Ouabain's Influence on TRPV4 Channels of Epithelial Cells: An Exploration of TRPV4 Activity, Expression, and Signaling Pathways.

Ouabain, a substance originally obtained from plants, is now classified as a hormone because it is produced endogenously in certain animals, including humans. However, its precise effects on the body remain largely unknown. Previous studies have shown that ouabain can influence the phenotype of epithelial cells by affecting the expression of cell-cell molecular components and voltage-gated potassium channels. In this study, we conducted whole-cell clamp assays to determine whether ouabain affects the activity and/or expression of TRPV4 channels. Our findings indicate that ouabain has a statistically significant effect on the density of TRPV4 currents (dITRPV4), with an EC50 of 1.89 nM. Regarding treatment duration, dITRPV4 reaches its peak at around 1 h, followed by a subsequent decline and then a resurgence after 6 h, suggesting a short-term modulatory effect related to on TRPV4 channel activity and a long-term effect related to the promotion of synthesis of new TRPV4 channel units. The enhancement of dITRPV4 induced by ouabain was significantly lower in cells seeded at low density than in cells in a confluent monolayer, indicating that the action of ouabain depends on intercellular contacts. Furthermore, the fact that U73122 and neomycin suppress the effect caused by ouabain in the short term suggests that the short-term induced enhancement of dITRPV4 is due to the depletion of PIP2 stores. In contrast, the fact that the long-term effect is inhibited by PP2, wortmannin, PD, FR18, and IKK16 suggests that cSrc, PI3K, Erk1/2, and NF-kB are among the components included in the signaling pathways.

Biomimetic Cardiac Tissue Models for In Vitro Arrhythmia Studies.

Cardiac arrhythmias are a major cause of cardiovascular mortality worldwide. Many arrhythmias are caused by reentry, a phenomenon where excitation waves circulate in the heart. Optical mapping techniques have revealed the role of reentry in arrhythmia initiation and fibrillation transition, but the underlying biophysical mechanisms are still difficult to investigate in intact hearts. Tissue engineering models of cardiac tissue can mimic the structure and function of native cardiac tissue and enable interactive observation of reentry formation and wave propagation. This review will present various approaches to constructing cardiac tissue models for reentry studies, using the authors' work as examples. The review will highlight the evolution of tissue engineering designs based on different substrates, cell types, and structural parameters. A new approach using polymer materials and cellular reprogramming to create biomimetic cardiac tissues will be introduced. The review will also show how computational modeling of cardiac tissue can complement experimental data and how such models can be applied in the biomimetics of cardiac tissue.

Ionic current changes underlying action potential repolarization responses to physiological pacing and adrenergic stimulation in adult rat ventricular myocytes.

This study aimed to simulate ventricular responses to elevations in myocyte pacing and adrenergic stimulation using a novel electrophysiological rat model and investigate ion channel responses underlying action potential (AP) modulations. Peak ion currents and AP repolarization to 50% and 90% of full repolarization (APD50-90 ) were recorded during simulations at 1-10 Hz pacing under control and adrenergic stimulation conditions. Further simulations were performed with incremental ion current block (L-type calcium current, ICa ; transient outward current, Ito ; slow delayed rectifier potassium current, IKs ; rapid delayed rectifier potassium current, IKr ; inward rectifier potassium current, IK1 ) to identify current influence on AP response to exercise. Simulated APD50-90 closely resembled experimental findings. Rate-dependent increases in IKs (6%-101%), IKr (141%-1339%), and ICa (0%-15%) and reductions in Ito (11%-57%) and IK1 (1%-9%) were observed. Meanwhile, adrenergic stimulation triggered moderate increases in all currents (23%-67%) except IK1 . Further analyses suggest AP plateau is most sensitive to modulations in Ito and ICa while late repolarization is most sensitive to IK1 , ICa , and IKs , with alterations in IKs predominantly stimulating the greatest magnitude of influence on late repolarization (35%-846% APD90 prolongation). The modified Leeds rat model (mLR) is capable of accurately modeling APs during physiological stress. This study highlights the importance of ICa , Ito , IK1, and IKs in controlling electrophysiological responses to exercise. This work will benefit the study of cardiac dysfunction, arrythmia, and disease, though future physiologically relevant experimental studies and model development are required.

