Tachycardia-Induced Changes in Na+ Current in a Chronic Dog Model of Atrial Fibrillation
Abstract We have previously shown that chronic rapid atrial activation (400 bpm) reduces atrial conduction velocity in dogs, contributing to the development of a substrate supporting sustained atrial fibrillation (AF). However, the cellular and ionic mechanisms underlying these functional changes have not been defined. We applied whole-cell patch-clamp techniques to atrial myocytes from dogs subjected to atrial pacing at 400 bpm for 7 days (P7, n=6) and 42 days (P42, n=5) and compared the results with those from sham-operated dogs similarly instrumented but without pacemaker activation (P0, n=6). Rapid atrial pacing allowed for the induction of sustained AF in 67% and 100% of dogs paced for 7 and 42 days, respectively, and significantly decreased conduction velocity under P7 and P42 conditions. In dogs paced for 7 days, Na+ current (INa) density was reduced by 28% at −40 mV (P<.0001, n=59 cells). INa changes were even more decreased under P42 conditions, by ≈52% at −40 mV (P<.0001): from −78.7±4.6 pA/pF (P0, n=28 cells) to −37.7±3.0 pA/pF (P42, n=43 cells). INa was significantly reduced at all voltages ranging from −65 to −10 mV. Voltage-dependent activation and inactivation properties, activation kinetics, and recovery from inactivation were not altered by rapid atrial pacing; however, inactivation kinetics were slowed. AF duration was related to mean INa in each dog (r2=.573, P<.001). We conclude that rapid atrial activation significantly reduces both conduction velocity and INa density. Since INa is a major determinant of conduction velocity, our data point to INa reduction as a potentially important mechanism contributing to the substrate for AF in this model.
Atrial fibrillation is the most frequently encountered arrhythmia in clinical practice and is likely to become more common with the aging of the population.1–3 To study AF, investigators have used short-term experimental models in which AF has been maintained either pharmacologically with acotinine4 or by vagal nerve stimulation.5–7 It has long been known that most clinical AF occurs in patients with atrial disease and is often associated with atrial dilation. These characteristics are absent in short-term AF models, limiting their relevance to clinical AF.
Recently, several chronic animal models of AF that are associated with chronic atrial cardiomyopathy and ultrastructural changes resembling those seen in clinical AF have been developed.8–10 One of these models uses rapid atrial pacing to cause 1:1 atrial activation at 400/min.10 In addition to producing clinically relevant atrial structural changes, this chronic dog model allows for studies of the relationship between time-dependent electrophysiological and underlying ionic changes during the development of AF. Furthermore, these models are relevant to the way in which sustained AF alters atrial properties to promote its own maintenance, a phenomenon known as “the domestication of AF” or “AF begets AF.”9
Wijffels et al9 noted that when AF was maintained electrically in chronically instrumented goats, the electrophysiological milieu changed gradually to allow for spontaneous maintenance of AF. ERP decreased to attain near-minimal values within 24 hours after the onset of AF induction, whereas changes in spontaneous AF persistence developed more slowly, stabilizing after ≈1 week. Morillo et al10 reported that chronic rapid atrial pacing (400/min) of dogs for 6 weeks decreased atrial ERP and allowed for the induction of sustained AF in >80% of the animals. They also noted P-wave prolongation after 6 weeks of pacing, but they did not assess conduction velocity directly or study the time course of electrophysiological changes. We recently found that chronic atrial tachycardia causes progressive decreases in atrial ERP, ERP adaptation to rate, and conduction velocity.11 The time course of changes in conduction velocity is somewhat slower than that of ERP alterations, appearing to account for continuing increases in AF susceptibility after ERP changes had stabilized.
Little information is available regarding the cellular changes underlying the changes in ERP and conduction velocity caused by chronic atrial tachycardia. In a previous study, we presented evidence for ionic remodeling of ICa as the mechanism underlying ERP changes.12 Phase-0 INa is an important determinant of conduction velocity in fast-channel tissues like the atria,13 but INa changes have not been examined in the atrial tachycardia model of AF.
