Altered Excitation-Contraction Coupling in Human Chronic Atrial Fibrillation

Abstract

This review focuses on the (mal)adaptive processes in atrial excitation-contraction coupling occurring in patients with chronic atrial fibrillation. Cellular remodeling includes shortening of the atrial action potential duration and effective refractory period, depressed intracellular Ca²⁺ transient, and reduced myocyte contractility. Here we summarize the current knowledge of the ionic bases underlying these changes. Understanding the molecular mechanisms of excitation-contraction-coupling remodeling in the fibrillating human atria is important to identify new potential targets for AF therapy.

Introduction

Atrial fibrillation (AF) is the most prevalent cardiac arrhythmia found in the clinical practice, affecting >2 million people in the United States alone.[1] AF is often associated with other cardiovascular disorders, such as coronary artery disease, valve dysfunction, congestive heart failure (CHF), and is characterized by significant morbidity. A key determinant of this morbidity is embolic stroke,[2] with loss of atrial contractility being one of the major causes of thrombus formation. AF is characterized by a rapid and irregular heartbeat caused when the atria quiver (fibrillate) erratically, sometimes faster than 200 times per minute.[2]

Several studies have investigated the molecular and ionic mechanisms involved in the remodeling occurring in the atria of patients with AF, and suggest that structural, electrophysiological, and contractile remodeling are critical factors in the disease progression, i.e., they contribute to the development of a substrate that facilitates the tendency for persistence of AF.[3,4] Structural remodeling involves changes in atrial myocyte and tissue morphology (e.g., cell hypertrophy, fibrosis).[3,5,6] Electrical remodeling includes changes in Ca²⁺ and K⁺ currents leading to shortening of the action potential (AP) duration (APD) and loss of APD rate-dependent adaptation.[6] A growing body of experimental evidence points to perturbations in intracellular Ca²⁺ handling as important players in AF-induced atrial remodeling,[7,8] with intracellular Ca²⁺ transients (CaTs) being reduced. Myofilament protein changes in AF are also likely to contribute to atrial contractile dysfunction.[9] However, the mechanisms leading to self-perpetuation of the arrhythmia and depressed cardiac contractility are yet poorly understood. Recently, Llach et al. have studied the basis of irregular beat-to-beat response of human atrial myocytes when subjected to elevations of the beating frequency (which often precedes cardiac arrhythmias) and suggested that stability or instability of the response was determined by the sarcoplasmic reticulum (SR) and L-type Ca²⁺ channel activities.[10]

In this review, we present the current knowledge about the changes occurring in excitation-contraction (E-C) coupling that characterize the remodeled human atrial myocytes from patients with chronic AF (cAF), and the postulated underlying ionic mechanisms.

Phenotypic Consequences of AF on AP, CaT, and Contractility

Myocytes from cAF patients are characterized by shorter APs (Fig. 1A) and effective refractory period (ERP), and loss of rate adaptation of both atrial repolarization (Fig. 1A) and refractoriness.[11-15] Typically, the human atrial AP duration at 90% repolarization (APD90) shortens when paced at faster frequencies, but in myocytes isolated from cAF patients this shortening is severely attenuated (Fig. 1A). CaT amplitude is strongly depressed in myocytes from cAF patients compared to those from subjects in sinus rhythm (Fig. 1B),[16] although the SR Ca2+ content is unaltered.[16-19] CaTs decay more slowly in cAF compared to sinus rhythm.[16,18] Elevated diastolic [Ca²⁺]_i has been reported and attributed to enhanced leak of Ca²⁺ from the SR.[17] Intracellular [Ca²⁺] measurements with aequorin light signals in atrial tissue from patients in sinus rhythm display a positive dependency of CaT amplitude on the pacing rate.[20] Our recently published mathematical model of the human atrial AP and CaT recapitulated this positive rate-dependence, and importantly showed that this is impaired when simulating cAF conditions.[16]

Figure 1. Altered E-C coupling in human AF. A) APs recorded at different pacing rates in a control human atrial myocyte (left) and in a cell from a cAF patient (right) (11). B) Intracellular Ca²⁺ transients measured in human atrial cells from sinus rhythm (left) and cAF (right) patients at physiological temperature. C) Twitch force measurements (22) in myocytes from sinus rhythm and cAF patients are shown at various doses of AVE0118 (I_Kur blocker).

