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Journal of Diabetes Research
Volume 2018, Article ID 8454078, 12 pages
https://doi.org/10.1155/2018/8454078
Research Article

The Pattern of mRNA Expression Is Changed in Sinoatrial Node from Goto-Kakizaki Type 2 Diabetic Rat Heart

1Department of Physiology, College of Medicine & Health Sciences, UAE University, Al Ain, UAE
2Department of Pharmacology, College of Medicine & Health Sciences, UAE University, Al Ain, UAE
3Cardiovascular Sciences, University of Manchester, Manchester, UK
4Department of Basic Medical Sciences, Mohammed Bin Rashid University of Medicine & Health Sciences, Dubai, UAE

Correspondence should be addressed to F. C. Howarth; ea.ca.ueau@htrawoh.sirhc

Received 10 June 2018; Revised 16 July 2018; Accepted 12 August 2018; Published 2 September 2018

Academic Editor: Michaelangela Barbieri

Copyright © 2018 F. C. Howarth et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background. In vivo experiments in Goto-Kakizaki (GK) type 2 diabetic rats have demonstrated reductions in heart rate from a young age. The expression of genes encoding more than 70 proteins that are associated with the generation and conduction of electrical activity in the GK sinoatrial node (SAN) have been evaluated to further clarify the molecular basis of the low heart rate. Materials and Methods. Heart rate and expression of genes were evaluated with an extracellular electrode and real-time RT-PCR, respectively. Rats aged 12-13 months were employed in these experiments. Results. Isolated spontaneous heart rate was reduced in GK heart (161 ± 12 bpm) compared to controls (229 ± 11 bpm). There were many differences in expression of mRNA, and some of these differences were of particular interest. Compared to control SAN, expression of some genes were downregulated in GK-SAN: gap junction, Gja1 (Cx43), Gja5 (Cx40), Gjc1 (Cx45), and Gjd3 (Cx31.9); cell membrane transport, Trpc1 (TRPC1) and Trpc6 (TRPC6); hyperpolarization-activated cyclic nucleotide-gated channels, Hcn1 (HCN1) and Hcn4 (HCN4); calcium channels, Cacna1d (Cav1.3), Cacna1g (Cav3.1), Cacna1h (Cav3.2), Cacna2d1 (Cavα2δ1), Cacna2d3 (Cavα2δ3), and Cacng4 (Cavγ4); and potassium channels, Kcna2 (Kv1.2), Kcna4 (Kv1.4), Kcna5 (Kv1.5), Kcnb1 (Kv2.1), Kcnd3 (Kv4.3), Kcnj2 (Kir2.1), Kcnk1 (TWIK1), Kcnk5 (K2P5.1), Kcnk6 (TWIK2), and Kcnn2 (SK2) whilst others were upregulated in GK-SAN: Ryr2 (RYR2) and Nppb (BNP). Conclusions. This study provides new insight into the changing expression of genes in the sinoatrial node of diabetic heart.

1. Introduction

Cardiovascular complications are widely reported in diabetic patients and may be associated with various cardiac arrhythmias and sudden cardiac death [15]. Although coronary artery disease and hypertension are risk factors for cardiovascular dysfunction in diabetic patients, there is also a risk of developing cardiac dysfunction that is independent of coronary atherosclerosis and hypertension [6]. Electrical disturbances have been widely reported in diabetic heart [7, 8]. Bolognesi et al. [9] reported that sinus bradycardia and QT prolongation can occur in insulin-treated diabetic patients with severe hypoglycemia. Abnormal functions of sinus node automaticity, third-degree atrioventricular block, and left bundle branch block occur more frequently in diabetic patients [1012]. Type 2 diabetic patients have an increased risk of supraventricular arrhythmias including atrial fibrillation [1, 1316], ventricular tachyarrhythmias, and ventricular fibrillation [3, 5, 7, 17]. Various studies have shown that QT prolongation is an independent risk factor for cardiovascular mortality in diabetic patients [2, 1821]. Howarth et al. [22] reported disturbances in the electrocardiogram including bradycardia and prolongation of the QRS and QT intervals in the GK rat. Soltysinska et al. [23] reported alterations in systolic and diastolic function and prolonged SAN recovery time in db/db diabetic mice. Hyperglycemia, a hallmark of diabetes mellitus, is associated with oxidative stress which in turn exacerbates inflammation and further exacerbates oxidative stress, which in turn may partly underlie QT prolongation and trigger ventricular arrhythmias [24, 25].

In the Zucker diabetic fatty rat, myocardial impulse propagation was impaired [26]. Little is known about the effects of type 2 diabetes mellitus (T2DM) on the electrophysiology of the SAN. In the streptozotocin- (STZ-) induced diabetic rat, SAN conduction, pacemaker cycle length, and action potential duration were prolonged [27, 28]. Various ion channels and ionic conductances including L-type and T-type Ca2+ current, hyperpolarization-activated “funny” current, Na+ current, Na+/Ca2+ exchange current, and various K+ currents are essential for the generation, propagation, and regulation of the SAN action potential [29]. Sarcoplasmic reticulum (SR) Ca2+ might also contribute to the generation and decay of the SAN action potential [30]. Structural and/or functional channelopathies may underlie some of the electrical abnormalities that have been reported in diabetic heart [22]. In order to further elucidate the molecular basis of these heart rhythm disturbances, we have investigated the pattern of more than 70 genes encoding proteins that are associated with the generation and conduction of electrical activity in the SAN in the GK type 2 diabetic heart. Results from this study will provide direction for future structural and functional studies of the electrical conduction system in the diabetic SAN.

2. Materials and Methods

2.1. Experimental Protocol

Ethical approval for this project was obtained from the Animal Ethics Committee, College of Medicine & Health Sciences, UAE University. Male GK and Wistar control rats were reared as previously described [31]. Rats were kept in cages, under a 12 h-12 h light-dark cycle, and had free access to food and tap water. Room temperature was kept between 21 and 25°C. Experiments commenced when the animals were 12–13 months of age. Blood glucose, after an overnight fast, and blood glucose 120 min after a glucose challenge (2 g/kg body weight, intraperitoneal) were measured in GK and age-matched controls. Prior to experiments, the body weight, heart weight, and the nonfasting blood glucose were also measured. The heart to body weight ratio was calculated.

2.2. Measurement of Heart Rate

Rats were sacrificed as previously described using a guillotine [32]. The chest was then opened, and the hearts were rapidly removed and mounted in Langendorff mode and perfused at a constant flow rate of 8 ml.g heart−1 min−1 at 36–37°C with normal Tyrode containing 140 mM NaCl, 5 mM KCl, 1 mM MgCl2, 10 mM glucose, 5 mM HEPES, and 1.8 mM CaCl2 and adjusted to pH 7.4 with NaOH bubbled with oxygen. An extracellular suction electrode was used to measure heart rate according to previously described techniques [33]. Action potentials were recorded in the left ventricle. Electrical signals were collected at 400 Hz. Signals were then amplified (ADInstuments, ML136 Bioamp), delivered to a Powerlab (ADInstruments, PL410), and displayed on a PC monitor. Analysis was performed using ADInstruments software version v 4.21 (ADInstruments, Australia).