The present study was designed to assess whether changes in INa density occur and whether they are related to changes in conduction velocity and the ability to maintain AF among dogs subjected to atrial pacing at 400/min for periods of up to 6 weeks. Specifically, we wished to determine (1) whether INa is altered by rapid atrial pacing, (2) whether any changes in current density are associated with and/or due to changes in the voltage-dependent properties and recovery kinetics of INa, (3) whether atrial conduction changes are related to INa alterations, and (4) whether changes in AF duration are related to changes in INa.
Materials and Methods
Pacemaker Insertion and Rapid Atrial Pacing
Adult mongrel dogs of either sex (28.5±2.3 kg, n=17) were anesthetized with sodium pentobarbital (30 mg/kg IV, followed by intravenous boluses of 4 mg/kg as needed). Artificial respiration was maintained via an endotracheal tube connected to a Harvard-type mechanical ventilator. Under sterile technique, a unipolar screw-in Medtronic J pacing lead (Medtronic Inc) was inserted through the right jugular vein, and the distal end of the lead was screwed in the right atrial appendage under fluoroscopic guidance, as previously described.10 Initial atrial capture was verified with the use of an external demand pacemaker (GBM 5880, Medtronic Inc). The proximal end of the pacing lead was then connected to a custom-modified implantable Medtronic pacemaker unit (model 8084), which was inserted into a subcutaneous pocket in the neck.
Twenty-four hours was allowed for the lead to stabilize. The pacemaker was then programmed to stimulate the atrium at 400/min (150-ms cycle length) with the use of 0.42-ms square-wave pulses at twice-threshold current. The atrium was continuously stimulated at 400/min for a pacing period of 7 days (P7, n=6) or 42 days (P42, n=5) days. The surface ECG was verified after 24 hours and then weekly to ensure continuous 1:1 atrial capture. Dogs subjected to rapid atrial pacing were compared with sham-operated dogs that were similarly instrumented but maintained without pacemaker activation for periods ranging from 7 to 42 days (P0, n=6). We have previously noted that the properties of sham-operated dogs are stable over a range of observation periods from 1 to 42 days.
On study days, dogs were reanesthetized with morphine (2 mg/kg SC) and α-chloralose (120 mg/kg IV bolus, followed by a continuous infusion of 29.25 mg · kg−1 h−1) and ventilated with room air supplemented with oxygen. Respiratory parameters were adjusted to maintain physiological arterial blood gases (Sao2 >90%; pH 7.38 to 7.44). Body temperature was maintained at 37°C with a circulating-water temperature control system. Polyethylene catheters were inserted into the left femoral artery and both femoral veins and kept patent with heparinized saline solution (0.9%). The left femoral artery catheter was used to measure arterial blood pressure. The right and left femoral vein catheters were used to infuse saline and α-chloralose, respectively. A median sternotomy was performed, and a pericardial cradle was created. Two bipolar polytetrafluoroethylene-coated stainless-steel electrodes were inserted into the right atrial appendage for recording and stimulation. A programmable stimulator (Digital Cardiovascular Instruments) was used to deliver 2-ms pulses at twice-threshold current. A P23 1D transducer (Statham Medical Instruments), electrophysiological amplifiers (Bloom Ltd), and a paper recorder (Astromed Model MT-95000) were used to record arterial blood pressure, six standard surface ECG leads, a right atrial electrogram, and stimulus artifacts.
In order to obtain conduction velocity measurements, one thin silicone sheet containing 40 bipolar electrodes with 1-mm interpolar and 6-mm interelectrode distances was sewn into position on the right atrial epicardial surface, covering the posterior aspect of the atrial appendage and the free wall. Each signal was filtered (30 to 400 Hz), digitized with 12-bit resolution and a 1-kHz sampling rate, and transmitted into a microcomputer (model 286, Compaq Computer). Software routines were used to amplify, display, and analyze each electrogram signal as well as to generate activation maps.14 Each electrogram was analyzed with computer-determined peak-amplitude criteria and was reviewed manually. The accuracy of activation time measurements was ±0.5 ms.