Our simulations indicated that APD rate adaptation in sinus rhythm atrial cells involves accumulation of intracellular Na⁺ ([Na⁺]_i) at high frequencies, which causes outward shifts in Na⁺/Ca²⁺ exchange and Na⁺/K⁺ pump currents. The model also predicted that E-C coupling remodeling in cAF would reduce Na⁺ accumulation, thus causing a blunted APD rate-dependent response.[16]

Baseline force of contraction of atrial trabeculae is also reduced in human cAF by approximately 70% compared to patients in sinus rhythm (Fig. 1C).[21-23]

Ionic Bases of Altered E-C Coupling in AF

The molecular bases of AF-induced alterations in E-C coupling are summarized in Table 1 and discussed in detail in the following paragraphs. E-C coupling remodeling can occur at the level of ion channels/transporters expression, or by modification of ion channel/transporter properties (for example, trafficking or phosphorylation). Furthermore, alterations of myofilament proteins may be involved in AF-induced hypocontractility.

Table 1. Molecular bases of altered E-C Coupling in human AF (changes vs. sinus rhythm) [Modified from [16]].

Cell Size/Structure
Size	Increased length and width (17)
Membrane capacitance	Increased (24)
Sarcolemmal Ion Channels
INa	No changes (15, 35) Steady-state inactivation shifted right (15) Slightly reduced current density (36) Late current increased (36)
ICaL	Reduced current density by ~50% (5, 11, 12, 18, 40) No changes in voltage dependence of activation and inactivation (11)
If	Increased mRNA levels (44)
Ito	Reduced density -80% in the RA -45% in the LA (5, 12, 15, 24, 45-47)
IKur	Reduced density -55% in the RA -45% in the LA (5, 24, 45, 47, 49) Unchanged (12, 15, 46)
IKs	Increased 2-fold (24)
IK1	Upregulated +100% (5, 12, 13, 15, 45)
IK,ACh	Increased basal current by receptor-independent, constitutively active component; increased (15) or reduced carbachol-activated current (13, 56, 57)
IKATP	Decreased (60) Increased (61)
Ca+ and Na+ handling
INCX	Upregulated (17, 18, 21, 34)
SERCA	Reduced maximal pump rate (18) and protein expression (34)
PLN	Enhanced PKA and CaMKII phosphorylation (34) Unaltered CaMKII-dependent phosphorylation (17)
RyR	Increased phosphorylation at PKA and CaMKII sites (17, 18, 67) resulting in increased channel open probability (68) and SR Ca2= leak (17, 111)
INKA	Unchanged function (65)
Ankyrin-B	Downregulated (70)
Protein kinases and phosphatases
CaMKII	Increased expression (32) and phosphorylation (17)
PKA	Similar activity in cAF vs. sinus rhythm (34)
PP1, PP2A	Higher activity (34)
Myofilaments	Reduced maximum rate of tension generation and maximum active tension, reduced passive tension, and increase in myofilament Ca+ sensitivity (9) No changes in maximum force and passive force, reduced rate of tension redevelopment (71) Increased phosphorylation of cMyBP-C (9) Decreased phosphorylation of cMyBP-C (34) No changes in cTnI phosphorylation (9, 34)

Figure 2 depicts simulated APs and CaT for sinus rhythm and cAF myocytes (from (16)) and the major ionic currents that are active during the cardiac cycle, and provides a graphical representation of the main changes occurring in the electrophysiological and Ca²⁺ handling processes in human AF.

Figure 2. Ionic bases of altered E-C coupling in AF. A) Simulated time courses of human atrial APs and contributing ionic currents are shown for sinus rhythm and cAF. Currents are listed on the left (with changes in cAF vs. sinus rhythm), and the genes encoding the channels are shown on the right. B) Simulated human atrial Ca²⁺ transients for sinus rhythm and cAF are shown with NCX and NKA currents (with changes in cAF vs. sinus rhythm on the left). C) Schematic representation of a human atrial myocyte illustrating the cAF-induced changes in Ca2+ handling proteins.

Atrial Cell Morphology

Cell capacitance of myocytes from cAF patients is greater than that of myocytes from SR patients, suggesting that AF cells are hypertrophied.[24] In fact, cells from AF patients are both longer and wider than those from patients in sinus rhythm.[17] Cell hypertrophy may contribute to cAF-induced global atrial dilation, along with changes of the extracellular matrix (with fibrosis and glycogen accumulation). Atrial dilation may itself have important consequences on cellular remodeling and alteration in protein composition and function of the atrial myocytes, as discussed later in this review.

It has recently been shown that atrial myocytes from human tissue sections exhibit extensive t-tubule networks.[25] The presence of t-tubules in the human atria (not detected in isolated human atrial myocytes),[26] may play an important role in determining the spatio-temporal properties of the intracellular CaT.[25] Notably, one can speculate that t-tubules could be subject to remodeling and contribute to perturbed E-C coupling in cAF, as suggested in sheep[27] and dog.[28] However, further investigations will be required to confirm this.