2.3. Expression of mRNA

Previously described techniques were used to evaluate the expression of genes encoding more than 70 proteins involved in electrical activity in the SAN [32, 3436]. After rats were sacrificed, the hearts were rapidly removed and placed in a plastic dish containing NaCl 140 mM, KCl 5.4 mM, MgCl2 1 mM, HEPES 5 mM, D-glucose 5.5 mM, and CaCl2 1.8 mM and adjusted to pH 7.4 with NaOH. The ventricles and the left atrium were removed, and the right atrium was opened to expose the SAN and crista terminalis. The SAN artery was used to identify the SAN. The SAN was exposed, and 2 mm biopsy samples of SAN were carefully collected from GK and control rat hearts according to previously described techniques [32]. The samples were placed in RNAlater (AM7021, Life Technologies, Carlsbad, CA, USA) and stored overnight at room temperature to allow thorough penetration of the tissue [32]. The following day, tissue samples were frozen at −20°C in readiness for further processing. Tissue samples were homogenized in homogenization microtubes containing 1.4 mm ceramic beads using a Precellys 24 tissue homogenizer (Bertin Technologies, USA). The homogenization protocol comprised 2 runs at 6500 rpm of 20 seconds each with a 15-second gap. The SV Total RNA Isolation System (Promega, Madison, USA) was used to isolate total RNA from the tissue, in accordance with the manufacturer’s instructions. Spectrophotometric techniques were used to measure the concentration and purity of the RNA samples. The absorbance was measured at 260 nm, and the ratio of absorbance was measured at 260 nm and 280 nm (ND-1000, NanoDrop). cDNA was generated using a 2-step RT-PCR procedure. Total RNA (500 ng) was converted into cDNA in a 25 μl PCR reaction with 10x RT Buffer 2.0 μl, 25x dNTP Mix (100 mM) 0.8 μl, 10x RT Random Primers 2.0 μl, MultiScribe™ Reverse Transcriptase 1.0 μl, RNase inhibitor 1.0 μl, and Nuclease-free H2O (High Capacity cDNA Reverse Transcription Kit, 4374966, Applied Biosystems, USA). A Veriti thermal cycler (Applied Biosystems, USA) was used to perform reverse transcription. The cycles were as follows: 10 min at 5°C, 120 min at 37°C, and 5 min at 85°C. Customized TaqMan Low Density Arrays (Format 32, 4346799, Applied Biosystems, USA) were used for gene expression assays. The TaqMan assays were preloaded in triplicate for each RNA sample. Rat glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was selected as the endogenous control [35, 37]. The expression of GAPDH was not significantly different () between GK and control SAN samples. 100 ng of cDNA was loaded, together with 2x TaqMan Gene Expression Master Mix (No AmpErase UNG, Applied Biosystems, USA), for a total of 100 μl per port. Two SAN tissue samples were combined for each real-time RT-PCR assay. Real-time RT-PCR was carried out in a Fast ABI Prism 7900HT Sequence Detection System (Applied Biosystems, USA). Standard mode PCR thermal cycling parameters were run as follows: 2 min at 50°C, 10 min at 94.5°C, and 40 cycles of 30 sec at 97°C and 59.7°C for 1 min. Analysis was performed using ABI Prism 7900HT SDS, v2.4 software. Statistical analysis was performed using SDS RQ Manager 1.1.4 software employing the 2−ΔΔCt method with a relative quantification RQmin/RQmax confidence set at 95%. The lists of target genes, encoded proteins, and description of the proteins are shown in Table 1.

Table 1: Target genes, encoded proteins, and description of proteins.
2.4. Statistics

Data were expressed as the mean ± SEM of “” observations. Statistical comparisons were performed using an independent samples -test (SPSS v. 20), and a “” value of less than 0.5 was considered to indicate a significant difference.

3. Results

3.1. General Characteristics

Nonfasting blood glucose and fasting blood glucose were significantly () elevated in GK rats compared to age-matched controls. After an overnight fast blood glucose, 120 min following a glucose challenge (2 g/kg body weight, intraperitoneal), was also significantly () elevated in GK compared to controls. Body weight and heart weight were significantly () increased, and heart weight/body weight ratio was significantly () reduced in GK compared to controls (Table 2).

Table 2: General characteristics of GK rats.
3.2. Heart Rate

Heart rate is shown in Figure 1. Heart rate was significantly () reduced in GK (161 ± 12 bpm, ) compared to control (229 ± 11 bpm, ) heart (Figure 1).

Figure 1: Heart rates in GK and control rats. Data are mean ± SEM, hearts.
3.3. Expression of mRNA

Figure 2 shows the expression of mRNA for gap junction proteins. Expression of Gja1 (2-fold), Gja5 (4-fold), Gjc1, and Gjd3 (4-fold) were all downregulated in GK compared to control SAN. Figures 3(a) and 3(b) show expression of mRNA for cell membrane transport and intracellular Ca2+ transport and regulatory proteins, respectively. Expression of Trpc1 and Trpc6 (6-fold) and Itpr13 (2-fold) were downregulated, and Ryr2 was upregulated in GK compared to control SAN. Figure 4 shows the expression of mRNA for hyperpolarization-activated cyclic nucleotide-gated channel proteins. Expression of Hcn1 (4-fold) and Hcn4 (5-fold) were downregulated in GK compared to control SAN. Figure 5 shows the expression of mRNA for calcium channel proteins. Expression of Cacna1d (3-fold), Cacna1g, Cacna1h (4-fold), Cacna2d1, Cacna2d3, and Cacng4 (3-fold) were downregulated in GK compared to control SAN. Figures 6(a) and 6(b) show the expression of mRNA for potassium channel proteins. Expression of Kcna2, Kcna4, Kcna5 (5-fold), Kcnb1, Kcnd3, Kcnj2, Kcnk1 (7-fold), Kcnk5 (3-fold), Kcnk6, and Kcnn2 were downregulated, and Kcnj11, Kcnj5, and Kcnk2 were upregulated in GK compared to control SAN. Figure 7 shows the expression of mRNA for various other proteins that might affect electrical activity in the SAN. Expression of Abcc9 and Nppb were upregulated in GK compared to control SAN.

Figure 2: Expression of genes encoding various gap junction proteins. Data are mean ± SEM, samples, each containing SANs from 2 hearts.
Figure 3: (a) Expression of genes encoding various membrane transport and (b) intracellular Ca2+ transport and regulatory proteins. Data are mean ± SEM, n = 6–9 samples, each containing SANs from 2 hearts.
Figure 4: Expression of genes encoding various hyperpolarization-activated cyclic nucleotide-gated channels. Data are mean ± SEM, n = 6–8 samples, each containing SANs from 2 hearts.
Figure 5: Expression of genes encoding various calcium channel proteins. Data are mean ± SEM, n = 7–9 samples, each containing SANs from 2 hearts.
Figure 6: (a) and (b) Expression of genes encoding various potassium channel proteins. Data are mean ± SEM, n = 8–9 samples, each containing SANs from 2 hearts.
Figure 7: Expression of genes encoding miscellaneous cardiac proteins. Data are mean ± SEM, n = 5–9 samples, each containing SANs from 2 hearts.