Human induced pluripotent stem cell-derived cardiomyocytes as an electrophysiological model: Opportunities and challenges-The Hamburg perspective.

Models based on human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) are proposed in almost any field of physiology and pharmacology. The development of human induced pluripotent stem cell-derived cardiomyocytes is expected to become a step forward to increase the translational power of cardiovascular research. Importantly they should allow to study genetic effects on an electrophysiological background close to the human situation. However, biological and methodological issues revealed when human induced pluripotent stem cell-derived cardiomyocytes were used in experimental electrophysiology. We will discuss some of the challenges that should be considered when human induced pluripotent stem cell-derived cardiomyocytes will be used as a physiological model.

Ion dynamics and the regulation of circadian cellular physiology.

Circadian rhythms in physiology and behavior allow organisms to anticipate the daily environmental changes imposed by the rotation of our planet around its axis. Although these rhythms eventually manifest at the organismal level, a cellular basis for circadian rhythms has been demonstrated. Significant contributors to these cell-autonomous rhythms are daily cycles in gene expression and protein translation. However, recent data revealed cellular rhythms in other biological processes, including ionic currents, ion transport, and cytosolic ion abundance. Circadian rhythms in ion currents sustain circadian variation in action potential firing rate, which coordinates neuronal behavior and activity. Circadian regulation of metal ions abundance and dynamics is implicated in distinct cellular processes, from protein translation to membrane activity and osmotic homeostasis. In turn, studies showed that manipulating ion abundance affects the expression of core clock genes and proteins, suggestive of a close interplay. However, the relationship between gene expression cycles, ion dynamics, and cellular function is still poorly characterized. In this review, I will discuss the mechanisms that generate ion rhythms, the cellular functions they govern, and how they feed back to regulate the core clock machinery.

Analysis of Electrophysiological Mechanisms of N-Deacetyllapaconitine Monochlorhydrate, the Main Metabolite of Lappaconitine Hydrobromide.

In in vitro experiments on isolated rat hippocampal neurons, we studied the electrophysiological mechanisms of the antiarrhythmic effects of N-deacetyllappaconitine monochlorhydrate (DALCh), active metabolite of lappaconitine hydrobromide (allapinin). Electrical activity of neurons was recorded by the patch-clamp method in the whole cell configuration. It was shown that DALCh increased the duration of both slow and fast depolarization phases and decreased the amplitude of the action potential. DALCh effectively inhibited transmembrane currents of Na+ ions and partially K+ ions through the corresponding transmembrane voltage-gated ion channels. Thus, DALCh, in contrast to lappaconitine hydrobromide, belongs not to 1C, but to the 1A class of antiarrhythmics according to the Vaughan-Williams classification.

Pathophysiological Responses to Conotoxin Modulation of Voltage-Gated Ion Currents.

Voltage-gated ion channels are plasma membrane proteins that generate electrical signals following a change in the membrane voltage. Since they are involved in several physiological processes, their dysfunction may be responsible for a series of diseases and pain states particularly related to neuronal and muscular systems. It is well established for decades that bioactive peptides isolated from venoms of marine mollusks belonging to the Conus genus, collectively known as conotoxins, can target different types and isoforms of these channels exerting therapeutic effects and pain relief. For this reason, conotoxins are widely used for either therapeutic purposes or studies on ion channel mechanisms of action disclosure. In addition their positive property, however, conotoxins may generate pathological states through similar ion channel modulation. In this narrative review, we provide pieces of evidence on the pathophysiological impacts that different members of conotoxin families exert by targeting the three most important voltage-gated channels, such as sodium, calcium, and potassium, involved in cellular processes.

Species-dependent differences in the inhibition of various potassium currents and in their effects on repolarization in cardiac ventricular muscle.

