Repolarization Level Research Articles

Validation of automatically measured monophasic action potential recordings

This study devised an automatic monophasic action potential (MAP) measurement program, and compared the computer-measured MAP durations with those measured manually by two independent observers in order to facilitate the analysis of MAP data obtained during a clinical electrophysiology study. The results were compared at various cycle lengths and during pharmacologic or physiologic interventions. This program identified the onset, plateau, and baseline accurately using the MAP data. The automatically measured MAP durations at the 90% repolarization level (MAPd) strongly correlated with those measured manually ( r=0.99). The MAPd shortened in parallel as the pacing cycle length was progressively shortened from 800 to 600, 400 and 300 ms .

The electrical restitution curve revisited: steep or flat slope--which is better?

The electrical restitution curve (ERC) traditionally describes the recovery of action potential duration (APD) as a function of the interbeat interval or, more correctly, the diastolic interval (DI). Often overlooked in modeling studies, the normal ventricular ERC is triphasic, starting with a steep initial recovery at the shortest DIs, a transient decline, and a final asymptotic rise to a plateau phase reached at long DIs. Recent studies have proposed that it would be advantageous to lower the slope of the ERC by drug intervention, as this might reduce the potential for electrical alternans and ventricular fibrillation. This review discusses the pros and cons of a flat versus steep slope of the ERC and draws attention to mechanisms thatjustify the (physiologically) steep slope, rather than a flat slope, as a better design against arrhythmias. Five potential mechanisms are discussed, which allows for a different interpretation of the effect of the slope on arrhythmogenicity. The most important appears to be the physiologic rate adaptive shortening of APD that, by reciprocal lengthening of the DI, allows the subsequent APD to move more quickly from the steep initial ERC phase onto the flat phase. A less steep initial ERC phase would protract the transition toward more fully recovered APD and, in fact, may perpetuate electrical alternans. The triphasic ERC time course in normal myocardium cannot be explained by or fitted to single exponentials or single ion channel recovery kinetics. A simple tri-ionic model is suggested that may help explain the shape of the ERC at various repolarization levels and place APD recovery into perspective with intracellular calcium recycling and recovery of contractile force.

Increased vulnerability of brain mitochondria in diabetic (Goto-Kakizaki) rats with aging and amyloid-beta exposure.

This study evaluated the respiratory indexes (respiratory control ratio [RCR] and ADP/O ratio), mitochondrial transmembrane potential (DeltaPsim), repolarization lag phase, repolarization level, ATP/ADP ratio, and induction of the permeability transition pore of brain mitochondria isolated from normal Wistar and GK diabetic rats of different ages (1.5, 12, and 24 months of age). The effect of amyloid beta-peptides, 50 micromol/l Abeta(25-35) or 2 micromol/l Abeta(1-40), on mitochondrial function was also analyzed. Aging of diabetic mice induced a decrease in brain mitochondrial RCR, ADP/O, and ATP/ADP ratios but induced an increase in the repolarization lag phase. Brain mitochondria from older diabetic rats were more prone to the induction of the permeability transition pore, i.e., mitochondria from 24-month-old diabetic rats accumulated much less Ca(2+) (20 micromol/l) than those isolated from 12-month-old rats (50 micromol/l) or 1.5-month-old rats (100 micromol/l). In the presence of 50 micromol/l Abeta(25-35) or 2 micromol/l Abeta(1-40), age-related mitochondrial effects were potentiated. These results indicate that diabetes-related mitochondrial dysfunction is exacerbated by aging and/or by the presence of neurotoxic agents such as amyloid beta-peptides, supporting the idea that diabetes and aging are risk factors for the neurodegeneration induced by these peptides.

The interaction of carbachol and strophantin on the electrical and mechanical events in electrically driven strips of guinea pig left atria.