Determination of AF Duration and Conduction Velocity
In paced dogs, a surface ECG was first recorded to confirm maintenance of 1:1 atrial pacing at 400/min. The pacemaker was then deactivated. In all groups, activation maps for conduction velocity measurements were obtained after 2 minutes of continuous pacing applied at the right atrial appendage at a short basic cycle length (150 ms), close to the cycle length of AF.
After data for conduction velocity measurements were obtained at all cycle lengths, AF was induced with a train of 10-Hz stimuli delivered to the right atrial appendage at four times threshold current and 2-ms duration. AF was defined as a rapid (>450/min) irregular atrial rhythm with varying atrial electrogram morphology. AF was considered sustained if it persisted for >45 minutes and was distinguished from atrial flutter on the basis of the irregularity of atrial electrogram morphology and frequency. In order to estimate the mean duration of AF, AF was induced 10 times if AF duration was <10 minutes. Two AF inductions were performed if episodes lasted between 10 and 45 minutes. If AF lasted for >45 minutes and no briefer episodes occurred, mean AF duration was based on this episode, and no further AF induction was attempted in order to avoid excessive prolongation of the experiment. Episodes of AF lasting >45 minutes were terminated by DC electrical cardioversion delivered with epicardial paddle electrodes. When electrical cardioversion was performed, a 30-minute rest period was allowed before continuing with the experimental protocol.
AF inducibility was assessed by noting the ability of single premature S2 stimuli at a 400-ms basic cycle length to induce the arrhythmia. AF was considered to be inducible in a given dog if AF was induced reproducibly by the S2 at a given coupling interval.
Activation maps were generated off-line after each experiment, and conduction velocity was determined by analyzing activation times at a series of four electrode sites in the direction of longitudinal propagation in the right atrial free wall. The distance of each site from the first of the series of electrodes was plotted against activation time, and conduction velocity was determined from the slope of the best-fit regression line.11,16 Activation maps were reviewed to ensure continuous longitudinal propagation, and only data with a clear linear relation (correlation coefficient, >.99) were accepted for analysis. The same sites were used for conduction velocity measurements for each experiment.
Cell Isolation and Solutions
At the end of the in vivo experimental protocol, the heart was excised and immersed in Tyrode’s solution containing (mmol/L) NaCl 136, CaCl2 2, KCl 5.4, MgCl 1.0, NaH2PO4 0.33, HEPES 5, and glucose 10 at room temperature (pH adjusted to 7.35 with NaOH). Canine atrial myocytes were isolated by enzymatic dissociation with the use of previously described methods.15 All solutions used for dissection and perfusion were equilibrated with 100% O2. The circumflex coronary artery was cannulated, and the left atrium was dissected free and perfused with Tyrode’s solution at 37°C for 5 minutes until the atrium was clear of blood. All leaking arterial branches were ligated with prolene thread to ensure adequate perfusion. The atrium was then perfused at 12 mL/min with nominally Ca2+-free Tyrode’s solution for 20 minutes. The solution was changed to one containing 110 U/mL collagenase (type II, Worthington) and 1% bovine serum albumin (Sigma Chemical Co). Softened tissue from a well-perfused region was removed with a forceps, gently triturated, and kept in a storage solution containing (mmol/L) KCl 20, KH2PO4 10, dextrose 10, l-glutamic acid 70, β-hydroxybutyric acid 10, taurine 10, and EGTA 10, along with 1% albumin (pH adjusted to 7.35 with KOH).
A small aliquot of the solution containing the isolated cells was placed in a 1-mL open perfusion chamber mounted on the stage of an inverted microscope. After 5 minutes to allow for cell adhesion to the chamber, the cells were perfused at 6 mL/min with a solution containing (mmol/L) NaCl 5, CsCl 132.5, MgCl2 1.0, CaCl2 1.0, HEPES 20, and glucose 11 (pH adjusted to 7.35 with CsOH). To provide optimal control of INa, the bath solution was kept at 17°C with the use of a Peltier effect device. The pipette solution used to record INa contained (mmol/L) NaCl 5, CsF 135, HEPES 5, EGTA 10, and Mg2ATP 5 (pH adjusted to 7.2 with CsOH).