4. Discussion

Previous in vivo biotelemetry experiments performed in GK type 2 diabetic rats have demonstrated disturbances in the electrocardiogram. These disturbances have included reduced heart rate which was associated with prolonged QRS and QT intervals [22]. The spontaneous heart beat in isolated perfused heart was also lower in GK compared to control rats which suggests that the bradycardia is at least partly attributed to an intrinsic defect in the electrical conduction system of the heart [33]. Changes in membrane potential that occur during the different phases of the SAN action potential are produced by changes in the movement of various ions (Na+, Ca2+, and K+) across the cell membrane and by the movement of Ca2+ in or out of the SR [30].

Disturbances in one or more of these ionic conductances would be expected to alter the electrophysiological properties of the SAN. In order to further clarify the molecular basis of the low heart rate in the GK rat, the expression of more than 70 genes that encode proteins that are associated with the generation and conduction of electrical activity in the GK SAN were evaluated.

Regarding the changes in mRNA of particular interest were (i) downregulation of Gja5 and Gjd3 (4-fold), (ii) downregulation of Trpc6 (6-fold), (iii) downregulation of Hcn4 (5-fold), (iv) downregulation of Cacna1d (3-fold), Cacna1h (4-fold), and Cacng4 (3-fold), (v) downregulation of Kcna5 (5-fold) and Kcnk1 (7-fold) in GK compared to control SAN, and (vi) upregulation of Nppb in GK compared to control SAN.

Gja5 (Cx40) and Gjd3 (Cx31.9) were downregulated in GK compared to control SAN. The connexins are structurally related proteins that assemble to form gap junctions which play an important role in cell-to-cell electrical communication and cardiac rhythmicity. At least 5 connexins (Connexin30.2, Connexin37, Connexin40, Connexin43, and Connexin45) are expressed in the heart, and each connexin displays regional and cell type-specific expression [38]. Downregulation of connexin proteins may result in impaired electrical conduction between cells and, hence, may have implications for electrical transmission among the cells of the SAN and between the SAN and other regions of the heart [27].

Trpc6 (TRPC6) and to a lesser extent Trpc1 (TRPC1) were downregulated in GK compared to control SAN. The transient receptor potential channels (TRPCs) are a large family of ion channels that are widely expressed in human tissue including the heart and the vasculature [39, 40]. TRPC1 and TRPC6 are stretch-activated, nonselective cation channels expressed in ventricular muscle from mouse heart [41]. The TRPCs have been found to play a role in cardiovascular disease [42]. Upregulation of TRPCs is involved in the pathophysiology of cardiac hypertrophy and heart failure [39, 43, 44]. Previous studies have demonstrated that cardiac hypertrophy upregulated TRPC6 and inhibition of TRPC6 suppressed agonist-induced hypertrophic responses [4547]. Downregulation of Trpc1 and Trpc6 in the GK SAN may have implications for the conduction of Na+ or Ca2+ current through nonselective TRPCs and in turn implications for the generation of action potentials in SAN cells [43, 48, 49].

The SAN generates action potentials automatically, and the cells of the SAN appear to have two separate but closely communicating mechanisms (often referred to as “clocks”). There is a “membrane clock” which consists of ion channels that include the hyperpolarization-activated cyclic nucleotide-gated channels (mainly HCN4), the L-type Ca2+ channels (mainly Cav1.3), and the T-type Ca2+ channels (mainly Cav3.1). There is also a “calcium clock” which consists of Ca2+-handling proteins that include the ryanodine receptor (RyR2), the sarcoplasmic reticulum-ATPase (SERCA2a), and the Na+/Ca2+ exchanger (NCX1) [50, 51]. Recent studies in human nodal cells have shown that when the clocks become uncoupled, SAN cells fail to generate spontaneous action potentials and β-adrenergic receptor stimulation, which in turn increases cyclic AMP concentration and was able to restore spontaneous, rhythmic action potentials [51]. HCN1–4 proteins are the structural components of the funny channels, and the funny current (If) is the main electrical driving force behind diastolic depolarization.

Hcn4 (HCN4) and to a lesser extent Hcn1 (HCN1) were downregulated in GK compared to control SAN. Shinagawa et al. [29] have shown that a variety of time-dependent and voltage-dependent ionic currents contribute to the rat action potential including the delayed rectifier K+ current, the L-type Ca2+ current, and Na+ current. Hcn4 and Hcn1 are variously expressed in rat atrioventricular node, SAN, right atrium, and ventricle [52]. In human, SAN Hcn4 accounts for 75%, Hcn1 for 21%, Hcn2 for 3%, and Hcn3 for 0.7% of the Hcn in human SAN [53]. It has been shown that Hcn4-related inheritable arrhythmias, resulting in four different mutations of the Hcn4 gene, are associated with reduced heart rate and various arrhythmias [54]. Hcn1 (HCN1) was reduced in GK compared to control SAN. In the absence of the pacemaker channel protein HCN1, mice display congenital SAN dysfunction that was characterized by low heart rate, sinus arrhythmia, increased SAN conduction and recovery time, and recurrent sinus pauses [55]. Downregulation of Hcn1 and Hcn4 might be expected to reduce the slope of the pacemaker potential, prolong the time to threshold potential, and partly underlie the bradycardia in the GK rat heart.

Cacna1d (Cav1.3), Cacna1h (Cav3.2), and Cacng4 (Cavγ4) were downregulated in GK compared to control SAN. Voltage-gated L-type and T-type Ca2+ channels are found in the heart. L-type and T-type Ca2+ currents are important in the generation of phase 4, and L-type Ca2+ current is also important in the generation of phase 0 of SAN and atrioventricular node action potentials. The T-type Ca2+ channels also have roles in cell growth and cardiovascular remodeling. Cacna1g (Cav3.1) and Cacna1h (Cav3.2) encode pore-forming alpha(1) subunits of the T-type Ca2+ channel in cardiac muscle [56]. Cacna1g (Cav3.1) and Cacna1h (Cav3.2) may be involved in the development of the heart’s electrical conduction system [57]. In adult heart, the T-type Ca2+ channel is widely distributed in the conduction system and plays an important role in the generation of the pacemaker depolarization in phase 4 of the SAN action potential [58]. Downregulation of Cacna1h (Cav3.2) might have consequences for the generation of the pacemaker potential and, therefore, the heart rate in the GK SAN. Interestingly, Cacna1c (Cav1.2) which mediates the generation of the pacemaker potential and action potential in the SAN was also modestly reduced in GK compared to control SAN. The gamma subunits of the L-type Ca2+ channel, of which there are 8 isoforms, differentially modulate Ca2+ channel function [59]. Reductions in expression of Cacna1d (Cav1.3) and Cacng4 (Cavγ4) would be consistent with depressed L-type Ca2+ current and modulation of the L-type Ca2+ channel activity and have implications for the pacemaker potential and generation of the action potential and, hence, heart rate in the GK rat heart.