We compared the electrophysiological and the mechanical actions of the cholinergic agonist carbachol (CARB) and the digitalis glycoside strophantin (STR) in a study on the electrically driven strips of guinea pig left atria. The respective control values for the resting membrane potential (RMP), action potential amplitude (APA), action potential duration at 20%, 50%, and 90% repolarization levels (APD20,50,90), and time to peak tension (TtPt) were of the order of 61.1 +/- 1.69 nmV, 104.64 +/- 1.28 mV, 28.5 +/- 1.22 msec, 51.0 +/- 1.51 msec, 106.91 +/- 3.81 msec, and 60.1 +/- 1.09 msec. Exposure to CARB (2 x 10(-7) M) rapidly reduced contractility, TtPt, APD, and APA to exceptionally lower levels within the initial 5 minutes and gradually caused a slight hyperpolarization in RMP. Strophantin (1 x 10(-7) M) caused contractility and RMP to increase slightly, APD and APA to reduce less markedly, but did not significantly affect TtPt. Under the influence of combined CARB/STR, the effect of CARB on the contraction amplitude (CA), TtPt, APD, and APA was attenuated and that on RMP was significantly potentiated (p < 0.05), whereas the effects of STR on CA, TtPt, APD, and APA were reversed (p < 0.05), and the effect on RMP was augmented. The action potential duration was less responsive to STR at all depolarization levels with respect to those of CARB and combined CARB/STR. Although the correlation between the time course of APD-20 and APD-90 for STR was poor, the correlation for CARB and combined CARB/STR was highly significant (r > 0.95). The correlation between the effects of CARB on CA and APD-90 was high but poor for the effect of combined CARB/STR. Striking to note is that the rate of induction of contractile changes with CARB (d(delta)/dt = 1.39 +/- 0.05%/sec), and its washout was more rapid (p < 0.05) in comparison to the rate of changes in APD-90 (0.98 +/- 0.05%/sec). This result, however, was not observed with combined CARB/STR. From the results we concluded that CARB and STR act as antagonists when used in combination, with the exception that only the effect on RMP was additive, with asignificant discordance between the rates of induction of effects on CA and APD-90, with the former being more rapid.

Electrophysiological properties of rainbow trout cardiac myocytes in serum-free primary culture.

A low-density primary culture of trout ventricular myocytes in serum-free growth medium was established and maintained for up to 10 days at 17 degrees C. The myocytes retained their normal rod shaped morphology, capacitive surface area of the sarcolemma (SL), and contractile quiescence. However, sarcolemmal cation currents changed significantly, some permanently, some transiently, after 8-10 days of culture. TTX-sensitive sodium current (I(Na)) and Ba(2+)-sensitive background inward rectifier potassium current (I(K1)) were permanently depressed to 24-28% of their control density measured in freshly isolated myocytes. In contrast, L-type calcium current (I(Ca)) was only transiently downregulated; after 2-3 days in culture, the density of the current was 32% of the control and recovered to the control value after 8-10 days in culture. The changes in membrane currents were reflected in the shape of the action potential (AP). After 2-3 days in culture, maximal overshoot potential and resting potential were significantly reduced, and the durations of the AP at 50 and 90% repolarization were significantly increased. These changes became significantly more pronounced after 8-10 days of culture, with the exception of AP duration at 50% repolarization level. The shortening of the early plateau phase may reflect an additional change to an outward current, presumably the rapid component of the delayed rectifier (I(Kr)). Although the present findings indicate that fish cardiac myocytes can be maintained in serum-free primary culture for at least 10 days at 17 degrees C, some but not all of the electrophysiological characteristics of the myocytes change markedly during culture. The changes in ion currents were not due to loss of sarcolemmal membrane and therefore are likely to represent altered expression of cation currents as an adaptive response to culture conditions.

Chronic Administration of Amiodarone Does Not Affect Na+/Ca2+ Exchange Current in Guinea Pig Cardiac Ventricular Myocytes

We investigated chronic effects of amiodarone on Na+/Ca2+ exchange current (INCX) and on the level of Na+/Ca2+ exchanger (NCX1) mRNA in guinea pig ventricular myocytes using the whole-cell clamp technique and RT-PCR analysis, respectively. Guinea pigs were intraperitoneally injected with 80 mg/kg per day of amiodarone or the vehicle (saline) for 1 or 4 weeks. Single ventricular cells were isolated from the hearts of both groups of animals. Action potential duration at 90% repolarization level was prolonged to 143% and 165% of the control values by treatment with amiodarone for 1 and 4 weeks, respectively. INCX density and the level of NCX1 mRNA were not significantly changed by chronic treatment with amiodarone. The level of thyroid hormone (T4) within the blood was not changed by the treatments. These results suggest that chronic treatment with amiodarone does not affect the Na+/Ca2+ exchanger, with respect to the level of its mRNA and current density in guinea pig ventricular myocytes.

