The cardenolides strophanthidin, digoxigenin and dihydroouabain act as activators of the human RORγ/RORγT receptors.

Two isoforms of a ligand-activated nuclear receptor, RORγ and RORγT, have been implicated in various physiological functions, including energy metabolism, circadian rhythm and immune system development. Using a stably transfected reporter cell line, we screened two chemical libraries and identified three cardenolides (natural, plant-derived pesticides) as activators of RORγ-dependent transcription. These compounds increased G6PC and NPAS2 expression in HepG2 cells, accompanied by increased occupancy of RORγ within the promoters of these genes. Further, strophanthidin, digoxigenin and dihydroouabain upregulated IL17A and IL17F expression and enhanced IL17 secretion in Th17 human lymphocytes. Molecular docking analyses of these compounds to the RORγ LBD showed favorable docking scores, suggesting that cardenolides may act as agonists of the receptor. Thus, our results provide new chemical structures for further development of RORγ-selective modulators with virtual therapeutic potential.

Mechanistic insight into the functional and toxic effects of Strophanthidin in the failing human myocardium.

BACKGROUND: Cardiac glycosides are characterized by a narrow therapeutic range with Ca2+-overload and arrhythmias occurring at higher concentrations. Data on cardiac glycosides in isolated failing human myocardium are scarce and the frequency-dependent actions and toxicity of Strophanthidin have not yet been characterized. AIMS: To determine inotropic responses and toxicity of Strophanthidin in failing human myocardium. METHODS AND RESULTS: Experiments were performed in trabeculae from 64 end-stage failing hearts. Developed force, and intracellular [Ca2+]i and [Na+]i were recorded with Strophanthidin (0.01 to 1 micromol/L; 37 degrees C, 1 Hz) and compared to interventions with distinct mechanisms of action (elevated [Ca2+]o, Isoproterenol, and EMD57033). The effects of Strophanthidin on force-frequency behaviour were also assessed. Strophanthidin exerted concentration-dependent positive inotropic effects. These were paralleled by increases in intracellular [Na+] as well as increasing [Ca2+]i-transients and SR-Ca2+-load. At high concentrations (>0.5 micromol/L), Strophanthidin caused afterglimmers and aftercontractions, with declining developed force despite further increasing [Ca2+]i-transients. The force-frequency-relationship and diastolic function at higher pacing rates was worsened by Strophanthidin in a concentration-dependent manner. CONCLUSIONS: Strophanthidin toxicity was dependent on concentration, calcium load, beating rate and beta-adrenergic receptor activation. Our data support the view that low doses, heart rate control and additional beta-adrenergic receptor blockade are essential in the use of cardiac glycosides in heart failure.

Determination of strophanthidin in ingesta and plant material by LC-MS/MS.

An LC-MS/MS method was developed for the semiquantitative determination of strophanthidin glycosides in ingesta from animals. Strophanthidin glycosides were simultaneously extracted and hydrolyzed to the strophanthidin aglycone using aqueous methanolic hydrochloric acid and the extracts cleaned up using solid-phase extraction. Extracts were analyzed using reverse-phase HPLC coupled with positive ion electrospray mass spectrometry. Characteristic product ion spectra were produced by fragmentation of the [M + H](+) precursor ion for each analyte. Quantitation was performed using the internal standard method with digitoxigenin serving as the internal standard. The method detection limit was calculated to be 0.075 microg/g, and the limit of quantitation was calculated to be 0.24 microg/g for strophanthidin in control rumen samples. This method was used in diagnostic investigations to confirm fatal strophanthidin glycoside poisonings in horses.

Induction of delayed afterdepolarizations and triggered arrhythmias in isolated Purkinje fibers: comparison of resibufogenin and acetylstrophanthidin.

