Nagammal Venkatesh

Sulfonylureas, ATP-sensitive K+ channels, and cellular K+ loss during hypoxia, ischemia, and metabolic inhibition in mammalian ventricle.

Sulfonylurea derivatives glibenclamide and tolbutamide are selective blockers of ATP-sensitive K+ (KATP) channels. However, their ability to prevent cellular K+ loss and shortening of action potential duration during ischemia or hypoxia in the intact heart is modest compared with their efficacy at blocking KATP channels in excised membrane patches. In the isolated arterially perfused rabbit interventricular septum, the increase in unidirectional K+ efflux and shortening of action potential duration during substrate-free hypoxia were effectively blocked by glibenclamide, but only by very high concentrations (100 microM); during hypoxia with glucose present, glibenclamide was only partially effective at reducing K+ loss. During total global ischemia (10 minutes), up to 100 microM glibenclamide or 1 mM tolbutamide attenuated shortening of action potential duration but only reduced [K+]0 accumulation by a maximum of 32 +/- 6%. In isolated patch-clamped guinea pig ventricular myocytes in which the whole-cell ATP-sensitive K+ current was activated by exposure to the metabolic inhibitors, glibenclamide (up to 100 microM) and tolbutamide (10 mM) were only partially effective at blocking the whole-cell ATP-sensitive K+ current (maximum block, 51 +/- 10% and 50 +/- 9%, respectively), especially when ADP was included in the patch electrode solution. In inside-out membrane patches excised from these myocytes, glibenclamide blocked unitary currents through KATP channels with a Kd of 0.5 microM and a Hill coefficient of 0.5 in the absence of ADP at the cytosolic membrane surface, but block was incomplete when 100 microM ADP (+2 mM free Mg2+) was present. ADP had a similar effect on block of KATP channels by tolbutamide. These findings suggest that free cytosolic [ADP], which rises rapidly to the 100 microM range during early myocardial ischemia and hypoxia, may account for the limited efficacy of sulfonylureas at blocking ischemic and hypoxic cellular K+ loss under these conditions.

ATP‐sensitive K+ channels and cellular K+ loss in hypoxic and ischaemic mammalian ventricle.

1. The contribution of ATP‐sensitive K+ (K+ATP) channels to the rapid increase in cellular K+ efflux and shortening of action potential duration (APD) during early myocardial ischaemia and hypoxia remains controversial, because for the first 10 min of ischaemia or hypoxia in intact hearts cytosolic [ATP] remains about two orders of magnitude greater than the [ATP] causing half‐maximal blockade of K+ATP channels in excised membrane patches. The purpose of this study was to investigate this apparent discrepancy. 2. During substrate‐free hypoxia, total, diastolic and systolic unidirectional K+ efflux rates increased by 43, 26 and 103% respectively after 8.3 min in isolated arterially perfused rabbit interventricular septa loaded with 42K+. APD shortened by 39%. From the Goldman‐Hodgkin‐Katz equation, the relative increases in systolic and diastolic K+ efflux rates were consistent with activation of a voltage‐independent K+ conductance. 3. During total global ischaemia, [K+]o measured with intramyocardial valinomycin K(+)‐sensitive electrodes increased at a maximal rate of 0.68 mM min‐1, which could be explained by a less than 26% increase in unidirectional K+ efflux rate (assuming no change in K+ influx), less than the increase during hypoxia. APD shortened by 23% over 10 min. 4. During hypoxia and ischaemia, cytosolic [ATP] decreased by about one‐third from 6.8 +/‐ 0.5 to 4.3 +/‐ 0.3 and 4.6 +/‐ 0.4 mM respectively, and free cytosolic [ADP] increased from 15 to 95 and approximately 63 microM respectively. 5. To estimate the percentage of activation of current through K+ATP channels (IK,ATP) necessary to double the systolic K+ efflux rate (comparable to the increase during hypoxia), K+ efflux during a single simulated action potential was measured by blocking non‐K+ currents under control conditions and after IK,ATP was fully activated by metabolic inhibitors. Activation of 0.41 +/‐ 0.07% of maximal IK,ATP was sufficient to double the systolic K+ efflux rate. The equivalent amount of constant hyperpolarizing current also shortened the APD in the isolated myocytes by 41 +/‐ 5%, compared to the 39% APD shortening observed during hypoxia in the intact heart. 6. The degree of activation of IK,ATP expected to occur during hypoxia and ischaemia was estimated by characterizing the ATP sensitivity of K+ATP channels in the presence of 2 mM‐free Mgi2+ and 0, 10, 100 and 300 microM‐ADPi in inside‐out membrane patches excised from guinea‐pig ventricular myocytes.(ABSTRACT TRUNCATED AT 400 WORDS)