Effects of a Typical IKr Channel Blocker Sematilide on the Relationship Between Ventricular Repolarization, Refractoriness and Onset of Torsades de Pointes

The effects of a typical I(Kr) channel blocker sematilide on the relationship between ventricular repolarization, refractoriness and onset of torsades de pointes (TdP) were studied using the canine isolated, blood-perfused ventricular septum preparation with monophasic action potential (MAP) recording. Intracoronary infusion of sematilide (10-100 microg/min) prolonged the repolarization phase and effective refractory period, the extent of which was greater in the former than in the latter, resulting in prolongation of terminal repolarization process. Prolonging the basic pacing cycle length from 400 to 600 ms and/or increasing the drug doses enhanced each of these actions. Reverse use-dependence was obvious in the drug-induced prolongation of MAP duration, but it was less clear in the effective refractory period. More importantly, during sematilide infusion, in preparations paced at longer basic cycle length of 600 - 2000 ms, TdP-like polymorphic ventricular tachycardia was repeatedly induced by an extra-stimulus applied on the terminal repolarization phase, which indicates the appearance of electrically vulnerable period. Prolonging the basic pacing cycle length and/or increasing the drug doses prolonged this electrically vulnerable period in parallel with the terminal repolarization phase. These results suggest that prolongation of the terminal repolarization process by sematilide would enhance the chance of conduction slowing at less complete repolarization levels, which may be associated with a high incidence of TdP induction.

Load-induced changes in repolarization: evidence from experimental and clinical data.

The high incidence of arrhythmias in patients with heart failure, hypertension, valvular heart disease, or mitral walve prolapse suggests a strong link between wall motion abnormalities and arrhythmias. A potential common mechanism underlying these observations may be that overload leads to electrophysiologic changes and facilitates arrhythmias. This article summarizes the interaction between changes in atrial and ventricular loading conditions and repolarization. Most experimental and clinical studies demonstrated 1) a reduction of action potential duration and refractoriness, 2) development of early afterdepolarizations, and 3) ectopic beats originating from these afterdepolarizations. Discrepancies between studies are related to different study designs, i.e., varying magnitude, velocity, and timing of increased load, the level of repolarization at which action potential duration is measured as well as different animal species. Direct effects of increased load on repolarization are most likely caused by activation of stretch-activated nonselective cation ion channels and changes in calcium handling. Current antiarrhythmic drug therapy is aimed at electrical disorders as the primary cause of arrhythmias. If mechanical disorders play a central role in the genesis of cardiac arrhythmias, future treatment should be directed at restoring a more normal mechanical function of the heart. Additional studies will further clarify the nature and clinical significance of load-related changes in repolarization and arrhythmogenesis.

Effects of dopamine on porcine myocardial action potentials and contractions at 37 °C and 32 °C

Background: Little information exists on the effects of drugs with cardiovascular action in hypothermia, and some findings have indicated paradoxic effects of dopamine in this setting. As we have not found any data on the electrophysiologic and contractile effects of dopamine on the heart in hypothermia, we decided to study this in pig myocardium, since pigs have a cardiovascular system more similar to that of humans than other animals.Methods: Excised muscle strips from pig ventricular septum were mounted in an organ bath. After 45 min of equilibration at 37 °C or 32 °C, resting and action potentials, time to peak contraction and contractile force were recorded during pacing with a frequency of 60/min. Dopamine at 4 μM or 8 μM was added and new recordings were made after 15 min.Results: Cooling to 32 °C caused a prolongation of contraction by 48% and the contractile force increased by 39%. The membrane action potential duration at 50% and 90% repolarization levels increased at 32 °C by 28% and 16% respectively. Dopamine significantly (P&lt;0.05) increased the contractile force and membrane action potential duration at 50% and 90% repolarization levels both in normothermia and in hypothermia, whereas the duration of the contraction was not significantly changed.Conclusion: Cooling to 32 °C significantly prolongs the myocardial action potential and the contraction duration. Dopamine increases the contractile force and prolongs the action potential both at 37 °C and at 32 °C.