Protein Kinases and Phosphatases

Intracellular CaT is dynamically regulated via phosphorylation by protein kinase A (PKA) and Ca/calmodulin-dependent protein kinase II (CaMKII) of key Ca²⁺ handling and regulatory proteins, such as L-type Ca²⁺ channels, ryanodine receptors (RyRs), and phospholamban (PLN).[29,30] In addition, sarcomere proteins and various sarcolemmal ion channels are targets of both PKA and CaMKII.[29,31] The phosphorylation state of target proteins is also controlled by serine/threonine protein-phosphatases that are differentially regulated in distinct cardiomyocyte microdomains. Thus, altered protein kinase and phosphatase activity may importantly contribute to E-C coupling remodeling in AF. Indeed, CaMKII has been found to be more expressed and more phosphorylated in human cAF.[17,32] Similar PKA activity was found in cAF vs. sinus rhythm in goats,[33] but El-Armouche et al. detected a higher total activity of type 1 and type 2A phosphatases in human cAF, causing inhomogeneous changes in protein phosphorylation in different cellular compartments.[34] This may specifically amplify PKA and CaMKII effects on certain targets without having significant effects on others (e.g., higher phosphatase activity/lower phosphorylation in thick vs. thin myofilaments, cell membrane vs. SR).[34] Thus there is growing interest in the potential role of CaMKII and protein phosphatase inhibitors in preventing arrhythmogenic remodeling in cAF.

Sarcolemmal Ion Channels

I_Na

The Na⁺ current (INa) plays a crucial role in cardiac E-C coupling by initiating the AP, and is also a major determinant of the cardiac AP propagation. Bosch et al. reported that INa density and voltage-dependence of activation were not altered in human AF,[15] the steady-state inactivation was shifted to the right,[15] and no changes were detected in mRNA levels of the Na⁺ channel gene SCN5A.[35] In contrast, Sossalla et al. provided recent evidence that expression of Nav1.5 and peak INa density is decreased (slightly) in the atrial myocardium of patients with cAF.[36]

Although it is unclear whether altered fast INa (Fig. 2A, 2nd row) contributes to the electrical remodeling in human AF, it has recently been shown that the late Na+ current component, INaL (inset), is significantly increased in cAF patients.[36] Sossalla et al.[36] proposed that this increase could be due to the increase in neuronal Na+ channel isoforms (Nav1.1 expression is increased), or mediated by CaMKII, which is increased in AF[17,32] and known to regulate INaL,[37] or caused by oxidative stress.[38,39] However, our simulations suggested that an increased INaL does not contribute significantly to repolarization in cAF, where the overall APD₉₀ was still shorter than that in normal healthy cells.[16] On the other hand, an increase in INaL may cause cellular Na+ and Ca²⁺ overload and lead to contractile dysfunction and electrical instability (via reverse-mode Na⁺/Ca²⁺ exchange).[29]

I_CaL

The L-type Ca²⁺ current (I_CaL) critically regulates E-C coupling by triggering SR Ca²⁺ release, and modulating AP shape and duration, i.e., maintaining the AP plateau.[29] Reduction in I_CaL density (-50% vs. sinus rhythm, Fig. 2A, 3rd row) is one of the most consistent electrophysiological features of electrical remodeling in human AF (as seen in (5, 11, 12, 16, 18, 40)).Christ et al.[40] demonstrated that decreased I_CaL density in cAF is not accompanied by altered expression of the corresponding α1c and β2a channel subunits (although other studies found different results),[41] and proposed that lower basal I_CaL is due to decreased channel phosphorylation in AF, which results from an altered ratio of protein kinase/phosphatase activity in favor of increased phosphatase activity. An analogous explanation was proposed for the blunted effect of CaMKII inhibition on ICaL in human cAF.[17] It has been shown that blocking I_CaL with nifedipine in normal human atrial cells results in an AP characteristic typically seen in AF[11] with respect to morphology, duration and impaired rate-dependent adaptation, i.e., reduction in I_CaL seems to be a critical component of the remodeled atrial electrical phenotype. However, Workman et al. found that nifedipine did not significantly alter ERP in sinus rhythm myocytes (although APD was shorter), thus supporting the idea that ICaL downregulation may not be sufficient by itself to explain the remodeled atrial electrical phenotype.[12]

I_CaT

There is no evidence of a T-type Ca²⁺ current (I_CaT) in human atrial myocytes.[42,43]

I_f

The hyperpolarization-activated pacemaker current, If, ion channel has been found to be increased in human AF compared to sinus rhythm, at least at the mRNA level,[44] and could contribute to ectopic atrial pacemaker activity. However, functional evidence for If involvement is lacking at present.