Compared to controls, the general characteristics of the GK rat included elevated nonfasting and fasting blood glucose and glucose intolerance, as evidenced by the raised blood glucose 120 min following a glucose challenge, after an overnight fast. RyR2 channels are found in sarcoplasmic reticulum and have important roles in cardiac myocyte contraction and the generation of autorhythmicity in SAN cells [51]. Recent studies have revealed a crucial role of RyR2 channels in the regulation of insulin release and glucose homeostasis [60]. RyR2 was upregulated in GK compared to CON SAN, and it would be interesting to investigate if the expression of pancreatic β-cell RyR2 is also altered in GK rat which in turn might partly underlie the hyperinsulinemia previously reported in GK compared to CON rat [61].

Kcna5 (Kv1.5) and Kcnk1 (TWIK1) were downregulated in GK compared to control SAN. Kcna5 (Kv1.5) encodes the potassium channel α-subunit Kv1.5 and is widely expressed in heart, brain, and vascular, airway, and smooth muscle cells [6264]. Within the heart, several studies have demonstrated the presence of Kv1.5 in atria, ventricle, and SAN [6567]. The cardiac ultrarapid outward (IKur) current, which is encoded by Kcna5, controls action potential duration and in particular repolarizing current [6870]. Remodeling of ion channels, including the voltage-gated potassium channel Kv1.5, is involved in the pathophysiology of atrial fibrillation [71]. Expression of Kv1.5 results in marked decreases in mouse ventricular myocyte action potential duration [72]. In vivo overexpression of Kv1.5 has been shown to shorten action potential duration, eliminate after depolarizations, and increase heart rate in mice with long QT syndrome [73]. Downregulation of Kcna5 might result in prolonged repolarization and duration of the action potential and, as a consequence, reduction in heart rate. Kcnk1 (TWIK1) is widely expressed in the human heart and brain [74]. TWIK1 channels may be involved in the regulation of the resting membrane potential and, hence, excitability of the cardiac myocyte [75, 76]. Downregulation of Kcnk1 might have implications for resting membrane potential and, hence, excitability of the SAN cell. Abnormal QT prolongation is not an uncommon feature in diabetic heart, and this is associated with reduction in the rapid delayed rectifier K+ current (IKr) in insulin-dependent diabetic heart [77]. Kcnh2, the gene that encodes expression of KCNH2 protein (also known as hERG1) that encodes the hERG channel, was not significantly altered in diabetic SAN.

Nppb (BNP) and to a lesser extent Nppa (ANP) were upregulated in GK compared to control SAN. BNP and ANP are secreted by the atria and ventricles and cause a reduction in blood pressure and cardiac hypertrophy. In the heart, BNP reduces ventricular fibrosis, and BNP and ANP are involved in the pathophysiology of heart failure, coronary heart disease, hypertension, and left ventricular hypertrophy [7880]. Increases in BNP and ANP in blood plasma and atrial tissues are associated with varying effects of these natriuretic peptides on the amplitude and kinetics of shortening and intracellular Ca2+ in ventricular myocytes from STZ-induced diabetic rat [81, 82]. Springer et al. [83] reported that BNP increased electrical conduction velocity and heart rate in isolated heart and in the SAN and also increased spontaneous action potential frequency in SAN cells. Upregulation of Nppb (BNP) and Nppa (ANP) may be associated with mechanisms that compensate for low heart rate in the GK type 2 diabetic rat heart.

It is of interest to compare gene expression in the current study with that in a recent study in SAN from the STZ-induced diabetic rat [32]. For example, Gja5 (Cx40) and Gjd3 (Cx31.9) were downregulated in GK and unaltered in STZ SAN; Trpc6 (TRPC6) was downregulated in GK and upregulated in STZ SAN; Ryr2 (RYR2) was upregulated in GK and unaltered in STZ SAN; Hcn1 (HCN1) and Hcn4 (HCN4) were downregulated in GK and unaltered in STZ SAN; Cacna1d (Cav1.3), Cacna1h (Cav3.2), and Cacng4 (Cavγ4) were downregulated in GK SAN; however, in the STZ study, Cacna1d was unaltered, Cacna1h was upregulated, and Cacng4 was downregulated; Kcna5 (Kv1.5) and Kcnk1 (TWIK1) were downregulated in GK SAN and unaltered in STZ SAN; Nppb (BNP) was upregulated in GK and in STZ SAN. These differences may be partly attributable to the different ages of the animals (GK/CON rats 12–13 months vs. STZ/CON rats 18 weeks of age) and the nature of diabetes (GK genetic vs. STZ chemical induced) in the two experimental models.

Heart failure and T2DM are two increasingly common and related diseases. MicroRNAs (miRs) play an important role in the pathogenesis of structural alterations in the failing heart [84]. Some patients, who are affected by heart failure and who have severe hemodynamic and electrical dysfunction, receive cardiac resynchronization therapy (CRT). Recent studies have shown that CRT is associated with alterations in expression of genes and miRs which regulate a variety of cardiac processes including cardiac apoptosis, cardiac fibrosis, cardiac hypertrophy, cardiac angiogenesis, and membrane channel ionic currents [85, 86]. In the future, interventions that are able to alter expression of miRs might become an important treatment modality for diabetes and heart failure.

The molecular biology results are based on quantitative PCR. The possibility of posttranscriptional modifications, for example, by miR, means that changes in gene expression might not necessarily result in corresponding changes in the expression of proteins. Previous studies have demonstrated that spironolactone regulates HCN protein expression through miR-1 in rats with myocardial infarction and interestingly, upregulation of miR-1 expression partially contributed to the posttranscriptional repression of HCN protein expression [87]. Recent studies have demonstrated that multiple miRs are involved in the regulation of SCN5A/Nav1.5 channel and its β1/SCN1B subunit which is responsible for the fast-activating, fast-inactivating sodium current in atrial and ventricular action potentials [88]. Further structural experiments will be required to investigate the expression of selected proteins and the consequences of these structural changes on the electrophysiology of ion channels in the diabetic heart.

It is hoped that results emerging from laboratory studies will eventually translate into treatment modalities, which might include manipulation of miRs, mRNAs, and associated conduction proteins, in order to normalize electrical dysfunction in the diabetic heart.

5. Conclusions

In conclusion, this study provides valuable insight into the differences in expression of genes that encode proteins that are involved in the generation and conduction of electrical activity in the SAN in the GK diabetic rat and will form the basis for future structural and electrophysiological studies.

Data Availability

The raw mRNA data and statistically analyzed data presented in the figures are available, should it be requested, or available from the corresponding author upon request.

Conflicts of Interest

The authors declare no competing financial or nonfinancial interests.

Acknowledgments

The authors received a research grant from the UAE University.