INa was recorded using the whole-cell configuration of the voltage clamp technique. Only quiescent rod-shaped cells showing clear cross striations were used. Small cells were selected for study to facilitate voltage control. Borosilicate glass microelectrodes (outer diameter, 1.0 mm) had resistances ranging from 0.8 to 1.5 MΩ when filled with pipette solution and were connected to a patch-clamp amplifier (Axopatch-1D, Axon Instruments). Data were sampled with an A/D converter (Digidata 1200, Axon Instruments) and stored on the hard disk of a computer for subsequent analysis. The sampling frequency was 10 kHz for INa measurements. Recordings were low pass–filtered at 2 to 5 kHz.
Tip potentials were zeroed before formation of the pipette-membrane seal. Several minutes after seal formation, the membrane was ruptured by gentle suction to establish the whole-cell configuration. After rupture of the cell membrane, pipette Rs was electrically compensated to minimize the capacitive surge on the current recording and the voltage drop across the clamped membrane. Rs was calculated by dividing the capacitive time constant, obtained by fitting the decay of the capacitive transient, by the calculated membrane capacitance (the time integral of the capacitive response to a 5-mV hyperpolarizing pulse from a holding potential of −60 mV, divided by the voltage drop). Cell capacitance averaged 64.3±2.3 pF in 130 cells. Before compensation, Rs averaged 5.1±0.2 MΩ, and the capacitive time constant was 306.1±14.7 μs. The mean Rs and capacitive time constant after compensation averaged 1.4±0.1 MΩ and 78.3±2.4 μs, respectively. Care was taken to ensure that the maximum voltage drop across the Rs in each cell did not exceed 5 mV, and the mean voltage drop averaged 4.2±0.2 mV in 130 cells. Cells for which the series voltage error exceeded 5 mV were rejected. Overall, under P0 (sham) conditions, 39% of the cells (18 of 46 cells) had to be discarded because of voltage control problems, caused mainly by very large current amplitude. Mean peak current amplitude, voltage drop, and Rs in cells rejected from the P0 group because of poor voltage control averaged 7969±741 pA, 9.9±1.2 mV, and 1.3±0.4 MΩ, respectively. In contrast, because of a decreased peak current amplitude, fewer P7 and P42 cells were rejected (7 of 66, or 11% of the cells, for P7; 2 of 45, or 4% of the cells, for P42). Cells with significant leak currents were rejected. The small residual leak was subtracted by a linear leakage compensation algorithm.
Statistical comparisons of multiple group means were obtained by ANOVA. Nonorthogonal decomposition using Dunnett’s test for multiple comparisons with one control was used to study changes due to different rapid atrial pacing durations.17 Bartlett’s test was used to analyze the homogeneity of variances between sham-operated and paced groups.18 The Kruskal-Wallis test was used for nonparametric data with unpaired measures. Linear regression was used to analyze the relationship between single dependent and independent variables, whereas stepwise multilinear regression was used to assess the dependence of a single dependent variable on multiple potential contributors to its variance. A nonlinear least-square curve-fitting program (CLAMPFIT in pCLAMP 6.0 or Sigma Plot) was used to perform curve-fitting procedures. All average results are expressed as mean±SEM, and a value of P<.05 was considered statistically significant.
Effects of Continuous Rapid Atrial Pacing on Duration and Inducibility of AF
As shown in the Table⇓, rapid pacing resulted in an increased duration of induced AF, which averaged 13±4 s in sham-operated (P0) dogs, 2108±479 s (P<.005 versus P0) in P7 dogs, and 2700±0 s (P<.0001 versus P0) in P42 dogs. The number of dogs with sustained AF (>45 minutes) also increased over time, with sustained AF occurring in 0% of P0 dogs compared with 67% and 100% of P7 and P42 dogs, respectively. Under P0 conditions, single premature atrial beats failed to induce AF in any dogs. As the duration of rapid pacing increased, the vulnerability of the atria to AF induction by single extrastimuli at a 400-ms basic cycle length increased, so that by 42 days of rapid pacing, AF was induced in all dogs (100%).
Conduction Velocity Changes Produced by Continuous Rapid Atrial Pacing
In addition to changes in AF duration and inducibility with premature beats, rapid atrial stimulation decreased conduction velocity. The Table⇑ indicates the time-dependent changes in atrial conduction velocity with atrial tachycardia. Conduction velocity was somewhat decreased after 7 days of tachycardia and showed larger, statistically significant decreases at 42 days.