Even though rodents are accessible model animals, their electrophysiological properties are deeply different from those of humans, making the translation of rat studies to humans rather difficult. We compared the mechanisms of ventricular repolarization in various animal models to those of humans by measuring cardiac ventricular action potentials from ventricular papillary muscle preparations using conventional microelectrodes and applying selective inhibitors of various potassium transmembrane ion currents. Inhibition of the IK1 current (10 µmol/L barium chloride) significantly prolonged rat ventricular repolarization, but only slightly prolonged it in dogs, and did not affect it in humans. On the contrary, IKr inhibition (50 nmol/L dofetilide) significantly prolonged repolarization in humans, rabbits, and dogs, but not in rats. Inhibition of the IKur current (1 µmol/L XEN-D0101) only prolonged rat ventricular repolarization and had no effect in humans or dogs. Inhibition of the IKs (500 nmol/L HMR-1556) and Ito currents (100 µmol/L chromanol-293B) elicited similar effects in all investigated species. We conclude that dog ventricular preparations have the strongest translational value and rat ventricular preparations have the weakest translational value in cardiac electrophysiological experiments.

Docosahexaenoic acid normalizes QT interval in long QT type 2 transgenic rabbit models in a genotype-specific fashion.

AIM: Long QT syndrome (LQTS) is a cardiac channelopathy predisposing to ventricular arrhythmias and sudden cardiac death. Since current therapies often fail to prevent arrhythmic events in certain LQTS subtypes, new therapeutic strategies are needed. Docosahexaenoic acid (DHA) is a polyunsaturated fatty acid, which enhances the repolarizing IKs current. METHODS AND RESULTS: We investigated the effects of DHA in wild type (WT) and transgenic long QT Type 1 (LQT1; loss of IKs), LQT2 (loss of IKr), LQT5 (reduction of IKs), and LQT2-5 (loss of IKr and reduction of IKs) rabbits. In vivo ECGs were recorded at baseline and after 10 µM/kg DHA to assess changes in heart-rate corrected QT (QTc) and short-term variability of QT (STVQT). Ex vivo monophasic action potentials were recorded in Langendorff-perfused rabbit hearts, and action potential duration (APD75) and triangulation were assessed. Docosahexaenoic acid significantly shortened QTc in vivo only in WT and LQT2 rabbits, in which both α- and ß-subunits of IKs-conducting channels are functionally intact. In LQT2, this led to a normalization of QTc and of its short-term variability. Docosahexaenoic acid had no effect on QTc in LQT1, LQT5, and LQT2-5. Similarly, ex vivo, DHA shortened APD75 in WT and normalized it in LQT2, and additionally decreased AP triangulation in LQT2. CONCLUSIONS: Docosahexaenoic acid exerts a genotype-specific beneficial shortening/normalizing effect on QTc and APD75 and reduces pro-arrhythmia markers STVQT and AP triangulation through activation of IKs in LQT2 rabbits but has no effects if either α- or ß-subunits to IKs are functionally impaired. Docosahexaenoic acid could represent a new genotype-specific therapy in LQT2.

Ion Channel Impairment and Myofilament Ca2+ Sensitization: Two Parallel Mechanisms Underlying Arrhythmogenesis in Hypertrophic Cardiomyopathy.

Life-threatening ventricular arrhythmias are the main clinical burden in patients with hypertrophic cardiomyopathy (HCM), and frequently occur in young patients with mild structural disease. While massive hypertrophy, fibrosis and microvascular ischemia are the main mechanisms underlying sustained reentry-based ventricular arrhythmias in advanced HCM, cardiomyocyte-based functional arrhythmogenic mechanisms are likely prevalent at earlier stages of the disease. In this review, we will describe studies conducted in human surgical samples from HCM patients, transgenic animal models and human cultured cell lines derived from induced pluripotent stem cells. Current pieces of evidence concur to attribute the increased risk of ventricular arrhythmias in early HCM to different cellular mechanisms. The increase of late sodium current and L-type calcium current is an early observation in HCM, which follows post-translation channel modifications and increases the occurrence of early and delayed afterdepolarizations. Increased myofilament Ca2+ sensitivity, commonly observed in HCM, may promote afterdepolarizations and reentry arrhythmias with direct mechanisms. Decrease of K+-currents due to transcriptional regulation occurs in the advanced disease and contributes to reducing the repolarization-reserve and increasing the early afterdepolarizations (EADs). The presented evidence supports the idea that patients with early-stage HCM should be considered and managed as subjects with an acquired channelopathy rather than with a structural cardiac disease.

