Effects of Electrical Remodeling on Atrial Fibrillation in Diabetes Mellitus

Atrial fibrillation (AF) is one of the most common arrhythmias in medical practice. Diabetes mellitus (DM) is one of the independent risk factors for atrial fibrillation. The increased morbility of atrial fibrillation in diabetes mellitus is related to both structural and electrical remodeling of atrium. Based on studies of atrial electrophysiological changes in diabetes mellitus, this article focuses on the electrical remodeling of atrial cardiomyocytes, including remodeling of sodium channels, calcium channels, potassium channels and other channels, to provide the basis for the clinical management of antiarrhythmic drugs in diabetic patients with atrial fibrillation.


Introduction
Atrial fibrillation (AF) is one of the most common arrhythmias in medical practice worldwide [1].Traditionally, AF can be classified into five patterns: first diagnosed, paroxysmal, persistent, long-standing persistent and permanent AF [2].AF patients have an increased risk of congestive heart failure and stroke, resulting in severe disability and death [3].Diabetes mellitus (DM) is one of major hazard factors for AF [4][5][6].Subclinical AF episodes occur frequently in type 2 diabetes mellitus (T2DM) patients and are associated with increased thromboembolic risk [7].Moreover, treatment of AF appears to be more challenging in patients with DM.Outcomes of AF ablation are worse in patients with DM compared to the general population and arrhythmia recurrence is significantly higher in the DM group compared to the non-DM group [8,9].Both AF and DM are currently prominent global public health issues [10].However, the underlying mechanisms of AF in DM have not been completely investigated.
DM predisposes to AF due to several factors, such as atrial remodeling, autonomic system dysfunction [11] and epigenetic regulation [12].Atrial remodeling includes structural remodeling and electrical remodeling.Atrial structural remodeling was found in both type 1 and type 2 DM animal models [5,13].There are evidences demonstrating that DM is associated with disordered arrangement and higher cross-sectional areas of atrial cardiomyocytes, as well as increased interstitial fibrosis and inflammation [5].Ultrastructural studies of DM cardiomyocytes also showed irregularly arranged myofibrils, degenerated Z-lines, and swollen, vacuolated mitochondria with fragmentation [14].In addition to structural remodeling, in-vestigators have studied the ionic mechanisms that underlie the electrical remodeling of AF in DM [15].This article reviews the remodeling of ion channels in atrial myocytes with DM and their related mechanisms, so as to provide the basis for the clinical treatment of antiarrhythmic drugs used in patients with diabetic AF.

AF Incidence and Alterations of Atrial Electrophysiological Characteristics in Diabetes
In previous studies, diabetic animals have shown a high susceptibility for induced AF, with a significantly higher incidence of AF and a longer AF duration after atrial burst stimulation [4][5][6][16][17][18][19][20][21][22].The electrocardiogram and electrophysiological parameters of diabetic animals have been reported in humans [23] and in different animal models [24].Diabetic animals often show irregularities in atrial depolarization as P-wave prolongation and increased P-wave dispersion, leading to impulse generation or conduction abnormalities [24].Lower heart rate (HR), prolonged rhythm-to-rhythm (RR) interval and similar Q wave-R wave-S wave (QRS) duration, onset of wave Q to the end of wave T (QT interval), QTc interval (corrected QTc interval) were observed in type 1 diabetic Sprague Dawley (SD) rats induced by streptozotocin (STZ) (6 weeks after treatment) [4].No significant differences were found in sinus cardiac length (SCL), left atrial (LA) effective refractory period (LA-ERP), right atrial (RA) ERP (RA-ERP), inter atrial conduction time (IACT), RA-ventricular conduction time (RA-VCT) and LA-ventricular conduction time (LA-VCT) between control and diabetic groups.In contrast, the conduction velocity of atria was slower and Genetically T2DM 20-week-old ZDF rats SCL↑, sinus node recovery time↑ and corrected sinus node recovery time↑ [18] Goto-Kakizaki rats HR↑, irregular P waves, separation of P and QRS waves [25] db/db diabetic mice (16 and 20 weeks old) AERP↑, duration of P-wave↑, interval of PR↑, and interval of RR↑ [13] T1DM, type 1 diabetes mellitus; T2DM, type 2 diabetes mellitus; STZ, streptozotocin; SD, Sprague Dawley; HFD, high fat diet; HR, heart rate; IACT, inter atrial conduction time; LA-ERP, left atrial-effective refractory period; RA-ERP, right atrial-effective refractory period; LA-CV, left atrial conduction velocity; SCL, sinus cardiac length; AERP, atrial effective refractory period.
conduction in homogeneity was notably increased in diabetic rats with a higher incidence of AF.In STZ-induced diabetic Wistar rats (8 weeks after treatment), IACT was longer, LA-ERP and RA-ERP were shorter than control rats [5].However, no significant differences were observed in atrioventricular Wenckebach cycle length (AV-WCL) and HR.The incidence of AF was also increased in a type 2 DM (T2DM) animal model [16].In high fat diet (HFD) and low dose STZ treated SD male rats, LA conduction velocity was significantly lower as shown by mapping images.
The IACT was longer and SCL, AV-WCL, RA-ERP and LA-ERP were not statistically different [17].In 20-weekold Zucker Diabetic Fatty (ZDF) rats, SCL, the time of sinus node recovery and corrected sinus node recovery were noteworthy longer than those in the control group [18].The electrocardiogram (ECG) of T2DM model Goto-Kakizaki (GK) rats showed accelerated HR, irregular P waves, separation of QRS and P waves, and partial blockade of electrical conduction [25].Atrial effective refractory period (AERP), duration of P-wave, and the time from the onset of the P wave until the R wave (PR interval) and RR interval were longer in db/db type 2 diabetic mice (16 and 20 weeks of age) [13].In summary, abnormal electrophysiological parameters were observed in both type 1 and type 2 DM models, including chemically induced type 1 DM, metabolic type 2 DM with mild to medium pancreatic injury followed by diet induced insulin resistance , and genetically hyperglycemic animals (Table 1, Ref. [4,5,13,[16][17][18]25]).