Metabolic regulation of cardiac ATP-sensitive K+ channels

SummaryActivation of ATP-sensitive K+ (KATP) channels has been implicated as a cause of increased cellular K+ efflux and action potential duration (APD) shortening during myocardial ischemia, hypoxia, and selective glycolytic inhibition, since selective KATP channel antagonists partially or completely block increased cellular K+ efflux and APD shortening under these conditions. During substrate-free hypoxia or myocardial ischemia in intact rabbit ventricle, unidirectional K+ efflux rate during systole approximately doubled and APD decreased by ≈40% after 10 minutes. In patch-clamped guinea pig ventricular myocytes, similar changes could be produced by activation of <0.5% of the maximal KATP channel conductance. Furthermore, from studying the desensitizing effects of ADPi on the ATP sensitivity of KATP channels in excised inside-out patches, it was estimated that the rapid changes in the cytosolic ATP/ADP ratio during ischemia and hypoxia were of sufficient magnitude to activate KATP channels to this degree. During selective glycolytic inhibition, however, the global cytosolic ATP/ADP ratio in intact heart remained normal despite an increase in cellular K+ efflux comparable to ischemia and hypoxia. In patch-clamped saponin-permeabilized ventricular myocytes, KATP channels were preferentially suppressed by glycolytic ATP production compared to ATP generated by mitochondria or by the creatinine kinase reaction, and functional glycolytic enzymes were found to be associated with KATP channels in excised membrane patches. We hypothesize that sarcolemma-associated glycolytic enzymes may be important in maintaining a high local cytosolic ATP/ADP ratio in the vicinity of KATP channels, where sarcolemmal ATPases are tending to depress the local ATP/ADP ratio.

Effects of Amiodarone and Desethylamiodarone on Rabbit Myocardial β-adrenoceptors and Serum Thyroid Hormones—absence of Relationship to Serum and Myocardial Drug Concentrations

Summary: The antiadrenergic actions of amiodarone (Am) are well known but its effect and that of its metabolite, desethylamiodarone (DAm), on β-receptor density (Bmax) and affinity (KD) are poorly defined. Thus, the acute and chronic effects of Am and DAm on myocardial β-receptors in rabbits were determined relative to changes in thyroid hormones and serum and tissue drug concentrations. Bmax and KD were measured by radio-ligand binding, thyroid hormones by RIA. and drug levels by HPLC. Compared with controls, intravenous Am (20 mg/kg) reduced Bmax by 23% (p < 0.05) and DAm (20 mg/kg) by 32% (p < 0.05). After 3 weeks of chronic drug, the corresponding value for Am was 24% (p < 0.05) versus 45% (p < 0.05) for DAm. The effect of DAm was significantly greater (p < 0.05) than that of Am, being comparable to that of Am ( −44%) after 6 weeks. In the case of Am, doubling the dose (and myocardial level) led to no further decrease in Bmax DAm also reduced Bmax more following chronic treatment than after acute administration (−45 versus −32%), a difference of borderline significance. Following 3 weeks of p.o. Am, T3 decreased 3% (NS) and reverse T3 (rT3) increased 90% (p < 0.05); after 6 weeks, the corresponding values were 25% (p < 0.05) and 181% (p < 0.01). After 1 week of p.o. DAm, T3 did not change but rT3 increased by 34% (p < 0.05); after 3 weeks the corresponding values were 21% (p < 0.01) and 64% (p < 0.01). Neither compound affected serum T4 levels. Thus, Am and DAm reduce Bmax acutely with a trend for further reduction after chronic therapy as a function of time but not of serum and myocardial drug levels.