Canine Myocytes Represent a Good Model for Human Ventricular Cells Regarding Their Electrophysiological Properties.

Due to the limited availability of healthy human ventricular tissues, the most suitable animal model has to be applied for electrophysiological and pharmacological studies. This can be best identified by studying the properties of ion currents shaping the action potential in the frequently used laboratory animals, such as dogs, rabbits, guinea pigs, or rats, and comparing them to those of human cardiomyocytes. The authors of this article with the experience of three decades of electrophysiological studies, performed in mammalian and human ventricular tissues and isolated cardiomyocytes, summarize their results obtained regarding the major canine and human cardiac ion currents. Accordingly, L-type Ca2+ current (ICa), late Na+ current (INa-late), rapid and slow components of the delayed rectifier K+ current (IKr and IKs, respectively), inward rectifier K+ current (IK1), transient outward K+ current (Ito1), and Na+/Ca2+ exchange current (INCX) were characterized and compared. Importantly, many of these measurements were performed using the action potential voltage clamp technique allowing for visualization of the actual current profiles flowing during the ventricular action potential. Densities and shapes of these ion currents, as well as the action potential configuration, were similar in human and canine ventricular cells, except for the density of IK1 and the recovery kinetics of Ito. IK1 displayed a largely four-fold larger density in canine than human myocytes, and Ito recovery from inactivation displayed a somewhat different time course in the two species. On the basis of these results, it is concluded that canine ventricular cells represent a reasonably good model for human myocytes for electrophysiological studies, however, it must be borne in mind that due to their stronger IK1, the repolarization reserve is more pronounced in canine cells, and moderate differences in the frequency-dependent repolarization patterns can also be anticipated.

Ion current profiles in canine ventricular myocytes obtained by the "onion peeling" technique.

The profiles of ion currents during the cardiac action potential can be visualized by the action potential voltage clamp technique. To obtain multiple ion current data from the same cell, the "onion peeling" technique, based on sequential pharmacological dissection of ion currents, has to be applied. Combination of the two methods allows recording of several ion current profiles from the same myocyte under largely physiological conditions. Using this approach, we have studied the densities and integrals of the major cardiac inward (ICa, INCX, INa-late) and outward (IKr, IKs, IK1) currents in canine ventricular cells and studied the correlation between them. For this purpose, canine ventricular cardiomyocytes were chosen because their electrophysiological properties are similar to those of human ones. Significant positive correlation was observed between the density and integral of ICa and IKr, and positive correlation was found also between the integral of ICa and INCX. No further correlations were detected. The Ca2+-sensitivity of K+ currents was studied by comparing their parameters in the case of normal calcium homeostasis and following blockade of ICa. Out of the three K+ currents studied, only IKs was Ca2+-sensitive. The density and integral of IKs was significantly greater, while its time-to-peak value was shorter at normal Ca2+ cycling than following ICa blockade. No differences were detected for IKr or IK1 in this regard. Present results indicate that the positive correlation between ICa and IKr prominently contribute to the balance between inward and outward fluxes during the action potential plateau in canine myocytes. The results also suggest that the profiles of cardiac ion currents have to be studied under physiological conditions, since their behavior may strongly be influenced by the intracellular Ca2+ homeostasis and the applied membrane potential protocol.

Implication of frequency-dependent protocols in antiarrhythmic and proarrhythmic drug testing.

It has long been known that the electrophysiological effects of many cardioactive drugs strongly depend on the rate dependent frequency. This was recognized first for class I antiarrhythmic agents: their Vmax suppressive effect was attenuated at long cycle lengths. Later many Ca2+ channel blockers were also found to follow such kinetics. The explanation was provided by the modulated and the guarded receptor theories. Regarding the duration of cardiac action potentials (APD) an opposite frequency-dependence was observed, i.e. the drug-induced changes in APD were proportional with the cycle length of stimulation, therefore it was referred as "reverse rate-dependency". The beat-to-beat, or short term variability of APD (SV) has been recognized as an important proarrhythmic mechanism, its magnitude can be used as an arrhythmia predictor. SV is modulated by several cardioactive agents, however, these drugs modify also APD itself. In order to clear the drug-specific effects on SV from the concomitant unspecific APD-change related ones, the term of "relative variability" was introduced. Relative variability is increased by ion channel blockers that decrease the negative feedback control of APD (i.e. blockers of ICa, IKr and IKs) and also by elevation of cytosolic Ca2+. Cardiac arrhythmias are also often categorized according to the characteristic heart rate (tachy- and bradyarrhythmias). Tachycardia is proarrhythmic primarily due to the concomitant Ca2+ overload causing delayed afterdepolarizations. Early afterdepolarizations (EADs) are complications of the bradycardic heart. What is common in the reverse rate-dependent nature of drug action on APD, increased SV and EAD incidence associated with bradycardia.