The purpose of this study was to compare the electrotoxicological effects of resibufogenin (RBG) (n = 14) with acetylstrophanthidin (AS) (n = 14) to induce delayed afterdepolarization (DAD) and triggered activity (TA), and their alteration of the electrophysiological properties in sheep cardiac Purkinje fibers using the extracellular electrograms, signal averaging, and standard microelectrode techniques simultaneously. The results indicated: 1) Lower toxic dose of RBG (0.52 mumol.L-1) and AS (0.25 mumol.L-1) induced intracellular and extracellular DAD (DAD-I and DAD-E) at pacing cycle length of 990 and 690 ms. 2) Higher toxic dose of RBG (2.6 mumol.L-1) and AS (5.0 mumol.L-1) induced DAD and TA, nonsustained or sustained premature action potential and oscillatory potentials; 3) At the beginning period of superfusing the drugs, both RBG and AS caused changes of the electrophysiological characteristics. This study demonstrates that the electro-toxicological characteristics and electrophysiological properties of RBG are similar to that of AS and suggests that RBG belongs to the family of digitalis-like drugs.

Role of oscillatory potential and pacemaker shifts in digitalis intoxication of the sinoatrial node.

BACKGROUND: Digitalis intoxication causes tachycardia, pacemaker shifts, and conduction disturbances in the sinoatrial (SA) node, but the mechanisms underlying these changes have not been clarified. We studied the role played by oscillatory potentials, electrical inhomogeneity, and calcium overload in cardiac steroid intoxication of the SA node. METHODS AND RESULTS: Guinea pig SA nodes (isolated from atrial tissue) were perfused in vitro. Transmembrane potentials and force were recorded. Strophanthidin (1 mumol/L) induced minor changes, although it was perfused for more than 30 minutes. In contrast, ouabain (0.5 mumol/L) and digoxin (1 mumol/L) intoxicated the SA node in 10-20 minutes. Ouabain and digoxin increased spontaneous rate and slope of diastolic depolarization, shifted the plateau to more negative values, and decreased the maximum diastolic potential. These cardiac steroids increased and then decreased contractile force and eventually caused the action potential and twitch to become irregular in amplitude and rhythm. In the presence of acetylcholine (ACh, 0.01-1 mumol/L), cardiac steroids decreased the resting potential, caused spontaneous activity, and increased force and, eventually, oscillatory potentials (Vos) and aftercontractions as well as overdrive excitation. To make the SA node electrically homogeneous (only slow responses), the SA node was perfused with high extracellular potassium concentration (with and without norepinephrine), tetrodotoxin (2.61 mumol/L), or lidocaine (50 mumol/L). Adding ouabain or digoxin to these solutions increased the rate but far less than in Tyrode's solution. Recovery in Tyrode's solution initially caused fast and irregular rhythms, which then subsided. Low extracellular calcium concentration ([Ca]o) (0.54 mmol/L) decreased force; adding ouabain markedly increased force and induced Vos. High [Ca]o (8.1 mmol/L) increased force; adding ouabain decreased force and made action potentials as well as contractions quite irregular. CONCLUSIONS: Ouabain and digoxin quickly intoxicate the SA node by inducing calcium overload and its manifestations (Vos, decrease in contractile force and aftercontractions), whereas strophanthidin does not, possibly because of the lack of a sugar moiety. The intoxication is less pronounced when sodium influx is decreased (slow responses), and this accounts for the shifts from dominant to subsidiary pacemakers. Marked conduction disturbances result from calcium overload, leading to the fractionation of SA node activity.

Effects of different cardiac steroids on intracellular sodium, inotropy and toxicity in sheep Purkinje fibers.