Activation of ATP-sensitive K+ channels by cromakalim. Effects on cellular K+ loss and cardiac function in ischemic and reperfused mammalian ventricle.

Pharmacological modulation of [K+]o accumulation and action potential changes during acute myocardial ischemia is under evaluation as a promising new antiarrhythmic and cardioprotective strategy during myocardial ischemia and reperfusion. We studied the effects of cromakalim, a K+ channel opener that activates ATP-sensitive K+ channels, in isolated arterially perfused rabbit interventricular septa subjected to ischemia and reperfusion and, through use of the patch clamp technique, in inside-out membrane patches excised from guinea pig ventricular myocytes. During aerobic perfusion, 5 microM cromakalim shortened action potential duration (APD) from 217 +/- 7 to 201 +/- 10 msec, had no effect on [K+]o, and reduced tension by 17 +/- 3% (n = 11). During ischemia, pretreatment with 5 microM cromakalim resulted in 1) more rapid APD shortening (71 +/- 9 versus 166 +/- 7 msec at 10 minutes and 63 +/- 12 versus 122 +/- 8 msec at 30 minutes), 2) similar [K+]o accumulation after 10 minutes (8.9 +/- 0.3 versus 9.6 +/- 0.5 mM) but a trend toward increased [K+]o accumulation after 30 minutes (11.0 +/- 1.7 versus 9.6 +/- 1.0 mM), and 3) similar times for tension to decline to 50% of control (2.14 +/- 0.16 versus 2.14 +/- 0.19 minutes) but shorter time to fall to 20% of control (4.34 +/- 0.33 versus 4.90 +/- 0.22 minutes; p = 0.003). After 60 minutes of reperfusion following 30 minutes of ischemia, recovery of function was similar, with a trend toward better recovery of developed tension (to 58 +/- 9% versus 39 +/- 10% of control; p = 0.18) and tissue ATP levels in cromakalim-treated hearts but no differences in APD or rest tension. Thus, 5 microM cromakalim had mild effects in normal heart but greatly accelerated APD shortening during ischemia without markedly increasing [K+]o accumulation, possibly because the more rapid APD shortening reduced the time-averaged driving force for K+ efflux through ATP-sensitive K+ channels. A significant cardioprotective effect during 30 minutes of ischemia plus 60 minutes of reperfusion could not be demonstrated in this model. In excised membrane patches studied at room temperature, the ability of cromakalim to activate ATP-sensitive K+ channels was significantly potentiated by 100 microM but not 15 microM cytosolic ADP, suggesting that in addition to the modest fall in cytosolic ATP during early ischemia, the rapid increases in cytosolic ADP may further sensitize cardiac ATP-sensitive K+ channels to activation by cromakalim.(ABSTRACT TRUNCATED AT 400 WORDS)

Electropharmacology of amiodarone: Absence of relationship to serum, myocardial, and cardiac sarcolemmal membrane drug concentrations