Adiponectin Decreases Gastric Smooth Muscle Cell Excitability in Mice.

Some adipokines known to regulate food intake at a central level can also affect gastrointestinal motor responses. These are recognized to be peripheral signals able to influence feeding behavior as well. In this view, it has been recently observed that adiponectin (ADPN), which seems to have a role in sending satiety signals at the central nervous system level, actually affects the mechanical responses in gastric strips from mice. However, at present, there are no data in the literature about the electrophysiological effects of ADPN on gastric smooth muscle. To this aim, we achieved experiments on smooth muscle cells (SMCs) of gastric fundus to find out a possible action on SMC excitability and on membrane phenomena leading to the mechanical response. Experiments were made inserting a microelectrode in a single cell of a muscle strip of the gastric fundus excised from adult female mice. We found that ADPN was able to hyperpolarize the resting membrane potential, to enhance the delayed rectifier K+ currents and to reduce the voltage-dependent Ca2+ currents. Our overall results suggest an inhibitory action of ADPN on gastric SMC excitation-contraction coupling. In conclusion, the depressant action of ADPN on the gastric SMC excitability, here reported for the first time, together with its well-known involvement in metabolism, might lead us to consider a possible contribution of ADPN also as a peripheral signal in the hunger-satiety cycle and thus in feeding behavior.

Postpartum hormones oxytocin and prolactin cause pro-arrhythmic prolongation of cardiac repolarization in long QT syndrome type 2.

AIMS: Women with long QT syndrome 2 (LQT2) have a particularly high postpartal risk for lethal arrhythmias. We aimed at investigating whether oxytocin and prolactin contribute to this risk by affecting repolarization. METHODS AND RESULTS: In female transgenic LQT2 rabbits (HERG-G628S, loss of IKr), hormone effects on QT/action potential duration (APD) were assessed (0.2-200 ng/L). Hormone effects (200 ng/L) on ion currents and cellular APD were determined in transfected cells and LQT2 cardiomyocytes. Hormone effects on ion channels were assessed with qPCR and western blot. Experimental data were incorporated into in silico models to determine the pro-arrhythmic potential. Oxytocin prolonged QTc and steepened QT/RR-slope in vivo and prolonged ex vivo APD75 in LQT2 hearts. Prolactin prolonged APD75 at high concentrations. As underlying mechanisms, we identified an oxytocin- and prolactin-induced acute reduction of IKs-tail and IKs-steady (-25.5%, oxytocin; -13.3%, prolactin, P < 0.05) in CHO-cells and LQT2-cardiomyocytes. IKr currents were not altered. This oxytocin-/prolactin-induced IKs reduction caused APD90 prolongation (+11.9%/+13%, P < 0.05) in the context of reduced/absent IKr in LQT2 cardiomyocytes. Hormones had no effect on IK1 and ICa,L in cardiomyocytes. Protein and mRNA levels of CACNA1C/Cav1.2 and RyR2 were enhanced by oxytocin and prolactin. Incorporating these hormone effects into computational models resulted in reduced repolarization reserve and increased propensity to pro-arrhythmic permanent depolarization, lack of capture and early afterdepolarizations formation. CONCLUSIONS: Postpartum hormones oxytocin and prolactin prolong QT/APD in LQT2 by reducing IKs and by increasing Cav1.2 and RyR2 expression/transcription, thereby contributing to the increased postpartal arrhythmic risk in LQT2.

Azologization of serotonin 5-HT3 receptor antagonists.