INa Changes Caused by Rapid Atrial Pacing
In order to avoid any contaminating effects of time-dependent changes in the INa I-V relationship, all studies were started 20 minutes after membrane rupture, with protocols performed in the same sequence in all groups of cells, beginning with the INa density-voltage relationship, followed immediately by analysis of voltage-dependent inactivation and, finally, time-dependent recovery from inactivation. Typical original current tracings from isolated canine atrial myocytes obtained from P0, P7, and P42 dogs are shown in Fig 1A⇓ to 1C. The data were obtained with a holding potential of −140 mV and a series of 5-mV depolarizing steps (0.1 Hz) to voltages ranging from −80 to −5 mV. Fig 1D⇓ shows mean INa density-voltage relations for P0 (n=28 cells), P7 (n=59 cells), and P42 (n=43 cells) cells. Maximum current was noted at −40 mV, and pacing did not alter the form of the INa density-voltage relation. INa density was reduced progressively and highly significantly by rapid atrial pacing. For example, at −40 mV, INa density averaged −78.7±4.6 pA/pF in P0 cells compared with −57.0±3.4 pA/pF (P<.0001 versus P0) in P7 cells and −37.7±3.0 pA/pF (P<.0001 versus P0) in P42 cells. INa was significantly reduced at voltages ranging from −65 to −10 mV under P42 conditions. As indicated by the data in the Table⇑, changes in INa occurred in consort with changes in conduction velocity, AF duration, and AF inducibility.
To evaluate possible mechanisms involved in the pacing-induced reduction in INa, we studied the voltage-dependent properties of the current. Fig 1E⇑ shows the voltage dependence of INa activation, which was calculated by dividing peak current during depolarizing test pulses by the driving force, with an estimated reversal potential of 0 mV as measured by extrapolating the terminal part of the I-V curve to the voltage-axis intercept. The data were well-fitted by a Boltzmann relation, with half-maximum activation voltage averaging −52.0±0.3, −54.6±0.3, and −50.0±0.2 mV under P0 (n=28 cells), P7 (n=59 cells), and P42 (n=43 cells) conditions, respectively (P=NS versus P0). The slope factor averaged 6.0±0.2, 6.3±0.3, and 6.0±0.2 mV for P0, P7, and P42 cells (P=NS versus P0). These results indicate that changes in INa density are not associated with or due to changes in the voltage dependence of INa activation.
A double-pulse protocol was used to assess the voltage dependence of INa inactivation, as shown in Fig 2⇓. A 1-s conditioning pulse to voltages between −140 and −65 mV was followed by a 30-ms test pulse to −40 mV (holding potential, −140 mV; frequency, 0.1 Hz). The peak current elicited by the test pulse was normalized to current without a prepulse. Panels A to C of Fig 2⇓ display typical original current recordings from atrial myocytes isolated from P0, P7, and P42 dogs. Fig 2D⇓ shows that voltage-dependent inactivation was complete at −80 mV under paced and nonpaced conditions. The results were well-fitted by a Boltzmann relation. Rapid pacing did not alter INa inactivation, with mean values for half-maximum activation voltage of −101.7±0.1, −101.7±0.3, and −101.0±0.2 mV in P0 (n=28 cells), P7 (n=59 cells), and P42 (n=43 cells) cells, respectively (P=NS versus P0). The slope factor averaged 5.8±0.1, 6.2±0.2, and 5.5±0.2 mV under P0, P7, and P42 conditions (P=NS versus P0).
The kinetics of INa activation and inactivation were analyzed by curve fitting to data obtained with the protocol shown in Fig 1⇑, inset. Monoexponential functions provided good fits to the data (Fig 3A⇓ to 3C). The resulting time constants shown in Fig 3D⇓ and 3E⇓ were voltage dependent. Although the activation time course was not altered by rapid pacing, inactivation kinetics slowed progressively in dogs paced for longer periods of time (n=28, 18, and 15 cells for kinetic analysis in P0, P7, and P42 dogs, respectively).