I_to and I_Kur

The Ca²⁺-independent transient outward K⁺ current (I_to) and the ultra-rapid delayed rectifier K⁺ current (I_Kur) dominate the early AP repolarization phase and confer the atrial AP a characteristic triangular shape. Human cAF is associated with strong reduction of Ito (Fig. 2A, 4th row) density[5,12,15,24,45-47] and downregulation of its channel α subunit Kv4.3.[35,48] I_Kur(Fig. 2A, 5th row) was reduced in cAF[24,45,47,49] paralleled by diminished expression of Kv1.5.[35,45,[48] However, others have reported no changes in I_Kur density.[12,15,46] Inconsistent results about IKur function have been commented on previously by Christ et al. and attributed to different strategies for identification of I_Kur (e.g., pharmacological or with Ito-inactivating prepulse), and to a fraction of IKur that is not accounted for by Kv1.5.[49] The reduction in Ito and I_Kur explains the slight prolongation in earlier phases of the AP (Fig. 2A, 1st row).[16,50]

It has been shown that CaMKII (increased in cAF) positively regulates Ito in human atrial myocytes in acute conditions, as the application of the CaMKII inhibitor KN-93 caused loss of channel function.[32] The authors speculated that, by reducing the extent of inactivation of Ito, upregulation of CaMKII during atrial fibrillation reduces Ca²⁺ influx and therefore minimizes Ca⁺ overload. On the other hand, CaMKII overexpression in cAF may impact channel expression, thus contributing to Ito downregulation, as recently shown in CaMKII-overexpressing transgenic mice.[51]

Experimental evidence suggests that block of IKur enhances force of contraction of isolated human atrial trabeculae both in patients in sinus rhythm and AF.[22,23,52] We have recently predicted that block of IKur results in prolongation and elevation of the AP plateau, which augments the CaT amplitude that would elicit a positive inotropic effect.[16] Taken together, these studies suggest that I_Kur might be a potentially useful atrial-specific target to potentially counteract hypocontractility associated with cAF. A slight AP prolongation associated to IKur blockade may also be beneficial.

Caballero et al. have recently looked at differences in current density and AF-induced alterations in the right vs. left human atrium. They found heterogeneity in the repolarizing currents between the atria in sinus rhythm, and demonstrated that cAF reduced the Ito amplitude and density more markedly in the left than in the right atrium, thus creating a right-to-left gradient, whereas IKur was more markedly reduced in the right than in the left atrium, thus dissipating the left-to-right gradient detected in sinus rhythm.[24] However, the data concerning intra-atrial heterogeneities in repolarizing currents in human atrial myocytes are still limited, and it is unclear whether and how these changes may contribute to the perpetuation of arrhythmia.[16]

I_Ks and I_Kr

The delayed rectifier K⁺ currents have proven much harder to record and study in isolated human atrial cells.[53] Nevertheless, their contribution is likely to be small in cells that lack an appreciable plateau phase (e.g., see current densities in Fig. 2A, 6th and 7th rows).[54] The block of the rapidly activating delayed rectifier K⁺ current, I_Kr, has been shown to prolong human atrial APD in the late phase of repolarization by a small amount,[23] and to date no experimental evidence has suggested its involvement in AF-induced electrical remodeling.

Recently, Caballero et al. provided the first demonstration that cAF significantly increased the amplitude of the slow delayed rectifier K⁺ current, I_Ks, in both atria.[24] They suggested that IKs increase could contribute to cAF-induced shortening of APD and to further promote fibrillatory conduction, especially with current accumulation at high frequencies.

I_K1 and I_K,ACh

The inwardly rectifying K⁺ current (I_K1) primarily controls the resting potential of the cardiac cell, and its much lower density in atrial than in ventricular myocytes[55] confers the atrial AP a more depolarized resting potential.[16] In cAF, increases in both current density[5,12,13,45,56] and mRNA levels[5,13] have been reported (Fig. 2A, 8th row). Increased I_K1 causes a more negative resting membrane potential in cAF vs. sinus rhythm human atrial myocytes.[13,16,56]

Patients with chronic AF exhibit agonist-independent constitutive I_K,ACh activity that contributes to the enhanced basal inward rectifier current and may result from abnormal channel phosphorylation by PKC.[13,56,57] Constitutively active I_K,ACh is considered to support the maintenance of AF, together with increased I_K1, by stabilizing reentrant activity sustained by rotors (faster activation, less meander).[58]

