vitro and is without contractile effect at clinically relevant concentrations. However, data from the combined application of a catecholamine and milrinone in isolated myocardium suggest that milrinone may induce a direct positive inotropic effect at clinical concentrations in the presence of augmented sympathetic neurohumoral stimulation.

Implication of Cyclic AMP in the Positive Inotropic Effects of Cyclic GMP-Inhibited Cyclic AMP Phosphodiesterase Inhibitors on Guinea Pig Isolated Left Atria

The present investigation was performed to characterize the positive inotropic actions of inhibitors of the cyclic GMP-inhibited cyclic AMP phosphodiesterase (CGI-PDE) on the electrically driven guinea pig left atria. With forskolin added at a concentration (1 x 10(-8)-3 x 10(-8) M) that in itself produced a 10.7% increase of the response to electrical stimulation, the EC50 value of isoproterenol was slightly but significantly (p less than 0.01) reduced from 1.5 to 0.9 nM without modification of the maximal developed tension (delta Emax). The positive inotropic effects of milrinone, piroximone, and SK&F 94120 were significantly enhanced by forskolin (percentage of increase at 3 x 10(-5) M CGI-PDE inhibitors concentration was milrinone 43, piroximone 154, and SK&F 94120 133). With 8-Br-cyclic GMP added (10(-4) M) that in itself exerted a small but significant negative inotropic effect (-0.12 g), the EC50 value of isoproterenol was significantly (p less than 0.01) increased from 1.5 to 3 nM without modification of the delta Emax value. The positive inotropic effects of CGI-PDE inhibitors were significantly depressed by 8-Br-cyclic GMP (percentage of decrease at 3 x 10(-5) M was milrinone 35, piroximone 63, and SK&F 94120 39). In identical conditions, neither forskolin nor 8-Br-cyclic GMP produced any significant alteration of the positive inotropic effects of elevated external Ca2+ or protoveratrine B. Together these results strongly suggest that the positive inotropic effect of CGI-PDE inhibitors is mediated by cyclic AMP in guinea pig left atria.(ABSTRACT TRUNCATED AT 250 WORDS)

Milrinone enhances cytosolic calcium transient and contraction in rat cardiac myocytes during beta-adrenergic stimulation

We have investigated the mechanism that underlies the absence of a positive inotropic effect of milrinone on rat myocardium. The twitch characteristics of enzymatically dissociated left ventricular myocytes from the adult rat and guinea pig were assessed by edge tracking during field stimulation. In some rat myocytes loaded with the ester derivative of the Ca 2+ probe Indo-1 we simultaneously measured changes in cell length and in the associated cytosolic Ca 2+ (Ca 1) transient. Our results show that in guinea pig myocytes bathed in 0.5 mM [Ca 2+] and field stimulated at 1 Hz, milrinone (10 μM) had a positive inotropic effect. In contrast milrinone had no effect on the contractile properties of rat myocytes studied under similar conditions and field stimulated at 0.2 Hz. In rat myocytes bathed in 0.5 mM [Ca 2+] and stimulated at 0.2 Hz isoproterenol (1 nM) increased the amplitude and shortened the duration of the contraction and of the associated Ca i transient; these effects of beta-adrenergic stimulation were further enhanced by the addition of milrinone (10 μM) in the presence of isoproterenol. Under conditions of higher cell Ca 2+ loading achieved by raising bathing [Ca 2+] to 1 mM and isoproterenol to 3 nM the positive inotropic effect of milrinone (10 μM) in rat myocytes saturated when spontaneous oscillatory Ca 2+ release appeared in the diastolic intervals between electrically stimulated twitches. Our results suggest that an enhancement in the baseline beta-adrenergic stimulation is required for milrinone to exercise a positive inotropic action on rat myocardial tissue.

Subsensitivity of the Failing Human Heart to Isoprenaline and Milrinoneo is Related to β-Adrenoceptor Downregulation

The number of cardiac beta-adrenoceptors and the positive inotropic effect of isoprenaline and milrinone were measured in cardiac membranes and isolated, electrically driven muscle strips from nonfailing donor hearts and from patients with mitral valve disease (NYHA II-III), ischemic heart disease, and dilated cardiomyopathy (NYHA IV). In nonfailing hearts, the number of beta-adrenoceptors were 41.5 fmol/mg protein (mean, n = 3). In ischemic heart disease and NYHA II-III, there was a loss of cardiac beta-adrenoceptors (22.1 fmol/mg protein, mean, n = 3; 23.2 +/- 2.7 fmol/mg protein, n = 30), respectively. In NYHA IV, there was a pronounced reduction of the number of cardiac beta-adrenoceptors to 12.1 +/- 1.5 fmol/mg protein (n = 15). The Kd value did not differ in either group. Correspondingly, the positive inotropic effect of isoprenaline was more pronounced in nonfailing myocardium, reduced in NYHA II-III and ischemic heart disease and almost blunted in NYHA IV. Similar results were observed with the phosphodiesterase inhibitor milrinone. A good correlation of the beta-adrenoceptor density to the maximal positive inotropic effect of isoprenaline and milrinone was observed. Neither the number of cardiac beta-adrenoceptors nor the positive inotropic effect of isoprenaline correlated with the age of the patients. We conclude that the number of cardiac beta-adrenoceptors and the positive inotropic effect of beta-adrenoceptor agonists are reduced in the failing human heart depending on the severity of heart failure. Furthermore, the positive inotropic effect of milrinone is also reduced and related to the reduction of beta-adrenoceptors. The lack of correlation with the age of the patients provides evidence for a predominant role of heart disease rather than aging in the reduction of beta-adrenoceptors and subsensitivity to cyclic AMP-increasing positive inotropic agents in the failing human heart.