Ion Channel Remodeling of Atrial Myocytes in Diabetes
The abnormal electrophysiological parameters in diabetes are closely related to the changes of action potentials (AP) in atrial myocytes.AP duration (APD) occurs in diabetic animals with higher APD 50 and APD 90 [18,25].Prolonged APDs result from up-or down-expression and activation or inactivation of different kinds of ion channels that form depolarizing and repolarizing currents, such as sodium channels, calcium channels, potassium channels, late sodium channels and other channels.

Sodium Channels
The voltage-gated sodium channel current (I Na ) is widely found in the atrium and is one of the most important depolarization cation channels in the cardiomyocyte membrane [26].It is the major determinant of the upstroke of AP.Proteins of Na V 1.5 (encoded by SCN5A gene) are responsible for I Na [27].In Akita type 1 diabetic mice, I Na was measured both in isolated RA and LA myocytes [21].Current-voltage (I-V) curves of atrial myocytes demonstrated that I Na was reduced.The decreased I Na density of diabetic mice occurred in association with a decline of maximum conductance (Gmax) and a mode straight shift of the voltage dependence of activation.Voltage dependence of inactivation was not altered in atrial myocytes of Akita mice.The expression of SCN5A mRNA and Na V 1.5 proteins were reduced in the atrium of Akita mice compared with normal controls.The alterations of I Na resulted in prolonged P-wave duration, and reduced atrial conduction velocity in Akita mice.Acute insulin treatment increased I Na due to enhanced insulin signaling through activation of phosphatidylinositol 3,4,5-triphosphate (PIP3).
In atrial myocytes of db/db mice [13], I Na amplitude, I Na steady-state activation, or fast and slow time constants of I Na activation were similar to control mice.The steadystate inactivation curve was shifted to the right, which suggested a larger window current.SCN5A mRNA and Na V 1.5 protein levels was similar in db/db atrium compared with control.In metabolic type 2 DM with STZ-injection fol-lowed by diet induced insulin resistance, the density of I Na were similar in the control and T2DM rat myocytes [24].
Sodium channels are activated during the depolarization phase and then rapidly deactivated.However, some channels reopen as a late or persistent sodium current (I Na-L ) that participate in repolarization [28].The basal I Na-L is mainly generated from the Na V 1.5 isoform and is regulated by calmodulin-dependent kinase II [29].Several studies show that the increase of I Na-L can markely prolong the duration of AP in cardiomyocytes and is important in the development of AF [30][31][32].I Na-L was increased in isolated atrial myocytes of DM mice compared to controls [33].In diabetic mice, the application of the I Na-L inhibitor (GS967) inhibited I Na-L , shortened APD, and reduced the incidence of AF by high-frequency electrical stimuli.A recent study in knock-in mice fed a high fat diet, which ablates phosphorylation of the Na V 1.5 channel and prevents augmentation of I Na-L , increased AF inducibility [31].In conclusion, the increased susceptibility to AF in diabetic mice was associated with increased I Na-L and the subsequent prolongation of AP.