Recently, Voigt et al. found significant left-to-right gradients in I_K1 and constitutively active I_K,ACh in patients with paroxysmal AF, which were dissipated in cAF, raising the idea that this may contribute to left-to-right dominant frequency gradients that are often more evident in paroxysmal AF vs. cAF.[56]

I_KATP

The ATP-sensitive K⁺ (I_KATP) channels generate an inward rectifying current that activates with a decrease in intracellular ATP concentration.[59] Gene expression and electrophysiological studies in patients with atrial fibrillation demonstrated reduced mRNA levels of Kir6.248 and current activation,[60] but increased current was also reported.[61] Interestingly, a KATP channel mutation has been shown to confer risk for adrenergic atrial fibrillation originating from the vein of Marshall,[62] and it has been proposed that KATP channel deficit could play a broader role in the pathogenesis of electrical instability.[63] It is also conceivable that metabolic and mechanosensitive gating of KATP channels could be altered with structural heart disease and atrial dilation, thus providing a substrate for the more common acquired form of atrial fibrillation.[63]

Ca²⁺ and Na⁺ Handling

I_NCX

The Na⁺/Ca²⁺ exchanger current (INCX) is the main Ca²⁺ extrusion and Na⁺ influx pathway in cardiac myocytes. It extrudes 1 Ca⁺ in exchange for 3 Na+, thus generating an inward current that influences cardiac repolarization and arrhythmogenesis.[29] Increased expression[18,21,34] and abnormal function of I_NCX protein[16,18] are implicated in human AF pathophysiology. An increase in I_NCX may be an adaptive response to cellular Ca²⁺ loading and contribute to diminish the Ca²⁺ overload induced by rapid atrial pacing (along with ICa downregulation). Indeed, the decay rate of caffeine-evoked CaT (attributable to Ca²⁺ removal by NCX) is shown to be faster in human cAF vs. sinus rhythm myocytes.[16-18] Note that simulated I_NCX during an AP is smaller in AF than in sinus rhythm (Fig. 2B, 2nd row), due to the reduced CaT (Fig. 2B, 1st row). Na⁺ overload-induced Ca²⁺ influx via reverse-mode NCX has been implicated in Ca²⁺ overload and related arrhythmogenesis, whereas increase Ca²⁺ extrusion via forward-mode has been linked to delayed-afterdepolarizations.[29,64] Indeed, Na⁺ and Ca²⁺ loading are more favored at increased atrial rates (with AF). However, more studies are needed to assess whether delayed afterdepolarizations (DADs) are important in initiating arrhythmias in AF, and the underlying role of NCX in mediating them, since an increased IK1in cAF will tend to oppose the occurrence of such DADs. These studies will help determine if blocking NCX represents a novel therapeutic strategy in suppressing arrhythmia triggers in cAF.

I_NKA

The Na⁺/K⁺ pump (NKA) is the main route of Na+ efflux in cardiac cells thus regulating intracellular [Na⁺]. By extruding 3 Na⁺ in exchange for 2 K⁺ it generates an outward current that is known to influence resting membrane popotential and repolarization.[29] Workman et al. found no difference in NKA pump current in myocytes from cAF patients compared to sinus rhythm, and concluded that INKA is not involved in AF-induced electrophysiological remodeling in patients.[65] Our simulations show different NKA current underlying the AP (Fig. 2B, 3rd row) because of altered Na+ loading in cAF. Intracellular [Na⁺] changes may contribute to the human cAF phenotype, as we postulated in our modeling study[16] but have not yet measured.

Ryanodine Receptors

RyRs directly control SR Ca²⁺ release in cardiac muscles, activating contraction during E-C coupling.[29] Spontaneous Ca^2+-release events (Ca²⁺ sparks) and Ca²⁺ waves through leaky RyR channels have been reported in myocytes from cAF patients[17,18,66,67] despite unaltered SR Ca²⁺ content. One potential contributor to RyR hyperactivity may be oxidative stress, which is known to play a critical role in AF pathophysiology[38] and increase RyR open probability. Neef et al. suggested that the CaMKII-dependent increase in SR Ca²⁺ leak caused by RyR hyperphosphorylation in AF is a potential arrhythmogenic mechanism,[17] because elimination of Ca²⁺ via inward INCX could lead to cell depolarization and cause DADs. Voigt et al. measured directly single RyRs isolated from cAF patients and demonstrated a higher channel open probability in cAF that responded to CaMKII inhibition.[68] Thus CaMKII inhibition may reduce the propensity for atrial arrhythmias.