As illustrated in Fig 4⇓, the recovery kinetics of INa were evaluated with the use of a train of 40 pulses from a holding potential of −140 to −40 mV (50-ms pulse duration, 200-ms interstimulus interval), followed by a 30-ms test pulse to −40 mV at various recovery intervals. The INa recovery time course was not altered by rapid atrial pacing. Recovery was well-fitted by a monoexponential function, with time constants of 7.2±0.4, 7.6±0.5, and 6.3±0.3 ms in 30, 44, and 29 cells from P0, P7, and P42 dogs, respectively (P=NS for P7 and P42 versus P0).
Relation Between INa, Conduction Velocity, and AF Duration Changes
One of our goals was to determine whether INa changes were related to alterations in AF duration. The logarithm of AF duration correlated with mean INa in individual dogs (r2=.573, P=.0004). When conduction velocity and INa were included as covariates in a stepwise multilinear regression, the logarithm of AF duration was best predicted by a model including both INa (P=.0004) and conduction velocity (P=.025).
In the present study, we evaluated the time course of changes in conduction velocity, INa, and duration of AF in dogs subjected to chronic atrial tachycardia. We have shown that rapid atrial pacing decreases both INa and conduction velocity, while causing progressive increases in the ability of single atrial extrasystoles to induce AF and in the duration of AF induced by burst pacing. Regression analysis suggests that changes in both INa and conduction velocity are associated with the progressive tendency to maintain AF and that alterations in conduction velocity occur along with changes in INa.
Conduction Velocity Changes Related to AF
In animal models, rapid atrial activation results in progressive increases in AF duration, with AF increasingly likely to be sustained after a week of pacing.10,11 Changes in AF duration are associated with a reduction in ERP and ERP adaptation to rate, changes often seen in patients with increased atrial vulnerability and a tendency to exhibit atrial reentrant arrhythmias.19 Similar results have also been obtained in various animal models using either rapid atrial pacing10,11 or electrically maintained AF.9,20
In addition to ERP changes, anomalies of intra-atrial conduction have long been known to be associated with clinical AF and are often indicated by P-wave prolongations during sinus rhythm.21 Increased P-wave durations were observed in rapidly paced dogs by Morillo et al10 and more recently by Elvan et al.22 An increased signal-averaged P-wave duration was detected in hyperthyroid patients with paroxysmal AF, with P-wave changes being a good predictor of the induction of AF in these patients.23 Our previous work suggested a role of conduction slowing in contributing to the maintenance of AF in dogs with maintained rapid atrial activation.11 The present study provides further evidence that chronic atrial tachycardia may itself decrease conduction velocity. Conduction slowing reduces the wavelength for reentry and increases the susceptibility to reentrant atrial arrhythmias.24 Therefore, our findings point to a potential role of atrial conduction slowing due to rapid atrial activation in the perpetuation of atrial arrhythmias.
INa Changes Related to AF
INa plays a major role in the upstroke of the action potential in atrial myocytes and has been found to be an important determinant of conduction velocity in cardiac tissue.13 Consistent with the role of INa in controlling conduction speed, we found that changes in conduction velocity accompany INa alterations. INa is therefore a candidate to underlie the conduction velocity changes induced by rapid atrial pacing in the chronic dog model.
The present study provides the first direct evidence, to our knowledge, that chronic rapid atrial pacing decreases not only conduction velocity but also INa density. Several biophysical properties of Na+ channels, including the voltage dependence of activation and inactivation, the time course of activation, and the rate of recovery from inactivation, were not altered in rapidly paced dogs. On the other hand, inactivation development slowed in paced animals. These observations differ somewhat from our findings regarding changes in ICa and Ito density in atrial cells from dogs subjected to rapid atrial pacing, which were not associated with any changes in voltage- or time-dependent current properties.12 Kääb et al25 similarly observed that pacing-induced ventricular failure causes a reduction in Ito density without changing other channel properties.25
One of the possible mechanisms underlying INa alterations in dogs subjected to chronic tachycardia may be related to [Ca2+]i changes. Increases in [Ca2+]i can downregulate Na+ channel expression.26 Evidence of cellular Ca2+ overload has been obtained in histological studies of atria subjected to very rapid atrial activation.27 The authors found mitochondrial swelling consistent with cellular Ca2+ overload and a protective effect of verapamil against electrical remodeling.27 It is possible that in response to a transient rise in Na+ entry during AF due to an increased frequency of depolarization, reverse-mode Na+-Ca2+ exchange may promote Ca2+ entry into myocytes, triggering the release of Ca2+ from the sarcoplasmic reticulum,28 thereby generating Ca2+ overload and leading to INa decrease. The decrease in INa density may thus serve an autoregulatory function by reducing total Na+ entry.