Plasma concentrations are often of major consideration in the evaluation of therapeutic efficacy of cardiovascular drugs. This approach is based on the assumptions that the concentration of the drug in the cardiac muscle is in equilibrium with the plasma drug level and that pharmacologic efficacy is proportional to the myocardial drug concentration. The more pronounced pharmacologic efficacy of amiodarone following chronic administration, despite low plasma drug concentrations, and the lesser effects of the drug after acute intravenous administration, when drug levels are maximum, has not been explained on the basis of the pharmacokinetic behavior of the drug. Data obtained from the transmembrane action potential recordings from rabbit ventricular myocardium were therefore correlated with drug concentrations in the serum, myocardium, and myocardial sarcolemma following acute intravenous administration and after 4 weeks of oral administration of 20 mg/kg/day of amiodarone. Following 15 minutes of acute drug administration, when amiodarone concentrations were maximal in the serum (4.72 +/- 1.23 micrograms/ml), cardiac muscle (34.5 +/- 7.6 micrograms/gm), and sarcolemma (1.94 mg/gm protein), the electrophysiologic changes were insignificant. However, following chronic treatment, when levels of amiodarone were low in the serum (0.05 +/- 0.01 micrograms/ml amiodarone, 0.25 +/- 0.08 micrograms/ml desethylamiodarone), cardiac muscle (1.91 +/- 0.9 micrograms/gm amiodarone, 1.35 +/- 1.33 micrograms/gm desethylamiodarone), and myocardial membranes (0.043 mg/gm protein [amiodarone], 0.097 mg/gm protein [desethylamiodarone], there was a 54.3% increase in action potential duration at 90% repolarization (p less than 0.01) and 65% increase in the effective refractory period (p less than 0.01) of rabbit ventricular myocardium.(ABSTRACT TRUNCATED AT 250 WORDS)

Hypothyroidism renders protection against lethal ventricular arrhythmias in a conscious canine model of sudden death

The protective effect of hypothyroidism against lethal ventricular tachyarrhythmias (VT) in the subacute phase of experimental myocardial infarction (MI) was investigated in 10 thyroidectomized dogs using a conscious model of sudden coronary death. Four weeks after surgical ablation of the thyroid, and having established biochemical hypothyroidism, anterior MI was produced by 120 min of occlusion-reperfusion of the left anterior descending coronary artery. In the subacute phase of MI, the inducibility of VT was investigated using programmed ventricular stimulation (PVS). and the effects on spontaneous development of ventricular fibrillation (VF) were studied by production of posterolateral ischemia at a site remote from the area of the previous infarction. Ischemia was produced by the passage of anodal direct current through a silver wire electrode implanted in the left circumflex coronary (LCX) artery. The results were compared to those from a cohort of 20 existing euthyroid controls that had undergone an identical experimental protocol. No differences were found in heart rate and other electrocardiographic parameters such as the PR, QRS. and QT (paced at 2.5 Hz) and the QTC interval between the hypo- and euthyroid groups. During PVS in the subacute phase of anterior MI, the measured threshold voltage and ventricular refractory periods were similar in both groups. The incidence of inducibility of VT was 100% in the euthyroid animals compared to 60% in the hypothyroid dogs, suggesting an antiarrhythmic effect of hypothyroidism. The incidence of sustained vs. non-sustained VT was similar in both groups. When ischemia was produced in the distribution of the LCX artery, lethal ischemic VF occurred in 4 of 10 (40%) hypothyroid dogs, whereas the incidence of ischemic VF was 90% in the euthyroid controls. The lower incidence of PVS-induced VT and the development of spontaneous VF suggests that hypothyroidism renders protection from cardiac rhythm disturbance in the subacute phase of MI.

effects of N-acetylprocainamide and Recainam in the Pharmacologic Conversion and Suppression of Experimental Canine Atrial Flutter : significance of Changes in Refractoriness and Conduction