SR Ca²⁺ ATP-ase and PLN

The SR Ca²⁺ ATP-ase (SERCA) is responsible for pumping Ca²⁺ back into the SR after Ca²⁺ release.[29] The endogenous inhibitor PLN regulates SERCA and releases its inhibition when phosphorylated by either PKA or CaMKII.[29,30] A decrease in SERCA activity, associated with smaller SERCA protein expression,[18,34] is evident in human cAF and explains the slower CaT decay compared to sinus rhythm.[16,18,34] On the other hand, reduced inhibition of SERCA by hyperphosphorylated PLN[34] in cAF could help to maintain a normal SR Ca²⁺ load despite increased RyR activity.

Ankyrin-B

Ankyrin-B (encoded by ANK2) is an adaptor protein expressed in excitable cells that targets ion channels (e.g., Na+ and Ca²⁺channels), transporters (e.g., NKA and NCX), and signaling molecules to specific membrane domains. In the heart, ankyrin-B loss-of-function mutations in humans lead to Long QT syndrome, AF, sinus node dysfunction and stress-induced ventricular arrhythmias.[69] Recently, reduced ankyrin-B expression has been demonstrated in atrial samples of patients with paroxysmal AF, and supported an association between ankyrin-B and AF.[70] A new potential molecular mechanism underlying ankyrin-associated AF has been proposed involving disrupted Cav1.3 (atrial L-type Ca²⁺ channels) membrane targeting in atrial myocytes.[70] It will be interesting to further explore the role of ankyrin in cAF.

Myofilaments

Altered Ca²⁺ handling (namely, downregulation of the L-type Ca²⁺ channels and increased Ca²⁺ extrusion via NCX) could account for the depressed contractility in remodeled atria, but a reduction of the maximum force generating capacity of the myofilaments and its Ca^2+-sensitivity may also be involved. Indeed, recent studies have highlighted the potential role of sarcomeric proteins in the cAF induced hypocontractility,[9,34,71] although results are somewhat controversial. Compared to sinus rhythm myofibrils, cAF myofibrils exhibited reduced maximum rate of tension generation and maximum active tension, reduced passive tension, and increased in myofilament Ca²⁺ sensitivity.[9] An earlier study did not show significant changes in maximum force and passive force, but did report reduced rate of tension redevelopment in cAF.[71] One major difference between the two studies is that the former used left atrial samples whereas the latter used right atrial samples.

Altered phosphorylation state of various myofilament proteins was found in cAF vs. sinus rhythm. Phosphorylation of the primary sarcomere target of PKA, cTnI, was not altered in cAF atria.[9,34] The expression of the slow β-myosin heavy chain isoform (cMyBP-C)[9,34,71] was upregulated in cAF, and its phosphorylation levels were found significantly increased[9] or decreased.[34] It has been suggested that discrepancies between these results may be explained by a decrease in cMyBP-C phosphorylation in cAF reflecting atrial dilatation rather than being a component of cAF.[9] Another potential reason is the use of samples from the left atrium in the former study and from the right atrium in the latter.

Further studies are needed to resolve these inconsistencies. Furthermore, cell shortening data that are currently missing in human atrial myocytes may help in linking these molecular changes to functional alterations.

It is becoming increasingly clear that studies of remodeling of human atrium by chronic AF are frequently and unavoidably influenced in part by multiple confounding clinical variables such as patient age, sex, disease history, and drug treatments. Furthermore, the changes in ion currents and APs should be considered to be associated with, rather than necessarily caused by, the chronic AF. Nevertheless, the concordance between these human chronic AF data and AF/atrial tachypacing-induced changes in animal models (see Table 1 in([72]))supports the view that chronic AF causes atrial electrophysiological remodeling in humans.

Consequences of Ventricular Dysfunction and Atrial Dilation on Human Atrial AP

Structural, electrophysiological, mechanical, metabolic and neurohumoral remodeling associated with cardiac disorders, such as coronary artery disease, CHF, and left ventricular systolic dysfunction (LVSD) may increase AF risk.[73] In atrial cells isolated from patients with CHF or LVSD, AP duration was either unchanged (Fig. 3A)[74,75] or increased (Fig. 3B).[76] In patients in sinus rhythm with reduced left ventricular ejection fraction (< 45%) APD₉₀ was shorter than in patients with higher ejection fraction,[75] and there was a significant correlation between cellular ERP shortening and decreasing left ventricular ejection fraction.[75] Furthermore, multivariate analysis adjusting for 10 relevant clinical covariates confirmed that LVSD was independently associated with atrial cellular ERP-shortening, which may, therefore, be expected to contribute to a predisposition to AF in these patients. The features of ionic remodeling in CHF or LVSD in human atrium are not fully understood. I_CaL was either decreased in patients with coronary artery disease, aortic valve disease, or mitral valve disease[77] or unchanged in LVSD or CHF patients.[75,78] Schreieck et al. found increased Ito in human atrial myocytes of patients with reduced left ventricular function, with no change in its voltage dependence or decay, but with enhanced reactivation.[74] However, this Ito increase may have been confounded by the lower proportion of patients treated with β-blockers in the reduced LV function group, since such treatment is associated with decreased I_to in human atrium.[79] In contrast, Workman et al. found that LVSD was associated with decreased I_to, a positive shift in its activation voltage, and no change in its decay kinetics.[75] Koumi et al. reported low resting membrane potential in atrial myocytes from CHF patients, possibly due to reduced density of I_K1 and I_K,ACh.[76] Workman et al. reported unchanged I_K1 in LVSD,[75] although Ba^2+-sensitive I_K1 or I_K,ACh were not measured. Unchanged atrial I_Kur has also been reported in human LVSD.[74,75]