The purpose of the present study was to examine the differences between cardiac steroids that might underlie the variations in toxic/therapeutic ratios that have been reported to occur in vitro as well as in vivo. We used Na(+)-sensitive microelectrodes to measure changes in intracellular Na+ activity (aiNa) associated with positive inotropic and toxic effects of acetylstrophanthidin (AS) and a semisynthetic agent, actodigin. Measurements of aiNa, twitch tension and transmembrane potential were made in sheep Purkinje fibers stimulated at 0.03, 1 and 2 Hz. Ca(+)+i overload toxicity was indicated by the presence of transient depolarizations (TD). The following results were obtained: 1) at a stimulation frequency of 1 Hz, aiNa was significantly higher at peak tension with AS (13.6 +/- 1.1 mM) than with actodigin (11.0 +/- 0.4 mM, P less than .01), yet TD occurred at the same aiNa (10.9 +/- 0.7 vs. 11.9 +/- 0.7 mM, respectively, N.S.); 2) at frequencies of 1 to 2 Hz, aiNa was lower when TD occurred (10.4 +/- 0.9 mM at 2 Hz) than at peak tension (12.1 +/- 0.8 mM, P less than .05) during exposure to AS, whereas aiNa was the same at peak tension (10.6 +/- 1.1 mM) and when TD occurred (10.5 +/- 1.1 mM, N.S.) during exposure to actodigin; 3) the degree of positive inotropy at a high stimulation frequency (2 Hz) was significantly greater with actodigin (about 12-fold increase in force compared to control) than with AS (about 6-fold increase in force).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of first and second generation sulphonylureas on cardiotoxicity of strophanthidin in rabbits.

The effects of the first generation sulphonylurea compound gliclazide and the second generation sulphonylurea compound glipizide on strophanthidin toxicity was investigated in rabbits. The sulphonylurea pretreated animals were intravenously infused with 23 mumol/kg strophanthidin until the appearance of the first ventricular ectopic beat and continued thereafter until the appearance of ventricular fibrillation. The first generation sulphonylurea gliclazide increased, while the second generation sulphonylurea glipizide decreased the strophanthidin toxicity in a dose dependent manner. It was concluded that instead of first generation sulphonylureas, second generation sulphonylureas must be preferred in cardiac glycoside treated diabetics, when sulphonylurea treatment is necessary.

Calcium oscillations in digitalis-induced ventricular fibrillation: pathogenetic role and metabolic consequences in isolated ferret hearts.

The pathophysiology of the ventricular fibrillation that complicates digitalis intoxication was investigated. In this and other calcium-overload states, oscillations of the intracellular free calcium concentration ([Ca2+]i) have been implicated as the cause of ventricular tachyarrhythmias. We addressed two questions: 1) Are [Ca2+]i oscillations obligatory in the pathogenesis of ventricular fibrillation during digitalis toxicity? 2) What are the metabolic consequences of [Ca2+]i oscillations? Ferret hearts (n = 20) were Langendorff-perfused at constant flow with oxygenated HEPES-buffered Tyrode's solution at 37 degrees C. Isovolumic left ventricular pressure was measured along with the extracellular electrogram or with simultaneous phosphorus nuclear magnetic resonance spectra. When strophanthidin (20 microM) was added during pacing at 3 Hz, the positive inotropic effect soon gave way to a decrease in developed force. The decrease in force was accompanied by an increase in inorganic phosphate concentration, a decrease in phosphocreatine concentration, and a slight acidosis. The rhythm changed to ventricular fibrillation after 12-25 minutes. This change was initially accompanied by further metabolic deterioration, but all metabolites reached steady state within 12-18 minutes of the onset of ventricular fibrillation. Fast Fourier transformation revealed the existence of periodic oscillations at 7-10 Hz in both the extracellular electrogram and the ventricular pressure during ventricular fibrillation. Ryanodine, an inhibitor of [Ca2+]i oscillations, abolished the pressure oscillations but not the voltage oscillations. Exposure to ryanodine significantly decreased the inorganic phosphate concentration and increased the phosphocreatine concentration (p less than 0.05) despite continuing exposure to strophanthidin. The results indicate that oscillations of [Ca2+]i are not required to sustain ventricular fibrillation, but when present, such oscillations contribute importantly to metabolic deterioration.