References

  1. T. Meinertz and K. Sydow, “Diabetes mellitus – risikofaktor für vorhofflimmern: potenzielle therapeutische implikationen,” Herz, vol. 39, no. 3, pp. 320–324, 2014. View at Publisher · View at Google Scholar · View at Scopus
  2. Z. Lu, Y. P. Jiang, C. Y. C. Wu et al., “Increased persistent sodium current due to decreased PI3K signaling contributes to QT prolongation in the diabetic heart,” Diabetes, vol. 62, no. 12, pp. 4257–4265, 2013. View at Publisher · View at Google Scholar · View at Scopus
  3. G. S. Hillis, J. Hata, M. Woodward et al., “Resting heart rate and the risk of microvascular complications in patients with type 2 diabetes mellitus,” Journal of the American Heart Association, vol. 1, no. 5, p. e002832, 2012. View at Publisher · View at Google Scholar · View at Scopus
  4. C. Pappone and V. Santinelli, “Cardiac electrophysiology in diabetes,” Minerva Cardioangiologica, vol. 58, no. 2, pp. 269–276, 2010. View at Google Scholar
  5. M. R. Movahed, M. Hashemzadeh, and M. Jamal, “Increased prevalence of ventricular fibrillation in patients with type 2 diabetes mellitus,” Heart and Vessels, vol. 22, no. 4, pp. 251–253, 2007. View at Publisher · View at Google Scholar · View at Scopus
  6. J. P. Piccini, L. Klein, M. Gheorghiade, and R. O. Bonow, “New insights into diastolic heart failure: role of diabetes mellitus,” The American Journal of Medicine, vol. 116, Supplement 1, no. 5, pp. 64–75, 2004. View at Publisher · View at Google Scholar · View at Scopus
  7. S. Mythri and H. Rajeev, “Left ventricular diastolic dysfunction (LVDD) & cardiovascular autonomic neuropathy (CAN) in type 2 diabetes mellitus (DM): a cross-sectional clinical study,” Journal of Clinical and Diagnostic Research, vol. 9, no. 1, pp. OC18–OC22, 2015. View at Publisher · View at Google Scholar · View at Scopus
  8. S. G. Kanorskiĭ and I. S. Kanorskaia, “Atrial fibrillation in patients with type 2 diabetes mellitus: specific features of development and antirecurrence therapy,” Kardiologiia, vol. 50, no. 7, pp. 31–37, 2010. View at Google Scholar
  9. R. Bolognesi, D. Tsialtas, M. G. Bolognesi, and C. Giumelli, “Marked sinus bradycardia and QT prolongation in a diabetic patient with severe hypoglycemia,” Journal of Diabetes and its Complications, vol. 25, no. 5, pp. 349–351, 2011. View at Publisher · View at Google Scholar · View at Scopus
  10. T. Wasada, K. Katsumori, S. Hasumi et al., “Association of sick sinus syndrome with hyperinsulinemia and insulin resistance in patients with non-insulin-dependent diabetes mellitus: report of four cases,” Internal Medicine, vol. 34, no. 12, pp. 1174–1177, 1995. View at Publisher · View at Google Scholar · View at Scopus
  11. M. R. Movahed, M. Hashemzadeh, and M. M. Jamal, “Increased prevalence of third-degree atrioventricular block in patients with type II diabetes mellitus,” Chest, vol. 128, no. 4, pp. 2611–4, 2005. View at Publisher · View at Google Scholar · View at Scopus
  12. E. Guzman, N. Singh, I. A. Khan et al., “Left bundle branch block in type 2 diabetes mellitus: a sign of advanced cardiovascular involvement,” Annals of Noninvasive Electrocardiology, vol. 9, no. 4, pp. 362–365, 2004. View at Publisher · View at Google Scholar · View at Scopus
  13. V. G. Lychev, E. B. Klester, and L. A. Plinokosova, “Arrhythmia in patients with chronic heart insufficiency and type 2 diabetes mellitus,” Klinicheskaia Meditsina, vol. 92, no. 3, pp. 38–42, 2014. View at Google Scholar
  14. O. Fatemi, E. Yuriditsky, C. Tsioufis et al., “Impact of intensive glycemic control on the incidence of atrial fibrillation and associated cardiovascular outcomes in patients with type 2 diabetes mellitus (from the Action to Control Cardiovascular Risk in Diabetes study),” The American Journal of Cardiology, vol. 114, no. 8, pp. 1217–1222, 2014. View at Publisher · View at Google Scholar · View at Scopus
  15. R. R. Huxley, K. B. Filion, S. Konety, and A. Alonso, “Meta-analysis of cohort and case-control studies of type 2 diabetes mellitus and risk of atrial fibrillation,” The American Journal of Cardiology, vol. 108, no. 1, pp. 56–62, 2011. View at Publisher · View at Google Scholar · View at Scopus
  16. X. Du, T. Ninomiya, B. de Galan et al., “Risks of cardiovascular events and effects of routine blood pressure lowering among patients with type 2 diabetes and atrial fibrillation: results of the ADVANCE study,” European Heart Journal, vol. 30, no. 9, pp. 1128–1135, 2009. View at Publisher · View at Google Scholar · View at Scopus
  17. C. Xie, J. Hu, L. J. Motloch, B. S. Karam, and F. G. Akar, “The classically cardioprotective agent diazoxide elicits arrhythmias in type 2 diabetes mellitus,” Journal of the American College of Cardiology, vol. 66, no. 10, pp. 1144–1156, 2015. View at Publisher · View at Google Scholar · View at Scopus
  18. A. J. Cox, A. Azeem, J. Yeboah et al., “Heart rate-corrected QT interval is an independent predictor of all-cause and cardiovascular mortality in individuals with type 2 diabetes: the diabetes heart study,” Diabetes Care, vol. 37, no. 5, pp. 1454–1461, 2014. View at Publisher · View at Google Scholar · View at Scopus
  19. A. B. Lehtinen, C. Newton-Cheh, J. T. Ziegler et al., “Association of NOS1AP genetic variants with QT interval duration in families from the diabetes heart study,” Diabetes, vol. 57, no. 4, pp. 1108–1114, 2008. View at Publisher · View at Google Scholar · View at Scopus
  20. B. Linnemann and H. U. Janka, “Prolonged QTc interval and elevated heart rate identify the type 2 diabetic patient at high risk for cardiovascular death. The Bremen diabetes study,” Experimental and Clinical Endocrinology & Diabetes, vol. 111, no. 4, pp. 215–222, 2003. View at Publisher · View at Google Scholar · View at Scopus
  21. M. Veglio, G. Bruno, M. Borra et al., “Prevalence of increased QT interval duration and dispersion in type 2 diabetic patients and its relationship with coronary heart disease: a population-based cohort,” Journal of Internal Medicine, vol. 251, no. 4, pp. 317–324, 2002. View at Publisher · View at Google Scholar · View at Scopus
  22. F. C. Howarth, M. Jacobson, M. Shafiullah, and E. Adeghate, “Long-term effects of type 2 diabetes mellitus on heart rhythm in the Goto-Kakizaki rat,” Experimental Physiology, vol. 93, no. 3, pp. 362–369, 2008. View at Publisher · View at Google Scholar · View at Scopus