The repolarization-excitability relationship in the human right atrium is unaffected by cycle length, recording site and prior arrhythmias

OBJECTIVESThe goal of this study was to determine the relationship between repolarization and excitability in the human atrium under various conditions.BACKGROUNDAction potential duration (APD) measurements from monophasic action potential (MAP) recordings provide a surrogate for measuring the effective refractory period (ERP) in human ventricle. The relationship between repolarization and refractoriness in human atrium and the effect of prior atrial fibrillation/flutter on the ERP/APD correlation are unknown.METHODSSeven patients with sinus rhythm and 15 patients after conversion of atrial flutter or fibrillation were evaluated. Monophasic action potentials were recorded at multiple right atrial sites and during different basic cycle lengths from 300 to 700 ms, while ERPs were determined by extrastimulus technique using the MAP recording-pacing combination catheter.RESULTSThere was a close correlation between ERP and APD at 70% repolarization (APD70, r = 0.97; p < 0.001) and 90% repolarization (APD90, r = 0.98; p < 0.001), respectively. Refractoriness occurred at a repolarization level of 72 ± 8%. The ERP/APD70and ERP/APD90ratios averaged 1.06 ± 0.10 and 0.86 ± 0.08, respectively. These ratios were nearly constant over the entire range of basic cycle lengths, between different sites in individual patients and between different patients. Patients cardioverted from atrial fibrillation or flutter exhibited no significant differences in the ERP/APD relationship compared with patients with sinus rhythm.CONCLUSIONSEffective refractory period and APD are closely related in the human right atrium. Using the MAP recording technique, atrial ERPs can be assessed by measurement of APDs. Effective refractory period is most closely reflected by APD70. Thus, MAP recordings allow investigation of the local activation and repolarization time course beat by beat, visualizing the excitable gap.

Effects of bimoclomol, the novel heat shock protein coinducer, in dog ventricular myocardium

The effects of the novel HSP-coinducer bimoclomol was studied on action potentials, ionic currents and [Ca 2+] i transients in isolated canine ventricular myocytes using conventional microelectrode techniques and whole cell voltage clamp combined with fluorescent [Ca 2+] i measurements. Contractility was studied in right ventricular trabeculae. All preparations were paced with a frequency of 0.2 Hz. Bimoclomol (100 μM) shortened action potential duration measured at 50% repolarization, but lengthened action potentials at the 90% repolarization level, decreased action potential amplitude and maximum depolarization velocity in a reversible manner. In voltage clamped myocytes, the drug activated a steady-state outward current at positive membrane potentials leaving the peak inward current unaffected. [Ca 2+] i transients, measured under voltage clamp control, were increased in amplitude and had accelerated decay kinetics in the presence of the compound, in addition to reduction of diastolic [Ca 2+] i. Bimoclomol significantly decreased the force of contraction in right ventricular trabeculae. Comparison of present data to previous results indicate that the cardiac effects of bimoclomol strongly depend on actual experimental conditions. The reduced contractility in spite of the increased amplitude of [Ca 2+] i transients suggests that 100 μM bimoclomol may decrease calcium sensitivity of the contractile apparatus.

Electrophysiological effects of bimoclomol in canine ventricular myocytes.