The electrophysiologic determinants of the pharmacologic conversion and the prevention of atrial flutter are poorly defined. This study investigated the effects of pharmacologically induced changes in atrial conduction velocity and refractoriness, in the conversion and suppression of atrial flutter induced in the open-chest anesthetized dog by intercaval crush and rapid atrial pacing. The effects of an intravenous infusion of the new class III antiarrhythmic drug N-acetylprocainamide (30 mg/kg over 15 min) and the class Ic antiarrhythmic drug recainam (10 mg/kg over 20 min followed by 10 mg/kg/h) were evaluated. N-acetylprocainamide restored sinus rhythm in 10 of 15 (66%) dogs, while recainam converted only 2 of 10 (20%). N-acetylprocainamide prevented reinduction in 3 (20%), while recainam was effective in none. In the atria, N-acetylprocainamide induced significant increases in effective refractory period (+27%, p less than 0.01), functional refractory period (+22%, p less than 0.01), and in atrial flutter cycle length (+13%, p less than 0.01). Recainam increased effective refractory period (+28%, p less than 0.01), functional refractory period (+20%, p less than 0.01), conduction time at atrial paced cycle length of 150 msec (+70%, p less than 0.01) and atrial flutter cycle length (+56%, p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Acute and chronic effects of ketanserin on the electrophysiologic properties of isolated rabbit ventricular myocardium: Particular reference to repolarization

The acute and chronic cellular electrophysiologic actions of ketanserin, a selective blocker of 5-hydroxytriptamine (5-HT2) receptor, were examined in the isolated rabbit ventricular muscle. This work was prompted by the recent observations that small numbers of patients treated with hypertension develop QTc prolongation, rarely associated with torsades de pointes. At 1.0 Hz stimulation, ketanserin, 10(-5) and 10(-4) M, prolonged the action potential duration (APD) (by 20% and 29%, respectively) and voltage-dependent refractoriness. At 10(-4) M, the maximal rate of rise of phase 0 of the action potential (Vmax) decreased 11%. Action potential amplitude and resting membrane potential were not affected by either concentration of ketanserin used (10(-6) to 10(-4) M). Trains of stimuli at rates of 1.0 Hz or higher led to an exponential decline in Vmax to a new plateau level. The time constant for the recovery of Vmax from the use-dependent block was 1.3 second. Chronic administration of ketanserin (40 mg/kg/day, intramuscularly) caused a significant prolongation of APD (106%; p less than 0.01) and voltage-dependent refractoriness without effects on the action potential amplitude, resting membrane potential, and Vmax. These data indicate that ketanserin exerts significant class I effects with mild class III effects when superfused acutely, whereas chronic administration of ketanserin exhibits marked class III effects. Both effects of the drug are likely to exert significant antiarrhythmic actions.

Digoxin−desethylamiodarone interaction in the rat: comparison with the effects of amiodarone

Summary: We have shown that there is a pharmacokinetic interaction between amiodarone and digoxin that results in an increase in steady-state serum and tissue concentrations of digoxin in rats. There is a linear correlation between serum levels of amiodarone, as well as desethylamiodarone, and steady-state serum digoxin levels in rats treated with amiodarone. Since desethylamiodarone is formed in amounts equal to that of the parent compound during chronic amiodarone therapy, we investigated the possibility of desethylamiodarone directly interacting with digoxin in rats. Rats that received digoxin alone showed a serum level of 0.32 ± 0.08 ng/ml, whereas those that received combination therapies showed a serum level of 3.25 ± 1.06 ng/ml (p < 0.001) with desethylamiodarone administration, and 3.00 ± 0.87 ng/ml with amiodarone administration. Concomitant administration of desethylamiodarone and digoxin increased digoxin concentration in the myocardium by 110% (p < 0.001), in the skeletal muscle by 208% (p < 0.001) and in the brain by 110% (p < 0.001). The corresponding figures for amiodarone-digoxin administration were 94% (p < 0.001), 172% (p < 0.001) and 80% (p < 0.001). The tissue/serum ratios of digoxin concentrations in the myocardium, skeletal muscle, and brain were decreased in the rats that received combination therapies, indicating reduced tissue binding of digoxin. The data indicate that desethylamiodarone interacts with digoxin in a manner similar to that of the parent compound.