INa is not the only determinant of conduction velocity. Changes in intercellular coupling may also be very important. Observations regarding changes in connexin expression in AF are preliminary and conflicting. One group noted reductions in connexin40 expression and no change in connexin43 in goats with chronic AF,20 whereas other investigators found an increase in connexin43 expression in dogs with AF.29 Further information is needed in order for time-dependent changes caused by rapid atrial activation in determinants of atrial conduction, including INa, connexin expression, and intercellular coupling, to be understood completely.
Potential Clinical Relevance
The present study is the first of which we are aware to evaluate the changes in INa caused by chronic rapid atrial activation. Our findings are relevant to mechanisms of altered atrial conduction and P-wave prolongation in clinical AF.21,23,30 Changes in INa may participate in the “domestication of AF” by slowing atrial conduction, whereas decreases in ICa appear also to contribute by accelerating atrial repolarization and abbreviating refractoriness.12 Thus, changes in both refractoriness and conduction appear to be involved in the promotion of AF maintenance by continuous atrial tachycardia.11 There is reason to believe that decreases in both INa and ICa may be due to Ca2+ overload. Preventing Ca2+ overload might thus prevent ionic remodeling and the domestication of AF. Evidence for such an effect on a short-term basis has recently been presented.27,31 These considerations point to the possibility of intervening to prevent the development of the substrate of AF in-stead of using the present pharmacological approaches, which target the electrophysiological properties of the substrate.
Limitations of Our Findings
In the present study, conduction velocity was measured in the right atrial free wall, while cells were isolated from the inferoposterior left atrial region. It is possible that conduction velocity measurements obtained from the right atrium are different from those elsewhere in the atria and cannot be related to changes observed in INa in cells obtained from other portions of the atria. However, this is unlikely, since conduction velocity measured in the right atrial free wall is highly correlated with overall conduction time and conduction velocity in Bachmann’s bundle under similar experimental settings.11
Voltage control can be a problem when studying INa. Great care was therefore taken to minimize voltage drop across the Rs by using small cells and pipettes with low resistance in order to clamp the membrane effectively. Cells that had a >5-mV voltage drop across the Rs were rejected. A significant number of cells obtained under control conditions (P0) had to be discarded because of excessive current amplitude and associated series voltage errors, whereas this was not the case for cells obtained from paced dogs. Since cells with larger currents tend to have larger voltage drops across the Rs, the rejection of cells with large series voltage errors would tend to result in an underestimation of INa in P0 dogs. If anything, this error would have reduced the difference between control and paced dogs and is therefore unlikely to have contributed to the differences we observed in INa between these groups.
Selected Abbreviations and Acronyms
|ERP||=||effective refractory period|
|I Ca||=||Ca2+ current|
|I Na||=||Na+ current|
|I to||=||transient outward current|
|P0||=||instrumentation without pacemaker activation|
|P7, P42||=||pacing for 7 and 42 days|
This study was supported by grants from the Medical Research Council of Canada, the Quebec Heart Foundation, and the Fonds de Recherche de l’Institut de Cardiologie de Montréal. Dr Gaspo is a research fellow from the Medical Research Council of Canada. Dr Bosch is a holder of a fellowship from the Deutsche Forschungsgemeinschaft. The authors wish to thank Emma De Blasio, Mirie Levi, and Nathalie Talbot for their skilled technical assistance and Carolyn Gillis and Caroll Boyer for secretarial assistance with the manuscript.
- Received March 21, 1997.
- Accepted September 5, 1997.
- © 1997 American Heart Association, Inc.
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