Concentration-dependent effects of bimoclomol, a novel heat shock protein (HSP) coinducer, were studied on the parameters of action potential and transmembrane ionic currents in enzymatically dispersed canine ventricular cardiomyocytes using conventional microelectrode and whole cell voltage clamp techniques. Bimoclomol (10-100 microM) decreased the maximum velocity of depolarization (Vmax) and amplitude of action potentials in a concentration-dependent manner. These effects were fully reversible after a 5-min period of washout in drug-free medium. Action potential duration measured at 50% or 90% level of repolarization (APD-50 and APD-90, respectively) was markedly shortened by bimoclomol. Both APD-50 and APD-90 were decreased, but the reduction in APD-50 was more pronounced. The APD-shortening effect of bimoclomol was significantly reduced in the presence of 20 nM charybdotoxin (inhibitor of the Ca-dependent K current) or 0.5 mM anthracene-9-carboxylic acid (inhibitor of the Ca-dependent Cl current) or 1 microM glibenclamide (inhibitor of the ATP-sensitive K current). In the presence of anthracene-9-carboxylic acid, APD-90 was lengthened by bimoclomol. The APD-shortening effect of bimoclomol was also partially antagonized by chelation of intracellular Ca2+ by application of the cell permeant form of BAPTA, or when using 10 mM EGTA-containing patch pipettes to record action potentials. The Vmax-depressant effect of bimoclomol was not affected by charybdotoxin, anthracene-9-carboxylic acid, glibenclamide, or BAPTA load. In voltage clamped cardiomyocytes bimoclomol (100 microM) had no effect on the amplitude of I(Ca), but decreased significantly the inactivation time constant of I(Ca) (from 19.8+/-1.6 ms to 16.8+/-1.2 ms at 0 mV). Bimoclomol also decreased significantly the amplitude of I(K1) (from -20.5+/-1.1 pA/pF to -16.6+/-0.8 pA/pF at -135 mV), causing reduction in slope of the negative branch of the I-V curve. At positive potentials, however, bimoclomol increased outward current. The bimoclomol-induced current, therefore, was studied in the presence of BaCl2, when I(K1) current was blocked. The bimoclomol-induced current had a reversal potential close to -90 mV. Bimoclomol (100 microM) had no effect on the amplitude or kinetic properties of the transient outward K current (I(to)) and the delayed rectifier K current (I(K)). It is concluded that bimoclomol exerts both Ca-independent (inhibition of I(Na) and I(K1), activation of the ATP-sensitive K current) and Ca-dependent effects (mediated by Ca-activated Cl and probably K currents) in canine ventricular myocytes.

Electrophysiological properties of ventricular muscle obtained from spontaneously diabetic mice.

The electrophysiological properties of cardiac muscle in KK/Ta mouse (hereafter referred to as KK mouse), an animal model of human non-insulin-dependent diabetes mellitus, were investigated, and the findings compared with those obtained from a non-diabetic control mouse (C57BL/6J mouse; referred to as B6 mouse). The ages of the B6 mice were 23.9 +/- 5.4 weeks (n = 24) and those of the KK mice used were 25.7 +/- 10.8 weeks (n = 34). The KK mice had mild obesity, hyperglycemia and hyperinsulinemia. Ventricular muscles from both mice were examined by light microscopy. Partial myocardial fibrosis and filament disorder in the ventricular muscles were found only in the KK mice. The resting membrane potential of the ventricular muscle was less negative in the KK mice than in the control mice. The maximum rate of rise in the upstroke of the action potential was significantly decreased in the KK mice compared with that of the control mice. These suggest a decrease in a time-independent K+ current (IK1) in the KK mice. The duration of the action potential (APD) at all levels of repolarization was significantly longer in the KK mice than in the B6 mice. A blocker of transient outward current (I(to)), 4-aminopyridine, significantly prolonged the APD of the B6 mice, but failed to prolong it in the KK mice, suggesting that Ito in the diabetic mice is very small. A Ca2+ channel blocker, CoCl2, dramatically lengthened all levels of APD in both groups, suggesting that there is no difference between B6 mice and KK mice in L-type Ca2+ current via Ca2+ channels. These suggest the malfunction or deficiency of ionic channels which carry, at least Ito and IK1 in diabetic mice.

Role of action potential shortening in the prevention of arrhythmias in canine cardiac tissue.

1. The effects of the K+ channel opener diazoxide and the oxime-containing Ca2+ and K+ channel blocker salicylaldoxime were tested in canine cardiac Purkinje tissue. 2. Both drugs shortened action potential duration (APD). For salicylaldoxime (0.1-1.0 mmol/L), the reductions in APD were statistically significant at the 25% level of repolarization (APD25) for 0.1 mmol/L (P < 0.05, n = 14) and 0.5 and 1.0 mmol/L (P < 0.01, n = 6), at the 50% level of repolarization (APD50) for 0.1 mmol/L (P < 0.05, n = 14) and 0.5 and 1.0 mmol/L (P < 0.01, n = 6) and at the 90% level of repolarization (APD90) for 0.5 and 1.0 mmol/L (P < 0.01, n = 6). In contrast, diazoxide (0.05-0.1 mmol/L) significantly shortened APD at all levels of repolarizations, with the APD50 and APD90 reduced most significantly (P < 0.01, n = 6) for higher concentrations of the drug (0.07-0.1 mmol/L). Both drugs significantly reduced the force of contraction. 3. Diazoxide (10 experiments) was more potent in suppressing strophanthidin-induced arrhythmias than salicylaldoxime (three of seven experiments). Salicylaldoxime reduced APD even further in the presence of diazoxide. 4. Although salicylaldoxime and diazoxide modulate different ion channels, it appears APD shortening may be a necessary, but insufficient, factor for the suppression of strophanthidin-induced arrhythmias.