Figure 3. Altered atrial APs in human ventricular dysfunction. A) Atrial APs from patients with moderate or severe LVSD (bottom) and from patients without LVSD (top) (75). B) Representative AP and response to ACh in isolated atrial myocytes from HF (top) and donor (bottom) hearts at a stimulation frequency of 1 Hz (76). C) Representative APs from a non-dilated (normal, left) and a dilated (right) atrium recorded at a pacing frequency of 1 Hz (80).

Cardiac dilatation is known to develop frequently during the course of cardiac failure.[80] In trabeculae and myocytes taken from dilated atria the AP was shorter and the plateau was markedly depressed (Fig. 3C) compared to trabeculae and myocytes from non-dilated atria.[80] However, it must be noted that the ventricular dysfunction was not quantified in these patients. AP changes were explained with more severely depressed I_CaL compared to the reduction in total outward current.[80]

Overall, the ionic bases of altered atrial function in patients with ventricular dysfunction, and how it predisposes to more frequent AF episodes culminating in cAF, remains poorly understood.

Autonomic Changes in Chronic AF and Related Myocardial Diseases

The autonomic nervous system, and particularly the relative activities of the sympathetic (adrenergic) and parasympathetic (cholinergic) branches, have a major influence on the occurrence of AF. Furthermore, chronic AF, and certain predisposing cardiac pathologies, remodel atrial electrophysiological responses to catecholamines and acetylcholine and thus influence the electrophysiological mechanisms of AF. β-adrenergic stimulation increases human atrial I_CaL,[11,41,42,81] I_Kur[79,82] and If,[83,84] has no effect on I_K1,[79] I_KA,Ch[85] or I_to,[79] and has markedly different effects on connexin conductance or expression, depending upon their main molecular correlate; i.e. Cx,[40] Cx,[43] Cx45.[86] The increased I_CaL and I_Kur, with lack of effect on other repolarizing currents, results in no net effect of β-stimulation on atrial APD,[90] as predicted by our model,[16] consistent with 5 of 6 reports in human atrial cells or tissues.[87] However, the increased I_CaL markedly elevates the AP plateau[81] and, coupled with increased [Ca²⁺]i from PLN phosphorylation by adrenergic stimulation,[88] favors non-reentrant activity such as afterdepolarizations.[87]

Human atrial studies of α-adrenergic stimulation are sparse: phenylephrine inhibited I_K1,[85,89] I_K,ACh[85] and IKur[82] also potentially promoting afterdepolarizations. Chronic AF consistently potentiates the effect of β-adrenergic stimulation to increase human atrial I_CaL.[11,40,43,90] While this could, in theory, increase the propensity for afterdepolarizations in the presence of catecholamines, chronic AF also markedly decreases basal I_CaL[6] and attenuates the effects of α-stimulation on I_K1 and I_K,ACh.[85] Chronic AF may also cause increased atrial adrenergic innervation, “neural remodeling”, in patients.[91] The effects of chronic AF on [Ca^2+-]_iresponses to adrenergic stimulation have yet to be studied in human atrium. Data on effects of myocardial diseases that predispose to AF, on human atrial adrenergic responses, are equivocal: the ability of β-stimulation to increase I_CaL was attenuated,[92,93] unchanged[75] or potentiated[77] in association with HF or LVSD. An attenuated I_CaL-increase was also reported in cells obtained from dilated atria from explanted hearts.[78] Attenuated β-responses may involve reduced β-receptor density or function.[87] Post-operative AF was not predicted by any change in the pre-operative atrial I_CaL response to β-stimulation.[94] Cholinergic elevation and increased levels of acetylcholine activate I_K,ACh and also antagonize effects of catecholamines on I_CaL, both shortening APD and ERP, thus promoting reentry.[95] Also, combined adrenergic/cholinergic-stimulation may produce “late phase EADs”,[96] possibly by concurrently shortening APD and increasing [Ca²⁺].[97] Chronic AF induces a constitutively active I_K,ACh in human atrium,[57] likely resulting from a PKC isoform switch.[98] However, the acetylcholine-mediated increase in atrial I_K,ACh[13,57] and shortening in atrial APD₁₃ were each attenuated in chronic AF, and the cholinergic receptors GIRK[1] and GIRK[4] were generally downregulated.[5] The attenuation of atrial cholinergic responses by chronic AF may be restricted to the right atrium.[56] The ability of acetylcholine to increase I_K,ACh and/or shorten atrial APD may be attenuated by HF, as shown in dogs.[99] However, corroborative data from human patients with HF or LVSD are sparse and confounded by the presence of chronic AF.[76] While much progress has been made, the complex and interacting influences of chronic AF and its predisposing myocardial pathologies on the involvement of the autonomic system in AF are yet to be resolved.