The effect of various hypoglycaemic sulphonylureas on the cardiotoxicity of glycosides and arrhythmogenic activity due to myocardial ischaemia.

The effects of different sulphonylureas on the electrical cardiac activity were studied in 145 rabbits and in 103 rats as well as in 278 digitalis-treated, non-smoker non-insulin-dependent diabetics on the same therapy at least during the previous three months. In rabbits and rats glibenclamide (0.0032-100 mumol. kg-1) decreased, while tolbutamide and carbutamide (0.008-1000 mumol. kg-1) increased strophantidin toxicity and myocardial ischaemia induced transitory ventricular fibrillation dose-dependently. The differences between the dose-response curve of glibenclamide and those of tolbutamide or carbutamide were significant. In digitalized non-insulin-dependent diabetics, multifocal ectopic ventricular beats could be observed in none among the 80 glibenclamide-treated diabetics, while in 12 cases of the 71 tolbutamide and in 10 cases of the 61 carbutamide treated diabetics. Two of the 66 non-insulin-dependent diabetics receiving only diet and 7 of the 278 age and sex matched, non-smoker, metabolically healthy patients had multifocal ectopic ventricular beats. No significant difference could be found between the therapeutical groups. It was concluded that instead of tolbutamide, glibenclamide must be preferred in digitalis-treated diabetics, when metabolic control is not satisfactorily achieved by diet and regime alone.

The effect of intracoronary lidocaine infusion on acetylstrophanthidin-induced ventricular arrhythmia in dogs.

Antiarrhythmic drugs frequently cause extracardiac side effects that limit their use. If intracoronary delivery of a lower dose of drug to an electrically unstable focus can control arrhythmias, systemic adverse effects of these agents might be avoided. We investigated whether intracoronary lidocaine can suppress ventricular arrhythmia induced by acetylstrophanthidin, a rapidly acting digitalislike agent. We isolated and then cannulated either the left anterior descending or the left circumflex coronary artery in 12 pentobarbital-anesthetized dogs. Sustained ventricular tachycardia that persisted for 11.4 +/- 8.6 minutes was reliably induced by the intracoronary infusion. In all of 24 trials, an intracoronary lidocaine bolus at 2% of the usual systemic dose (0.77 mg/30 sec) abolished digitalis-induced ventricular tachycardia for an average of 2.0 +/- 1.8 minutes. This effect was not observed after a saline bolus. We conclude that an intracoronary bolus of low-dose lidocaine can suppress acetylstrophanthidin-induced ventricular arrhythmia.

Pressure-induced cardiac hypertrophy: changes in Na+,K+-ATPase and glycoside actions in cats.

Effects of myocardial hypertrophy caused by pressure overload on sarcolemmal Na+,K+-ATPase and positive inotropic action of strophanthidin were examined in cats. Partial ligation of the main pulmonary artery for four weeks resulted in right ventricular hypertrophy with no significant changes in left ventricular muscle. Hypertrophy was associated with a reduction in the number of active Na+,K+-ATPase units. Affinity of the remaining enzyme for [3H]ouabain was unchanged. No apparent right or left shift in dose-response curve for the positive inotropic effect of strophanthidin was observed and toxic concentrations of strophanthidin were unchanged; however, the degree of the positive inotropic effect produced by high concentrations of strophanthidin was significantly smaller in hypertrophied muscle. Moreover, decreases in developed tension rather than tachyarrhythmias was the predominant form of toxicity observed in hypertrophied muscle. These results indicate that myocardial hypertrophy reduces the number of active Na+,K+-ATPase units per milligram protein, decreases maximal positive inotropic effect of strophanthidin, and alters the prevailing form of strophanthidin toxicity.

Interaction of ischemia and reperfusion with subtoxic concentrations of acetylstrophanthidin in isolated cardiac ventricular tissues: effects on mechanisms of arrhythmia.