Electrophysiological changes in rat ventricular and atrial myocardium at different stages of experimental diabetes.

Action potential configuration in ventricular and atrial myocardium, as well as rate-dependent changes in ventricular action potential duration (APD) were studied and compared in healthy and diabetic rats. Diabetes was induced by a single injection of streptozotocin (STZ, 65 mg kg(-1) i.v.). Conventional microelectrode techniques were applied to record action potentials after the establishment of diabetes (2, 6, 10 and 18 weeks after STZ-treatment). Untreated age-matched animals were used as controls. Both depolarization and repolarization were significantly retarded following STZ-treatment. However, the time course of development of diabetic changes in atrial and ventricular myocardium was different. APD was significantly lengthened from week 2 of diabetes in ventricular, but only from week 6 in atrial preparations. In atrial myocardium, lengthening of APD was more pronounced at early rather than late phases of repolarization. The maximum rate of depolarization (Vmax) was significantly reduced from the 6th week of diabetes in both preparations. No differences were observed in action potential amplitude (except at week 18) and in the resting membrane potential in diabetic rats. Diabetic ventricular preparations showed a positive APD-frequency relationship at any level of repolarization, in contrast to control muscles, where APD25 and APD50 values lengthened. But APD75 and APD90 values were not changed significantly with increase in the pacing frequency. The results indicate that development of diabetic alterations are not fully identical in atrial and ventricular myocardium of the rat, probably owing to differences in density and kinetics of ionic currents responsible for atrial and ventricular action potentials.

Mechanisms of stretch-induced changes in [Ca2+]i in rat atrial myocytes: role of increased troponin C affinity and stretch-activated ion channels.

To study the effects of stretch on the function of rat left atrium, we recorded contraction force, calcium transients, and intracellular action potentials (APs) during stretch manipulations. The stretch of the atrium was controlled by intra-atrial pressure. The Frank-Starling behavior of the atrium was manifested as a biphasic increase of the contraction force after increasing the stretch level. The development of the contraction force after step increase of the stretch (intra-atrial pressure from 1 to 3 mm Hg) was accompanied by the increase in the amplitude of the calcium transients (P<0.05, n=4) and decrease in the time constant of the Ca2+ transient decay. The APs of the individual myocytes were also affected by stretch; the duration of the AP was decreased at positive voltages (AP duration at 15% repolarization level, P<0.001; n=13) and increased at negative voltages (AP duration at 90% repolarization level, P<0. 01; n=13). To study the mechanisms causing these changes we developed a mathematical model describing [Ca2+]i and electrical behavior of single rat atrial myocytes. Stretch was simulated in the model by increasing the troponin (TnC) sensitivity and/or applying a stretch-activated (SA) calcium influx. We mimicked the Ca2+ influx by introducing a nonselective cationic conductance, the SA channels, into the membrane. Neither of the 2 plausible mechanosensors (TnC or SA channels) alone could produce similar changes in the Ca2+ transients or APs as seen in the experiments. The model simulated the effects of stretch seen in experiments best when both the TnC affinity and the SA conductance activation were applied simultaneously. The SA channel activation led to gradual augmentation of Ca2+ transients, which modulated the APs through increased Na+/Ca2+-exchanger inward current. The role of TnC affinity change was to modulate the Ca2+ transients, stabilize the diastolic [Ca2+]i, and presumably to produce the immediate increase of the contraction force after stretch seen in experiments. Furthermore, we found that the same mechanism that caused the normal physiological responses to stretch could also generate arrhythmogenic afterpotentials at high stretch levels in the model.

Class I antiarrhythmic drugs alter the severity of myocardial stunning by modulating ATP-sensitive K+ channels in guinea pig ventricular muscles.