Oxidative Stress and Inflammation-related Changes in Human AF

Patients undergoing cardiac surgery often experience post-operative AF. It has been shown that these patients did not exhibit the electrophysiological remodeling seen in patients with cAF as far as Ca²⁺ and K⁺ currents and AP characteristics are concerned,[94] whereas altered atrial Ca²⁺ handling in post-operative AF patients has not yet been studied. However, Van Wagoner et al. showed that patients with the highest I_CaL density pre-surgery, were associated with post-operative AF, thus indirectly suggesting a role for Ca²⁺overload, mediated via oxidative/inflammatory stress, as a possible trigger.[11] This is because increased levels of inflammatory markers are often recorded after cardiac surgery, and recent evidence suggests oxidative stress may play an important role in the pathogenesis and perpetuation of post-operative AF.[100]

Several studies have shown increased myocardial oxidative stress associated with AF.[38,101] In addition, inflammatory markers such as interleukin-6 and C-reactive protein have been found elevated in AF patients.[101,102] Evidence suggests that in several pathophysiological conditions inflammation and oxidative stress are highly interrelated, whereby inflammation augments oxidative stress and viceversa, and may be involved in AF pathogenesis. Importantly, oxidant and inflammatory mechanisms may contribute to the described structural, electrophysiological, and contractile remodeling that favors maintenance of AF,[103] and thus could be considered as targets for AF-treatment, as discussed in more comprehensive reviews on the topic.[101,104] Inflammatory processes may contribute to atrial injury resulting in myocyte hypertrophy and fibrosis. Furthermore, several Ca²⁺ channels and transporters are the subject of redox modulation.[105] For example, oxidative stress may play an important role in I_Ca changes, as it has been shown that S-nitrosylation of the L-type Ca²⁺ channel α subunit is increased in AF, and exogenously applied glutathione partially restores the AF-related I_Ca reduction.[106] We also discussed above the potential contribution of oxidation in AF-associated RyR hyperactivity. Several K⁺ channels (e.g., I_to[107] and I_KATP) are also sensitive to redox state. Kv1.5 currents are inhibited by oxidation by S-nitrosylation,[108] which may contribute to IKur suppression in AF. Redox-dependent modulation of Na+ channel activity has also been reported.[109] Additionally, oxidative stress may affect myofilament protein function[38] and influence the activity of protein kinases (e.g., CaMKII[110]) and phosphatases that alter E-C coupling via phosphorylation of target proteins and are also redox sensitive. Suppressing E-C coupling remodeling with anti-inflammatory and antioxidant drugs (such as glucocorticoids and statins) has proven clinically useful in some cases in preventing AF recurrence.[104] In sum, we are still limited in our understanding regarding how oxidative/inflammatory stress influences E-C coupling, particularly in the context of chronic AF, and further studies are warranted.

Conclusions

Chronic AF is associated with altered expression and activity of numerous sarcolemmal ion channels, transporters, Ca²⁺ handling and myofilament proteins. Understanding the ionic mechanisms underlying E-C coupling remodeling in fibrillating human atria, and distinguishing compensatory responses from maladaptive mechanisms, may allow for identification of new therapeutic targets to improve electrical and contractile function in cAF patients.

Acknowledgements

Supported by NHLBI Grants P01HL039707, P01HL087226, and the Leducq Foundation (to Dr. José Jalife), NHLBI Grants P01-HL080101, R37-HL30077-29 and the Leducq Foundation (to Dr. Donald Bers). We would like to thank Drs. José Jalife, Donald Bers, and Dobromir Dobrev for useful feedback and discussions.