The aim of this study was to determine if "ischemia" and/or reperfusion potentiate digitalis toxicity through effects on oscillatory afterpotentials. Isolated canine Purkinje tissue-papillary muscle preparations were studied using standard microelectrode techniques. Tissues were superfused for 10 min with an "ischemic" solution that mimicked hypoxia, acidosis, elevated lactate, zero substrate and normo- or hyperkalemia. Reperfusion with "normal" Tyrode's solution was then reinstated for 60 min. Next, subthreshold oscillatory after potentials were induced with acetylstrophanthidin (ACS) and the protocol was repeated with ACS in all solutions. Without ACS, ischemic conditions with 4 mM KCl caused depolarization of Purkinje and muscle tissues. Reperfusion resulted in hyperpolarization of Purkinje tissue followed by mild depolarization, and then recovery. Purkinje tissue exposed to ischemic conditions with hyperkalemia responded similarly, except that hyperpolarization upon reperfusion was absent. In the presence of ACS, ischemic conditions with 4 mM KCl abolished oscillatory afterpotentials and caused marked depolarization of Purkinje tissue. Reperfusion decreased the coupling intervals and increased the amplitude of oscillatory afterpotentials relative to pre-ischemic levels, and frequently elicited arrhythmic activity. Arrhythmias ceased and tissues recovered by 60 min of reperfusion. Ischemic conditions incorporating hyperkalemia also abolished ACS-induced oscillatory afterpotentials and delayed their reappearance upon reperfusion. All other reperfusion responses were similar. This study demonstrates that "ischemic" suppresses oscillatory afterpotential-mediated effects of digitalis in canine Purkinje tissue, whereas reperfusion potentiates oscillatory afterpotential-induced arrhythmias.

Digitalis-induced mechanical toxicity: protection by slow Ca++ channel blockers.

In the in vitro perfusion of the isolated heart, toxic doses of cardiac glycosides produce an inotropic response which is followed by a decline in contractile force and an increase in the resting tension. Several reports in the literature indicate that the subsequent decline in contractile force may be related to cardiac cellular Ca++ overload. The purpose of the present study was to determine if the slow Ca++ channel blockers such as verapamil and nifedipine, which block Ca++ influx through voltage-dependent gated channels, can reduce or prevent the digitalis-induced decline in contractile force (mechanical toxicity). Langendorff preparations of isolated perfused guinea pig heart were used for the present study. The data obtained demonstrate that 1 to 2 microM ouabain in the perfusion medium produced mechanical toxicity in the hearts after an initial inotropic response. Verapamil or nifedipine, when combined with ouabain in the perfusion medium, increased the magnitude of the inotropic response and delayed or abolished the mechanical toxicity in a dose-dependent manner. No changes in the sarcolemmal Na+,K+-adenosine triphosphatase or ouabain binding were observed in the presence of verapamil or nifedipine. The data suggest that simultaneous use of verapamil or nifedipine may protect against digitalis-induced mechanical toxicity.

Arrhythmogenic dose of acetylstrophanthidin unchanged by beta-sympathomimetics in conscious dogs.

The enhanced arrhythmogenic risk of combined treatment with cardiac glycosides and beta-sympathomimetics is referred in some textbooks, but only a few detailed studies on in vivo models are available. We therefore investigated this problem in conscious dogs in an intraindividual study. We determined the dose of acetylstrophanthidin (intravenous infusion of 5 mcg/kg per min), which provoked ventricular premature beats with and without concomitant treatment with the partial beta-agonistic compounds doxaminol (3 mg/kg p.o.), prenalterol (0.4 or 1.0 mg/kg p.o.) or isoprenaline (0.31 +/- 0.100 mcg/kg per min). In some dogs a coronary artery was narrowed in order to reduce the coronary blood supply. The arrhythmogenic dose of acetylstrophanthidin was nearly the same in all the groups investigated (range from 52.1 +/- 5.66 to 59.9 +/- 3.23 mcg/kg). Whereas the arrhythmogenic dose of acetylstrophanthidin was unchanged by beta-sympathomimetics, the combination of the glycoside and each of the beta-agonistic drugs increased the contractile force more than did either single compound. We therefore conclude that the arrhythmogenic risk of the combination of glycosides and beta-sympathomimetics may be--at least in experimental models--less than has been suggested in the past.