Enhancement of the effectiveness of milrinone to increase force of contraction by stimulation of cardiac beta-adrenoceptors in the failing human heart.

The positive inotropic effect of milrinone was investigated in isolated, electrically driven (1 Hz) human papillary muscle strips from nonfailing myocardium (control) and from patients with moderate (NYHA II-III) and severe (NYHA IV) heart failure. In the control hearts, milrinone increased force of contraction to about the same degree as Ca2+ at 15 mmol/l. In NYHA II-III, the positive inotropic effect was significantly reduced compared with the control hearts. In NYHA IV, the effectiveness (maximal increase in force of contraction) and potency (as judged from the EC50 values) were significantly less than in NYHA II-III and controls. Preexposure of the preparations to isoprenaline in NYHA II-III and NYHA IV shifted the concentration-response curve for milrinone to the left and, moreover, restored the effectiveness of the compound (i.e., similar positive inotropic effect as Ca2+). No difference in the cAMP-phosphodiesterase inhibition by milrinone could be detected between controls, NYHA II-III, and NYHA IV. It is concluded that diminished basal cAMP production is responsible for reducing the effectiveness of milrinone in the failing human heart. A diminished inhibition of cAMP-phosphodiesterase does not play a role. The fact that stimulation of cardiac beta-adrenoceptors increased the effectiveness of milrinone provides evidence that combined application of catecholamines and phosphodiesterase inhibitors may be useful in the treatment of patients with terminal heart failure.

The Effect of Extracellular Ca2+ and Related Ions on the Cardiac Action of Milrinone

The biphasic single dose, dose-response curve of milrinone was sensitive to [Ca2+]0. At concentrations of 1.8 nM Ca2+ or less this biphasic response is observed but at [Ca2+]0 of 4.5 mM or more the dose response curve becomes monotonic. The inotropic response to milrinone in contrast to norepinephrine is highly sensitive to the extracellular [Ca2+]0. At low [Ca2+]0 of 0.15 mM milrinone could produce a negative inotropic effect. The positive inotropic effect of milrinone was proportional to [Ca2+]0 up to 2.7 mM. With [Ca2+]0 above 3.6 mM and low [Na+]0, the inotropic response to milrinone was reduced. These effects were due to increased [Ca2+]i and not due to the increase in contractile force produced by Ca2+. The positive inotropic effect of milrinone in contrast to norepinephrine is increased with an increase in [K+]0 possibly due to the depolarization produced by K+. The positive inotropic response to 10 micrograms of milrinone when [Ka+]0 = 4 mM was not significantly changed by Ca2+ channel blocking agents. In depolarized tissue (20 mM K+) the electropharmacological and contractile effects of milrinone are blocked by verapamil and ruthenium red. This suggests that under these conditions different mechanisms of Ca2+ channel activation are operative. Substitution of Sr2+ for Ca2+ increased contractile force and prolonged time to peak tension and relaxation time. Milrinone decreased time to peak tension but had no detectable effect on relaxation time. The results are discussed and it is suggested that milrinone acts on Ca2+ channels in the sarcolemma and intracellularly by increasing cyclic AMP which activates Ca2+ release and uptake from the sarcoplasmic reticulum.

Mechanism of the positive inotropic effect of milrinone in cultured embryonic chick ventricular cells

Milrinone, a potent positive inotropic and vasodilating agent, has shown promise in the clinical treatment of congestive heart failure, but significant controversy about its mechanism of action exists. To approach these mechanistic problems in a non-innervated, non-diffusion-limited system, the effects of milrinone on cultured embryonic chick ventricular cells were examined. At 37 degrees C in physiologic buffer, milrinone produced a rapid, concentration-dependent increase in amplitude of contraction that was 45% of the maximum increment in contraction produced by elevated extracellular calcium; the EC50 was 8 microM. This peak response was quantitatively similar to the contractile response produced by isobutyl methylxanthine, a potent phosphodiesterase inhibitor. Milrinone inhibited 70% of total phosphodiesterase activity of cultured ventricular cells with an EC50 of 11 microM. Exposure to 1 X 10(-4) M milrinone resulted in rapid increase in cyclic AMP content to levels greater than 100% above control within 4 min. The same concentration also produced a 43% increase in the rate of transsarcolemmal 45Ca uptake. The stimulation of 45Ca uptake rate was similar to the response produced by 1 microM isoproterenol and could be completely abolished by 10 microM verapamil. Thus, in cultured embryonic chick myocardial cells, the positive inotropic effect of milrinone is largely, if not entirely, attributable to phosphodiesterase inhibition, leading to intracellular cyclic AMP accumulation and stimulation of transsarcolemmal calcium influx via the slow calcium channel.

Additive positive inotropic effects of milrinone, ouabain and calcium in diseased human ventricular myocardium.

The interactions of milrinone, ouabain and calcium on force of contraction in isolated, contracting human papillary muscle strips were measured. Milrinone (EC50, 8 X 10(-5)M) increased force of contraction maximally by 2.8 +/- 0.8 mN at 5 X 10(-4)M; significantly less than either ouabain (1 X 10(-7)M; 4.8 +/- 0.5 mN increase) or calcium (15 mM; 6.2 +/- 0.6 mN increase). A submaximal, but not a maximal, inotropic effect of ouabain could be increased by the addition of milrinone; in contrast, both ouabain and calcium increased the maximal inotropic effect of milrinone by 1.7 +/- 0.2 mN and 2.7 +/- 0.3 mN, respectively. The combined inotropic effect of milrinone with either ouabain of 4.2 +/- 0.3 mN or calcium of 5.6 +/- 0.4 mN was not different from that with calcium or ouabain alone. We conclude that further positive inotropic effects should be expected when digitalis is given to patients with congestive heart failure who are already optimally treated with milrinone.