The effects of various class I antiarrhythmic drugs and glibenclamide were examined on the recovery of contraction during reperfusion, in relation to the action potential duration (APD) seen during ischemia. Action potential and contractile tension were recorded from isolated guinea pig right ventricular muscles perfused with oxygenated Tyrode solution via the coronary artery. Ten minutes of no-flow ischemia shortened the APD at 90% of repolarization level (APD90) to 58% of control (pre-ischemic values). The APD90 was completely restored after 60 min of reperfusion. The developed tension was abolished during ischemia and recovered to 87% of control after 60 min of reperfusion. In the presence of Vaughan Williams class Ia drug cibenzoline (5 microM) or an ATP-sensitive potassium (K(ATP)) channel blocker glibenclamide (10 microM), the shortening of the APD90 during ischemia was significantly attenuated. However, the recovery of developed tension was significantly inhibited. Class Ic drug pilsicainide (10 microM) did not affect the ischemia-induced shortening of the APD90 or the recovery of developed tension after reperfusion. In the presence of class Ib drug mexiletine (10 microM), the shortening of the APD90 during ischemia was significantly facilitated. The recovery of developed tension in the presence of mexiletine tended to be improved, although the difference was not statistically significant. The developed tension measured after the 60 min reperfusion period following 20 min of no-flow ischemia was markedly depressed, indicating the presence of myocardial stunning. Mexiletine and pilsicainide significantly improved the recovery of developed tension and diminished the stunning. We conclude that cibenzoline and glibenclamide, which block cardiac K(ATP) channels inhibit contractile recovery after reperfusion by attenuating the shortening of APD during ischemia. In contrast, mexiletine, which activates K(ATP) channels (in addition to blockade of Na+ channels) improves contractile recovery by facilitating the shortening of APD during ischemia.

Different inhibition patterns of tedisamil for fast and slowly inactivating transient outward current in rat ventricular myocytes.

Tedisamil has been described as a selective inhibitor of a fast inactivating transient outward current (i(to,f)) in rat ventricular myocytes. Because recent reports demonstrated the existence of a second slowly inactivating transient component (i(to,s)) we investigated i(to,s) and differentiated the effects of tedisamil on both transient outward current components and their influence on action potential duration. Standard electrophysiological techniques were used for whole cell recordings at 24-26 degrees C from enzymatically isolated myocytes. Inhibition of i(to,f) by tedisamil was the result of an acceleration of inactivation at positive test potentials with a concentration for half-maximal inhibition (EC50) of 4-7 micromol/l, which is confirmatory to reports from other investigators. Our new results show that i(to,s) is more sensitive to tedisamil with an EC50 of 0.5 micromol/l. Furthermore the pattern of i(to,s) inhibition is different compared with i(to,f), because inactivation of i(to,s) is not accelerated by tedisamil. Instead the amplitude of the steady state inactivation curve of i(to,s) is attenuated which indicates a reduction of maximally available current. I(to,s) was evaluated by three different methods as time-dependently inactivating current (7.5 s test pulse duration), voltage-dependently inactivated current and tedisamil-sensitive current. All approaches yield similar inactivation curves. The potential for halfmaximal inactivation of i(to,s) lies about 35 mV more negative than that for i(to,f) and the slope factor (K = -23 mV) is different to that of i(to,f) (K = -3 mV). Effectiveness of tedisamil-induced modulation of i(to,f) and i(to,s) on action potential repolarization was tested. Action potentials stimulated at 0.5 Hz were not prolonged by 1 micromol/l tedisamil (dominant i(to,s) block) at a repolarization level of 0 mV but prolonged to about 120% of control at -70 mV. This indicates that i(to,f) was sufficient to guarantee a regular early repolarization whereas decrease of i(to,s) delayed the final repolarization. In conclusion, the observation that tedisamil inhibits i(to,f) and i(to,s) differently supports the hypothesis that the two i(to)-components are related to two different channel populations expressed in rat ventricular myocytes.