Effects of verapamil on the arrhythmogenic action of acetylstrophanthidin.

Although coadministration of verapamil and digoxin results in significant increases in plasma glycoside concentrations, evidence of digitalis toxicity appears to be infrequent with this combination. To evaluate the effect of verapamil on electrophysiologic toxicity from digitalis, 5 anesthetized dogs were instrumented for physiologic recording and given acetylstrophanthidin by intravenous infusion until evidence of toxicity appeared. Each animal was then treated with verapamil intravenously, with mean steady-state plasma levels of 177 +/- 30 ng/ml, and acetylstrophanthidin infusion repeated; after return of sinus rhythm, the verapamil infusion was increased (producing mean levels of 379 +/- 50 ng/ml) and acetylstrophanthidin given a third time. Prior to verapamil dosing, ventricular ectopy was the manifestation of glycoside toxicity; following the first verapamil infusion, only 20% of the dogs developed ectopy, the remainder having second- or third-degree atrioventricular (AV) block, or AV junctional tachycardia. With the higher verapamil dose, AV block or junctional tachycardia occurred in all animals during acetylstrophanthidin infusion. In addition, the dose of glycoside required to produce electrophysiologic toxicity was significantly increased by verapamil. Therefore, verapamil appears to exert a protective effect against the development of digitalis-induced arrhythmia, possibly by suppressing delayed afterpotential generation, and significantly increases the dose of digitalis required to produce AV block.

Calcium overload and strophanthidin-induced mechanical toxicity in cardiac Purkinje fibers.

In cardiac Purkinje fibers, strophanthidin increases and then decreases contractile force. The relationship between the decrease in force and calcium overload was studied by recording the electrical and mechanical activity under conditions known to increase calcium overload or its effects. Inhibitors of oxidative phosphorylation reduced the positive inotropy of strophanthidin and enhanced the decrease in force. These inhibitors also reduced the inotropic effect of high calcium. Increasing intracellular calcium by decreasing extracellular sodium concentration also resulted in a decrease in the strophanthidin inotropy. When arrhythmia was delayed, strophanthidin induced contracture and this was favored by blockers of glycolysis and by enhancing cellular calcium. Some of these effects were also observed in ventricular muscle fibers but at higher strophanthidin concentrations. The results suggest that the decline in contractile force during strophanthidin exposure is related to calcium overload, although it is made clear that in Purkinje fibers contractile force and resting force may be independently affected under suitable conditions.

The effect of local anaesthetics on strophanthidin toxicity in canine cardiac Purkinje fibres.

1. Canine Purkinje fibres were superfused in vitro and the electrical and mechanical effects of the local anaesthetics benzocaine and procaine were studied in the absence and in the presence of other agents.2. Both benzocaine (1 x 10(-4)-5 x 10(-4)m) and procaine (6 x 10(-5)-2.5 x 10(-4)m) decreased slightly the amplitude of the upstroke and markedly the duration of the action potential. The plateau was shifted to more negative values and the force of contraction was decreased. These effects were dose-dependent.3. The local anaesthetics abolished the spontaneous activity induced by strophanthidin (5 x 10(-7)m) by flattening the oscillatory potential in diastole and increased the force of contraction under these circumstances.4. The local anaesthetics significantly delayed the time of the onset of the spontaneous activity induced by strophanthidin (10(-6)m). Also, the intensity of the stimuli had to be increased and the rate of discharge of intoxicated fibres was slower in the presence of local anaesthetics. In contrast, the positive inotropic effect was little affected.5. The local anaesthetics reduced but did not block the inotropic action of norepinephrine and high Ca; and did not abolish (while Mn did) small action potential in 27 mm-K-depolarized fibres.6. In fibres treated with local anaesthetics, lowering [Ca](o) did not result in a force rebound and administration of caffeine or exposure to low Na resulted in a larger increase in force.7. In fibres loaded with Ca, local anaesthetics caused an increase in force.8. Local anaesthetics decreased the force more when the external Na concentration was lower.9. This work shows that the local anaesthetics alter the mechanical performance of Purkinje fibres and lead to a depression of the strophanthidin induced oscillatory potential. As for the mechanism for these changes, the present experiments support the hypothesis that local anaesthetic agents have an antiarrhythmic action by decreasing intracellular Na and therefore intracellular Ca as suggested by Perry, McKinney & DeWeer (1978) on the basis of experiments on nerve.