Calcium Channels
The voltage gated calcium channel is another important cation influx channel.The L-type calcium current (I CaL ) contributes to a depolarizing current which is actived during the repolarization phase of AP.It is responsible for the maintenance of the platform stage.Proteins of Ca V 1.1~1.4 and Ca V 3.1~3.3are responsible for I CaL and T-type Ca 2+ currents, respectively.
In atrial myocytes of STZ-induced diabetes, the maximum current density of I CaL was significantly higher compared with control.The steady-state I CaL activation curve was shifted to the left and the activation slope factor was decreased, while the inactivation curve was shifted to the right and the inactivation slope factor was higher in the diabetic group [6].These results suggested the I CaL was easily activated and was difficult to be inactivated in DM.Ca V 1.2 protein expression was also increased in the diabetic atrium.Selective inhibition of protein kinase C (PKC)-β using ruboxistaurin (RBX) can reduce nuclear factor kappa-B (NF-κB)/transforming growth factor-β (TGF-β)/Ca V 1.2 expression and I CaL activation, and inhibit abnormal atrial remodeling in diabetic rats.In ZDF rats, the protein expression level of Ca V 1.2 in the atrium and current density of I CaL were significantly lower in the atrial myocytes, while the kinetics of I CaL were similar to the control group [18].In the atrium of metabolic type2 diabetic rats, Ca V 1.2 mRNA and protein expression were significantly decreased, whereas the level of Ca V 3.1 was upregulated [14].I CaL was reduced and the T-type Ca 2+ current was increased in diabetic atrial myocytes.Long term rosuvastatin treatment alleviated these pathological changes in diabetic rats.The results of studies involving I CaL have not been consistent, and may be related to the use of different animal models and the du-ration of diabetes.

Potassium Channels
There are several types of potassium channels in cardiomyocytes.It has been reported that the main repolarizing potassium currents (I K ) are transient outward potassium currents (I to ), rapid-delayed rectifier potassium currents (I Kr ), slow-delayed rectifier potassium currents (I Ks ) and steady-state potassium currents (I ss ) in the human heart ventricle, while they are fast transient-outward potassium currents (I to, f ), ultra-rapid delayed rectifier potassium current (I Kur ) and I Ks currents in the atrium.I to and I Kur participate in the phase 1 repolarization process of myocardial AP, and I Kr , I Ks participate in the phase 2 and phase 3 repolarization process of AP.Proteins of K V 4.2 (encoded by KCND2) and K V 4.3 (encoded by KCND3) are responsible for I to , and K V 1.5 proteins (encoded by KCNA5) are responsible for I Kur [34].
Bohne et al. [13] found atrial I K , mainly including I to and the I Kur , were decreased in atrial myocytes of db/db mice.The decrease of I to occurred in association with reductions in the expression of KCND2 mRNA and K V 4.2 proteins (mRNAs for KCND3 were reduced and K V 4.3 proteins were similar).The reduction in I Kur was not related to mRNA or protein expression (no differences in KCNA5 mRNA or K V 1.5 protein levels).There were no differences in calcium-activated potassium currents in atrial myocytes of db/db mice.Atrial current density of I to and I Kur in ZDF diabetic rats was less than that in controls and the expression levels of the protein K V 4.3 and K V 1.5 were significantly downregulated [18].No significant differences were found in the kinetics of I to .Polina et al. [21] also found I K carried by K V 1.5 channels were reduced in type 1 diabetic Akita mice.They measured I K in atrial myocytes with and without a prepulse to inactivate I to .Peak total I K was reduced in diabetic atrial myocytes while I to (the difference currents between measurements with and without the inactivating prepulse) were similar between wildtype and diabetic mice.The I Kur , as measured by 4-aminopyridine sensitive I K , was reduced, and western blot showed no differences in K V 4.2 and K V 4.3 protein levels of the atrium from wild-type and diabetic mice; however K V 1.5 protein was reduced with no difference in mRNA expression.Inward rectifier K + currents (I k1 ) mainly affected resting membrane potential.No significant difference in I k1 densities were found between control and diabetic atrial myocytes [13,24].
There are numerous studies showing that small conductance calcium-activated potassium channels (SK channels) play important roles in diabetic AF.The SK channels have three isoforms including SK1 (K Ca 2.1, encoded by KCNN1), SK2 (K Ca 2.2, encoded by KCNN2) and SK3 (K Ca 2.3, encoded by KCNN3).The SK currents were significantly reduced and the AP duration was prolonged in atrial myocytes of GK rats [25].Compared with control rats, the expression of SK2 channel was decreased and the expression of the SK3 channel was increased in atrial myocytes of GK rats.Metformin reversed SK channel alterations in the diabetic atrium.Liu et al. [35] also reported that SK2 protein levels was decreased and SK3 protein elevels were increased in the atrium of T2DM rats.Metformin treatment prevents the SK channel alterations by inhibiting the PKC/extracellular signal regulated kinase pathway.Long term treatment of metformin also upregulated the SK2 channel and downregulated the SK3 channel by inhibiting the nicotinamide adeninedinucleotide phosphate oxidase 4/p38 mitogen-activated protein kinase (MAPK) signaling pathway [36].