Stretch-induced voltage changes in the isolated beating heart: importance of the timing of stretch and implications for stretch-activated ion channels

It is now well recognized that myocardial stretch can cause arrhythmias due to stretch-induced depolarizations. The effects of transient stretch applied during the various phases of the cardiac action potential have not been investigated. This study (1) examined the effects of short stretch pulses and sustained stretch on the monophasic action potential (MPA) repolarization time course and diastolic potential, (2) examined the arrhythmic response to differently timed stretch pulses, and (3) tested by comparison with computer simulations whether these effects are compatible with stretch-activated channel characteristics known from patch-clamp studies. We studied the MAP changes elicited by short transient stretch pulses applied at different times during the cardiac cycle to 8 isolated Langendorff-perfused rabbit hearts. The left ventricle (LV) was instrumented with a fluid-filled balloon, the volume of which was altered rapidly and precisely by means of a computer-controlled linear motor-driven piston. MAPs were recorded simultaneously from one right ventricular (RV) and two LV sites while short volume pulses of increasing amplitude were applied to the LV at variable delays after the last of 8 regular electrical pacing stimuli. The effect of pulsatile volume pulses applied at different phases of electrical systole and diastole was compared to the effect of sustained stretch pulses (60 s duration) of the same amplitude. The experimental results were compared with computer simulations of stretch-induced effects on the action potential to further validate the experimentally measured effects with theoretical predictions based on the Oxford Heart model with added stretch channel terms. Stretch pulses applied during early systole caused a brief transient repolarization during the LV MAP plateau phase, with a maximal amplitude of 24 +/- 10% of the total MAP amplitude. Stretch pulses at the end of the MAP caused a transient depolarization, with a maximal amplitude of 13 +/- 5%. These oppositely polarized stretch effects crossed over during a transitional range of repolarization (mean 65 +/- 9% of repolarization) when stretch produced neither transient repolarizations nor depolarizations. Only stretch pulses applied at a mean repolarization level of 77 +/- 5% or later led to arrhythmias, preceded by transient depolarizations. No corresponding de- or repolarizations were seen in MAPs recorded simultaneously from the unstretched RV. The effects of long pulses on the MAP waveform were nearly identical to an overlay plot of the effects of many differently timed short transient pulses. When the stretch-induced voltage changes in the MAP were plotted against the repolarization level at which they were produced, a linear relationship was found (mean correlation coefficient r = 0.97; P < 0.0001) with a reversal at approximately half the total MAP amplitude. The computer simulations of the influence of stretch-activated channels reproduced both the effects of short and sustained stretch seen in the MAP recordings. We demonstrated in the isolated beating heart that the electrophysiologic effects of sudden myocardial stretch depend on the timing of the stretch relative to electrical systole or diastole. These findings are in agreement with patch clamp studies on stretch-activated ion channels which showed a linear current/voltage relation with a reversal potential between -20 and -30 mV. Only stretch pulses applied at the end of the action potential or during diastole elicit ectopic beats as a result of transient depolarizations, while stretch pulses applied during phase 2 and 3 cause transient repolarizations or no effect, respectively.

Class III antiarrhythmic effects of ceruloplasmin on rat heart

Recently it has been shown that ceruloplasmin presents a protective action against reperfusion-induced arrhythmias in the isolated perfused rat heart, an effect that is lost when the protein is denaturated by heat. The present study was carried out to see whether ceruloplasmin can alter electrophysiological properties such as ventricular effective refractory periods, conduction time, and action potential duration calculated at 50, 75, and 90% levels of repolarization (APD50, APD75, APD90). To check the specificity of the electrophysiological effects of ceruloplasmin, we have also compared them with those of heat-denatured ceruloplasmin, superoxide dismutase, catalase, deferoxamine, and albumin. In isolated rat hearts, ceruloplasmin (0.25-3 microM) (n = 8 for each concentration) was shown to increase the effective refractory period in a concentration-dependent manner by 26 to 89%. Conduction time was not significantly altered. Heat-denatured ceruloplasmin (0.50-3 microM) (n = 8 for each concentration) increased the effective refractory period by 33 to 70% and did not affect the conduction time. In contrast, superoxide dismutase (1-4 microM), catalase (1-2 microM), deferoxamine (500 microM-1 mM), and albumin (1-4 microM) (n = 8 for each substance and for each concentration) had no significant effect on effective refractory period and conduction time at any dose, suggesting that the ceruloplasmin effect might be specific. In rat ventricular preparations, ceruloplasmin (1 microM) also induced a constant prolongation of APD50 (52%), APD75 (64%), and APD90 (41%) after 15 min of infusion (n = 6). The prolongation of effective refractory period and of action potential duration, by native and heat-denatured ceruloplasmin, suggests that this substance has specific class III effects, although this cannot entirely account for its antifibrillatory action at reperfusion in isolated rat hearts.