The influence of reduced serum potassium level on the toxicity of some cardenolides in guinea pigs.

Recent experiments on the pharmacological properties of the semisynthetic cardiotonic steroid strophanthidin-3-bromoacetate (SBA) have challenged the well-known potassium digitalis antagonism in isolated heart muscle preparations. In order to establish these results in vivo, the minimum lethal doses (LDmin) of ouabain (OUA), digoxin (DO), digotoxin (DT), k-strophanthidin (STR) and SBA were determined by the infusion toxicity method in guinea pigs at normokalemia and hypokalemia. The experimentally induced decrease of the serum potassium concentration (5.0 mmoles/l vs. 3.3 mmoles/l) significantly reduced the LDmin of DO (1.42 vs. 1.05 mumoles/kg), DT (1.78 vs. 1.24 mumoles/kg) and STR (20.16 vs. 15.98 mumoles/kg), whereas the LDmin of OUA (0.37 vs. 0.34 mumoles/kg) was not altered. Contrary, the LDmin of SBA was even slightly, but not significantly increased during hypokalemia (16.77 vs. 19.04 mumoles/kg). In addition, from the experimental data an optimum time of infusion (Topt), corresponding to the LDmin, can be derived, which is equivalent to the time for optimum "utilization" of the drug. The obtained sequence: STR less than OUA less than DO less than DT less than SBA represents the well-known differences in the onset of the pharmacological action in man resp. animal. Hypokalemia in general resulted in a shortening of the Topt, thus indicating a more rapid "utilization" of the drug tested. The above differences of the cardenolide action at reduced serum potassium concentration may be dependent on the recently reported divergent influence of potassium on the association- resp. dissociation rate constants for the interaction of these drugs with their binding site at the Na+-K+-ATPase.

Interactions of norepinephrine and strophanthidin in cardiac Purkinje fibers.

The actions and interactions between strophanthidin and norepinephrine on electrical and mechanical events were studied in cardiac canine Purkinje fibers perfused in vitro. The results obtained show that norepinephrine (but not strophanthidin) shifts phase 1 and the beginning of the plateau to a more positive potential, an effect reduced by beta-receptor blockade. Norepinephrine increases the contractile force to the maximal value sooner than strophanthidin. In low Ca solution, the inotropic action of strophanthidin far exceeds that of norepinephrine. Norepinephrine does not cause spontaneous discharge in the driven fibers, whereas strophanthidin eventually induces fast spontaneous rhythms. When norepinephrine is given in the presence of strophanthidin (or vice versa) spontaneous activity is induced which consists either of a slow rhythm (characterized by large action potentials) or of a fast rhythm (characterized by small action potentials). The latter is typically induced by strophanthidin alone but occurs sooner in the presence of both agents. Propranolol prevents the onset of the slow rhythm but not that of the fast rhythm. It is concluded that norepinephrine and strophanthidin increase force by different mechanisms and that the potentiation between these agents in causing spontaneous discharge may involve an enhancement of diastolic depolarization(slow rhythm) or of the oscillatory potential (fast rhythm).