  23. E. Soltysinska, T. Speerschneider, S. V. Winther, and M. B. Thomsen, “Sinoatrial node dysfunction induces cardiac arrhythmias in diabetic mice,” Cardiovascular Diabetology, vol. 13, no. 1, p. 122, 2014. View at Publisher · View at Google Scholar · View at Scopus
  24. C. Sardu, G. Carreras, S. Katsanos et al., “Metabolic syndrome is associated with a poor outcome in patients affected by outflow tract premature ventricular contractions treated by catheter ablation,” BMC Cardiovascular Disorders, vol. 14, no. 1, p. 176, 2014. View at Publisher · View at Google Scholar · View at Scopus
  25. M. Dong and J. Ren, “What fans the fire: insights into mechanisms of leptin in metabolic syndrome-associated heart diseases,” Current Pharmaceutical Design, vol. 20, no. 4, pp. 652–658, 2014. View at Publisher · View at Google Scholar · View at Scopus
  26. K. B. Olsen, L. N. Axelsen, T. H. Braunstein et al., “Myocardial impulse propagation is impaired in right ventricular tissue of Zucker diabetic fatty (ZDF) rats,” Cardiovascular Diabetology, vol. 12, no. 1, p. 19, 2013. View at Publisher · View at Google Scholar · View at Scopus
  27. F. C. Howarth, N. Nowotny, E. Zilahi, M. A. El Haj, and M. Lei, “Altered expression of gap junction connexin proteins may partly underlie heart rhythm disturbances in the streptozotocin-induced diabetic rat heart,” Molecular and Cellular Biochemistry, vol. 305, no. 1-2, pp. 145–151, 2007. View at Publisher · View at Google Scholar · View at Scopus
  28. M. Watanabe, H. Yokoshiki, H. Mitsuyama, K. Mizukami, T. Ono, and H. Tsutsui, “Conduction and refractory disorders in the diabetic atrium,” American Journal of Physiology-Heart and Circulatory Physiology, vol. 303, no. 1, pp. H86–H95, 2012. View at Publisher · View at Google Scholar · View at Scopus
  29. Y. Shinagawa, H. Satoh, and A. Noma, “The sustained inward current and inward rectifier K+ current in pacemaker cells dissociated from rat sinoatrial node,” The Journal of Physiology, vol. 523, no. 3, pp. 593–605, 2000. View at Publisher · View at Google Scholar · View at Scopus
  30. O. Monfredi, H. Dobrzynski, T. Mondal, M. R. Boyett, and G. M. Morris, “The anatomy and physiology of the sinoatrial node—a contemporary review,” Pacing and Clinical Electrophysiology, vol. 33, no. 11, pp. 1392–1406, 2010. View at Publisher · View at Google Scholar · View at Scopus
  31. M. Smail, L. al Kury, M. A. Qureshi et al., “Cell shortening and calcium dynamics in epicardial and endocardial myocytes from the left ventricle of Goto-Kakizaki type 2 diabetic rats,” Experimental Physiology, vol. 103, no. 4, pp. 502–511, 2018. View at Publisher · View at Google Scholar · View at Scopus
  32. Z. Ferdous, M. A. Qureshi, P. Jayaprakash et al., “Different profile of mRNA expression in sinoatrial node from streptozotocin-induced diabetic rat,” PLoS One, vol. 11, no. 4, article e0153934, 2016. View at Publisher · View at Google Scholar · View at Scopus
  33. F. C. Howarth, M. Shafiullah, and M. A. Qureshi, “Chronic effects of type 2 diabetes mellitus on cardiac muscle contraction in the Goto-Kakizaki rat,” Experimental Physiology, vol. 92, no. 6, pp. 1029–1036, 2007. View at Publisher · View at Google Scholar · View at Scopus
  34. K. A. Salem, T. E. Adrian, M. A. Qureshi, K. A. Parekh, M. Oz, and F. C. Howarth, “Shortening and intracellular Ca2+ in ventricular myocytes and expression of genes encoding cardiac muscle proteins in early onset type 2 diabetic Goto-Kakizaki rats,” Experimental Physiology, vol. 97, no. 12, pp. 1281–1291, 2012. View at Publisher · View at Google Scholar · View at Scopus
  35. K. A. Salem, M. A. Qureshi, V. Sydorenko et al., “Effects of exercise training on excitation-contraction coupling and related mRNA expression in hearts of Goto-Kakizaki type 2 diabetic rats,” Molecular and Cellular Biochemistry, vol. 380, no. 1-2, pp. 83–96, 2013. View at Publisher · View at Google Scholar · View at Scopus
  36. E. M. Gaber, P. Jayaprakash, M. A. Qureshi et al., “Effects of a sucrose-enriched diet on the pattern of gene expression, contraction and Ca2+ transport in Goto-Kakizaki type 2 diabetic rat heart,” Experimental Physiology, vol. 99, no. 6, pp. 881–893, 2014. View at Publisher · View at Google Scholar · View at Scopus
  37. A. D’Souza, F. C. Howarth, J. Yanni et al., “Chronic effects of mild hyperglycaemia on left ventricle transcriptional profile and structural remodelling in the spontaneously type 2 diabetic Goto-Kakizaki rat,” Heart Failure Reviews, vol. 19, no. 1, pp. 65–74, 2014. View at Publisher · View at Google Scholar · View at Scopus
  38. H. S. Duffy, A. G. Fort, and D. C. Spray, “Cardiac connexins: genes to nexus,” in Cardiovascular Gap Junctions, S. Dhein, Ed., vol. 42 of Advances in Cardiology, pp. 1–17, Adv Cardiol, Basel, Karger, 2006. View at Publisher · View at Google Scholar · View at Scopus
  39. H. Watanabe, M. Murakami, T. Ohba, K. Ono, and H. Ito, “The pathological role of transient receptor potential channels in heart disease,” Circulation Journal, vol. 73, no. 3, pp. 419–427, 2009. View at Publisher · View at Google Scholar · View at Scopus
  40. R. S. Bon and D. J. Beech, “In pursuit of small molecule chemistry for calcium-permeable non-selective TRPC channels – mirage or pot of gold?” British Journal of Pharmacology, vol. 170, no. 3, pp. 459–474, 2013. View at Publisher · View at Google Scholar · View at Scopus
  41. M. L. Ward, I. A. Williams, Y. Chu, P. J. Cooper, Y. K. Ju, and D. G. Allen, “Stretch-activated channels in the heart: contributions to length-dependence and to cardiomyopathy,” Progress in Biophysics and Molecular Biology, vol. 97, no. 2-3, pp. 232–249, 2008. View at Publisher · View at Google Scholar · View at Scopus
  42. J. Rowell, N. Koitabashi, and D. A. Kass, “TRP-ing up heart and vessels: canonical transient receptor potential channels and cardiovascular disease,” Journal of Cardiovascular Translational Research, vol. 3, no. 5, pp. 516–524, 2010. View at Publisher · View at Google Scholar · View at Scopus
  43. M. Nishida and H. Kurose, “Roles of TRP channels in the development of cardiac hypertrophy,” Naunyn-Schmiedeberg's Archives of Pharmacology, vol. 378, no. 4, pp. 395–406, 2008. View at Publisher · View at Google Scholar · View at Scopus
  44. R. Guinamard and P. Bois, “Involvement of transient receptor potential proteins in cardiac hypertrophy,” Biochimica et Biophysica Acta, vol. 1772, no. 8, pp. 885–894, 2007. View at Publisher · View at Google Scholar · View at Scopus