Other Channels
Howarth et al. [37] evaluated gene expression in the sinoatrial node of GK rats and found hyperpolarization-activated cyclic nucleotide-gated channels (HCN) were downregulated.The reduction of HCN isoforms were also reduced in the sinoatrial node of diabetic rats induced with STZ injection, indicating HCN might be an important contributor to the dysfunction of sinoatrial node in DM [38].mRNA and protein expressions of hyperpolarizationactivated cyclic nucleotide-gated channel 2 (HCN2) were reduced exclusively in the ventricles of STZ rats [39].However, HCN2 expression in the atrium of STZ rats and H9c2 cells treated with high glucose were unchanged.
Higher protein expression levels of Na + -Ca 2+ exchanger current (NCX) were observed in the STZ-induced diabetic group [6].Yang et al. [4] observed the electrophysiological abnormalities of diabetic rats were accompanied by more severe oxidative stress and higher protein expression of NCX in the atrium.The protein level of NCX in the atrial tissue of diabetic rats was upregulated without alterations in mRNA.Allopurinol (a xanthine oxidase inhibitor) intervention can downregulate its protein level, which indicates that NCX activation plays a key role in diabetic electrical remodeling of the atrium, and antioxidant treatment improves electrical remodeling by inhibiting NCX expression.

Conclusions
AF contributes to increase morbidity and mortality, especially in the DM population.Rhythm control is important to treat AF [40] and catheter ablation is the most effective treatment for AF [2].However, success rate of ablation in diabetic patients remains lower compared to the general population particularly for those with persistent AF [8].This is likely due to the complex substrate of AF in patients with diabetes, which may be related to chronic inflammation [41], sarcoplasmic endoplasmic reticulum calcium ATPase (SERCA) levels [42] or epigenetics, such as altered expression of microRNA [12] in AF patients.Antiinflammatory agents may reduce AF recurrence post ablation [41].Selective microRNA therapy, by upregulation or downregulation by microRNA, may be used to treat AF to prevent cardiac structural and electrical remodeling [12].Various remodeling of ion channels occurs in diabetes, including the sodium channels, calcium channels, potassium channels and others, resulting in abnormal electrophysiological parameters of the atrium and increases the incidence of AF (Fig. 1).However, how these ion channels are regulated in the diabetic atrium is not fully understood.Therefore, molecular mechanisms of atrial electrical remodeling in diabetes need to be further explored, which may provide new targets for prevention and treatment of AF in diabetes mellitus.
help and advice on writing the manuscript.All authors read and approved the final manuscript.

Fig. 1 .
Fig. 1.Overview of ion channels remodeling contributing to the action potential alteration of the atrial myocytes in diabetes mellitus.In diabetes mellitus, INa, INa-L, ICa, Ito, IKur, SK, NCX in atrium myocytes are altered, resulting in prolonged action potential duration and reduced atrial conduction velocity, increased incidence of atrial fibrillation.INa, the voltage-gated sodium channel current; INa-L, the late sodium current; ICa, the voltage gated calcium current; Ito, the transient outward potassium currents; IKur, the ultra-rapid delayed rectifier potassium current; SK, that small conductance calcium-activated potassium chan-nels; NCX, the Na + -Ca 2+ exchanger current.