  45. C. Vindis, R. D’Angelo, E. Mucher, A. Nègre-Salvayre, A. Parini, and J. Mialet-Perez, “Essential role of TRPC1 channels in cardiomyoblasts hypertrophy mediated by 5-HT2A serotonin receptors,” Biochemical and Biophysical Research Communications, vol. 391, no. 1, pp. 979–983, 2010. View at Publisher · View at Google Scholar · View at Scopus
  46. M. Nishida, K. Watanabe, M. Nakaya, and H. Kurose, “Mechanism of cardiac hypertrophy via diacylglycerol-sensitive TRPC channels,” Yakugaku Zasshi, vol. 130, no. 3, pp. 295–302, 2010. View at Publisher · View at Google Scholar · View at Scopus
  47. M. Nishida, K. Watanabe, Y. Sato et al., “Phosphorylation of TRPC6 channels at Thr69 is required for anti-hypertrophic effects of phosphodiesterase 5 inhibition,” The Journal of Biological Chemistry, vol. 285, no. 17, pp. 13244–13253, 2010. View at Publisher · View at Google Scholar · View at Scopus
  48. M. T. Harper, J. E. C. Londono, K. Quick et al., “Transient receptor potential channels function as a coincidence signal detector mediating phosphatidylserine exposure,” Science Signaling, vol. 6, no. 281, article ra50, 2013. View at Publisher · View at Google Scholar · View at Scopus
  49. Y. H. Sun, Y. Q. Li, S. L. Feng et al., “Calcium-sensing receptor activation contributed to apoptosis stimulates TRPC6 channel in rat neonatal ventricular myocytes,” Biochemical and Biophysical Research Communications, vol. 394, no. 4, pp. 955–961, 2010. View at Publisher · View at Google Scholar · View at Scopus
  50. O. Monfredi, V. A. Maltsev, and E. G. Lakatta, “Modern concepts concerning the origin of the heartbeat,” Physiology (Bethesda), vol. 28, no. 2, pp. 74–92, 2013. View at Publisher · View at Google Scholar · View at Scopus
  51. K. Tsutsui, O. J. Monfredi, S. G. Sirenko-Tagirova et al., “A coupled-clock system drives the automaticity of human sinoatrial nodal pacemaker cells,” Science Signaling, vol. 11, no. 534, 2018. View at Publisher · View at Google Scholar
  52. Y. Ou, X. L. Niu, and F. X. Ren, “Expression of key ion channels in the rat cardiac conduction system by laser capture microdissection and quantitative real-time PCR,” Experimental Physiology, vol. 95, no. 9, pp. 938–945, 2010. View at Publisher · View at Google Scholar · View at Scopus
  53. Y. F. Xiao, N. Chandler, H. Dobrzynski et al., “Hysteresis in human HCN4 channels: a crucial feature potentially affecting sinoatrial node pacemaking,” Sheng Li Xue Bao, vol. 62, no. 1, pp. 1–13, 2010. View at Google Scholar
  54. M. Baruscotti, G. Bottelli, R. Milanesi, J. C. DiFrancesco, and D. DiFrancesco, “HCN-related channelopathies,” Pflügers Archiv - European Journal of Physiology, vol. 460, no. 2, pp. 405–415, 2010. View at Publisher · View at Google Scholar · View at Scopus
  55. S. Fenske, S. C. Krause, S. I. H. Hassan et al., “Sick sinus syndrome in HCN1-deficient mice,” Circulation, vol. 128, no. 24, pp. 2585–2594, 2013. View at Publisher · View at Google Scholar · View at Scopus
  56. N. Niwa, K. Yasui, T. Opthof et al., “Cav3.2 subunit underlies the functional T-type Ca2+ channel in murine hearts during the embryonic period,” American Journal of Physiology-Heart and Circulatory Physiology, vol. 286, no. 6, pp. H2257–H2263, 2004. View at Publisher · View at Google Scholar · View at Scopus
  57. E. Mizuta, M. Shirai, K. Arakawa et al., “Different distribution of Cav3.2 and Cav3.1 transcripts encoding T-type Ca2+ channels in the embryonic heart of mice,” Biomedical Research, vol. 31, no. 5, pp. 301–5, 2010. View at Publisher · View at Google Scholar · View at Scopus
  58. K. Ono and T. Iijima, “Cardiac T-type Ca2+ channels in the heart,” Journal of Molecular and Cellular Cardiology, vol. 48, no. 1, pp. 65–70, 2010. View at Publisher · View at Google Scholar · View at Scopus
  59. L. Yang, A. Katchman, J. P. Morrow, D. Doshi, and S. O. Marx, “Cardiac L-type calcium channel (Cav1.2) associates with γ subunits,” The FASEB Journal, vol. 25, no. 3, pp. 928–936, 2011. View at Publisher · View at Google Scholar · View at Scopus
  60. G. Santulli, G. Pagano, C. Sardu et al., “Calcium release channel RyR2 regulates insulin release and glucose homeostasis,” Journal of Clinical Investigation, vol. 125, no. 11, p. 4316, 2015. View at Publisher · View at Google Scholar · View at Scopus
  61. K. Witte, K. Jacke, R. Stahrenberg et al., “Dysfunction of soluble guanylyl cyclase in aorta and kidney of Goto-Kakizaki rats: influence of age and diabetic state,” Nitric Oxide, vol. 6, no. 1, pp. 85–95, 2002. View at Publisher · View at Google Scholar · View at Scopus
  62. L. K. Svoboda, K. G. Reddie, L. Zhang et al., “Redox-sensitive sulfenic acid modification regulates surface expression of the cardiovascular voltage-gated potassium channel Kv1.5,” Circulation Research, vol. 111, no. 7, pp. 842–853, 2012. View at Publisher · View at Google Scholar · View at Scopus
  63. E. Koutsouki, R. S. Lam, G. Seebohm et al., “Modulation of human Kv1.5 channel kinetics by N-cadherin,” Biochemical and Biophysical Research Communications, vol. 363, no. 1, pp. 18–23, 2007. View at Publisher · View at Google Scholar · View at Scopus
  64. W. Guo, K. Kamiya, M. Hojo, I. Kodama, and J. Toyama, “Regulation of Kv4.2 and Kv1.4 K+ channel expression by myocardial hypertrophic factors in cultured newborn rat ventricular cells,” Journal of Molecular and Cellular Cardiology, vol. 30, no. 7, pp. 1449–1455, 1998. View at Publisher · View at Google Scholar · View at Scopus
  65. H. Dobrzynski, S. M. Rothery, D. D. R. Marples et al., “Presence of the Kv1.5 K+ Channel in the Sinoatrial Node,” Journal of Histochemistry & Cytochemistry, vol. 48, no. 6, pp. 769–780, 2000. View at Publisher · View at Google Scholar · View at Scopus
  66. D. Fedida, J. Eldstrom, J. C. Hesketh et al., “Kv1.5 is an important component of repolarizing K+ current in canine atrial myocytes,” Circulation Research, vol. 93, no. 8, pp. 744–751, 2003. View at Publisher · View at Google Scholar · View at Scopus
  67. E. Wettwer, “Is there a functional correlate of Kv1.5 in the ventricle of canine heart and what would it mean for the use of IKur blockers?” British Journal of Pharmacology, vol. 152, no. 6, pp. 835–837, 2007. View at Publisher · View at Google Scholar · View at Scopus
  68. Z. Yang, C. F. Browning, H. Hallaq et al., “Four and a half LIM protein 1: a partner for KCNA5 in human atrium,” Cardiovascular Research, vol. 78, no. 3, pp. 449–457, 2008. View at Publisher · View at Google Scholar · View at Scopus
  69. S. M. Schumacher, D. P. McEwen, L. Zhang, K. L. Arendt, K. M. van Genderen, and J. R. Martens, “Antiarrhythmic drug-induced internalization of the atrial-specific K+ channel Kv1.5,” Circulation Research, vol. 104, no. 12, pp. 1390–1398, 2009. View at Publisher · View at Google Scholar · View at Scopus
  70. S. M. Schumacher-Bass, E. D. Vesely, L. Zhang et al., “Role for myosin-V motor proteins in the selective delivery of Kv channel isoforms to the membrane surface of cardiac myocytes,” Circulation Research, vol. 114, no. 6, pp. 982–992, 2014. View at Publisher · View at Google Scholar · View at Scopus
  71. X. H. Ou, M. L. Li, R. Liu et al., “Remodeling of Kv1.5 channel in right atria from Han Chinese patients with atrial fibrillation,” Medical Science Monitor, vol. 21, pp. 1207–1213, 2015. View at Publisher · View at Google Scholar · View at Scopus
  72. H. Li, W. Guo, H. Xu, R. Hood, A. T. Benedict, and J. M. Nerbonne, “Functional expression of a GFP-tagged Kv1.5 α-subunit in mouse ventricle,” American Journal of Physiology-Heart and Circulatory Physiology, vol. 281, no. 5, pp. H1955–H1967, 2001. View at Publisher · View at Google Scholar
  73. M. Brunner, S. A. Kodirov, G. F. Mitchell et al., “In vivo gene transfer of Kv1.5 normalizes action potential duration and shortens QT interval in mice with long QT phenotype,” American Journal of Physiology-Heart and Circulatory Physiology, vol. 285, no. 1, pp. H194–H203, 2003. View at Publisher · View at Google Scholar
  74. F. Lesage, R. Reyes, M. Fink, F. Duprat, E. Guillemare, and M. Lazdunski, “Dimerization of TWIK-1 K+ channel subunits via a disulfide bridge,” The EMBO Journal, vol. 15, no. 23, pp. 6400–6407, 1996. View at Google Scholar
  75. H. Chen, F. C. Chatelain, and F. Lesage, “Altered and dynamic ion selectivity of K+ channels in cell development and excitability,” Trends in Pharmacological Sciences, vol. 35, no. 9, pp. 461–9, 2014. View at Publisher · View at Google Scholar · View at Scopus
  76. L. Ma, X. Zhang, and H. Chen, “TWIK-1 two-pore domain potassium channels change ion selectivity and conduct inward leak sodium currents in hypokalemia,” Science Signaling, vol. 4, no. 176, p. ra37, 2011. View at Publisher · View at Google Scholar · View at Scopus
  77. Y. Zhang, J. Xiao, H. Wang et al., “Restoring depressed HERG K+ channel function as a mechanism for insulin treatment of abnormal QT prolongation and associated arrhythmias in diabetic rabbits,” American Journal of Physiology-Heart and Circulatory Physiology, vol. 291, no. 3, pp. H1446–H1455, 2006. View at Publisher · View at Google Scholar · View at Scopus
  78. M. Volpe, S. Rubattu, and J. Burnett Jr, “Natriuretic peptides in cardiovascular diseases: current use and perspectives,” European Heart Journal, vol. 35, no. 7, pp. 419–425, 2014. View at Publisher · View at Google Scholar · View at Scopus
  79. L. R. Potter, A. R. Yoder, D. R. Flora, L. K. Antos, and D. M. Dickey, “Natriuretic peptides: their structures, receptors, physiologic functions and therapeutic applications,” in cGMP: Generators, Effectors and Therapeutic Implications, H. H. H. W. Schmidt, F. Hofmann, and J. P. Stasch, Eds., vol. 191 of Handbook of Experimental Pharmacology, pp. 341–366, Springer, Berlin, Heidelberg, 2009. View at Publisher · View at Google Scholar · View at Scopus
  80. M. F. McGrath, M. L. Kuroski de Bold, and A. J. de Bold, “The endocrine function of the heart,” Trends in Endocrinology & Metabolism, vol. 16, no. 10, pp. 469–477, 2005. View at Publisher · View at Google Scholar · View at Scopus
  81. F. C. Howarth, N. A. Shamsi, M. A. Qaydi et al., “Effects of brain natriuretic peptide on contraction and intracellular Ca2+ in ventricular myocytes from the streptozotocin-induced diabetic rat,” Annals of the New York Academy of Sciences, vol. 1084, no. 1, pp. 155–165, 2006. View at Publisher · View at Google Scholar · View at Scopus
  82. F. C. Howarth, A. Adem, E. A. Adeghate et al., “Distribution of atrial natriuretic peptide and its effects on contraction and intracellular calcium in ventricular myocytes from streptozotocin-induced diabetic rat,” Peptides, vol. 26, no. 4, pp. 691–700, 2005. View at Publisher · View at Google Scholar · View at Scopus
  83. J. Springer, J. Azer, R. Hua et al., “The natriuretic peptides BNP and CNP increase heart rate and electrical conduction by stimulating ionic currents in the sinoatrial node and atrial myocardium following activation of guanylyl cyclase-linked natriuretic peptide receptors,” Journal of Molecular and Cellular Cardiology, vol. 52, no. 5, pp. 1122–1134, 2012. View at Publisher · View at Google Scholar · View at Scopus
  84. R. Marfella, C. di Filippo, N. Potenza et al., “Circulating microRNA changes in heart failure patients treated with cardiac resynchronization therapy: responders vs. non-responders,” European Journal of Heart Failure, vol. 15, no. 11, pp. 1277–1288, 2013. View at Publisher · View at Google Scholar · View at Scopus
  85. C. Sardu, M. Barbieri, M. R. Rizzo, P. Paolisso, G. Paolisso, and R. Marfella, “Cardiac resynchronization therapy outcomes in type 2 diabetic patients: role of microRNA changes,” Journal of Diabetes Research, vol. 2016, Article ID 7292564, 8 pages, 2016. View at Publisher · View at Google Scholar · View at Scopus
  86. C. Sardu, G. Paolisso, and R. Marfella, “Letter by Sardu et al “regarding article, circulating microRNA-30d is associated with response to cardiac resynchronization therapy in heart failure and regulates cardiomyocyte apoptosis: a translational pilot study”,” Circulation, vol. 133, no. 6, p. e388, 2016. View at Publisher · View at Google Scholar · View at Scopus
  87. H. D. Yu, S. Xia, C. Q. Zha, S. B. Deng, J. L. Du, and Q. She, “Spironolactone regulates HCN protein expression through micro-RNA-1 in rats with myocardial infarction,” Journal of Cardiovascular Pharmacology, vol. 65, no. 6, pp. 587–592, 2015. View at Publisher · View at Google Scholar · View at Scopus
  88. H. Daimi, E. Lozano-Velasco, A. Haj Khelil et al., “Regulation of SCN5A by microRNAs: miR-219 modulates SCN5A transcript expression and the effects of flecainide intoxication in mice,” Heart Rhythm, vol. 12, no. 6, pp. 1333–1342, 2015. View at Publisher · View at Google Scholar · View at Scopus