The serotonin 5-hydroxytryptamine 3 receptor (5-HT3R) plays a unique role within the seven classes of the serotonin receptor family, as it represents the only ionotropic receptor, while the other six members are G protein-coupled receptors (GPCRs). The 5-HT3 receptor is related to chemo-/radiotherapy provoked emesis and dysfunction leads to neurodevelopmental disorders and psychopathologies. Since the development of the first serotonin receptor antagonist in the early 1990s, the range of highly selective and potent drugs expanded based on various chemical structures. Nevertheless, on-off-targeting of a pharmacophore's activity with high spatiotemporal resolution as provided by photopharmacology remains an unsolved challenge bearing additionally the opportunity for detailed receptor examination. In the presented work, we summarize the synthesis, photochromic properties and in vitro characterization of azobenzene-based photochromic derivatives of published 5-HT3R antagonists. Despite reported proof of principle of direct azologization, only one of the investigated derivatives showed antagonistic activity lacking isomer specificity.

Ion Channel Function During Oocyte Maturation and Fertilization.

The proper maturation of both male and female gametes is essential for supporting fertilization and the early embryonic divisions. In the ovary, immature fully-grown oocytes that are arrested in prophase I of meiosis I are not able to support fertilization. Acquiring fertilization competence requires resumption of meiosis which encompasses the remodeling of multiple signaling pathways and the reorganization of cellular organelles. Collectively, this differentiation endows the egg with the ability to activate at fertilization and to promote the egg-to-embryo transition. Oocyte maturation is associated with changes in the electrical properties of the plasma membrane and alterations in the function and distribution of ion channels. Therefore, variations on the pattern of expression, distribution, and function of ion channels and transporters during oocyte maturation are fundamental to reproductive success. Ion channels and transporters are important in regulating membrane potential, but also in the case of calcium (Ca2+), they play a critical role in modulating intracellular signaling pathways. In the context of fertilization, Ca2+ has been shown to be the universal activator of development at fertilization, playing a central role in early events associated with egg activation and the egg-to-embryo transition. These early events include the block of polyspermy, the completion of meiosis and the transition to the embryonic mitotic divisions. In this review, we discuss the role of ion channels during oocyte maturation, fertilization and early embryonic development. We will describe how ion channel studies in Xenopus oocytes, an extensively studied model of oocyte maturation, translate into a greater understanding of the role of ion channels in mammalian oocyte physiology.

Methylene Blue Counteracts H2S-Induced Cardiac Ion Channel Dysfunction and ATP Reduction.

We have previously demonstrated that methylene blue (MB) counteracts the effects of hydrogen sulfide (H2S) cardiotoxicity by improving cardiomyocyte contractility and intracellular Ca2+ homeostasis disrupted by H2S poisoning. In vivo, MB restores cardiac contractility severely depressed by sulfide and protects against arrhythmias, ranging from bundle branch block to ventricular tachycardia or fibrillation. To dissect the cellular mechanisms by which MB reduces arrhythmogenesis and improves bioenergetics in myocytes intoxicated with H2S, we evaluated the effects of H2S on resting membrane potential (Em), action potential (AP), Na+/Ca2+ exchange current (INaCa), depolarization-activated K+ currents and ATP levels in adult mouse cardiac myocytes and determined whether MB could counteract the toxic effects of H2S on myocyte electrophysiology and ATP. Exposure to toxic concentrations of H2S (100 µM) significantly depolarized Em, reduced AP amplitude, prolonged AP duration at 90% repolarization (APD90), suppressed INaCa and depolarization-activated K+ currents, and reduced ATP levels in adult mouse cardiac myocytes. Treating cardiomyocytes with MB (20 µg/ml) 3 min after H2S exposure restored Em, APD90, INaCa, depolarization-activated K+ currents, and ATP levels toward normal. MB improved mitochondrial membrane potential (∆ψm) and oxygen consumption rate in myocytes in which Complex I was blocked by rotenone. We conclude that MB ameliorated H2S-induced cardiomyocyte toxicity at multiple levels: (1) reversing excitation-contraction coupling defects (Ca2+ homeostasis and L-type Ca2+ channels); (2) reducing risks of arrhythmias (Em, APD, INaCa and depolarization-activated K+ currents); and (3) improving cellular bioenergetics (ATP, ∆ψm).

Natriuretic Peptide Receptor-C Protects Against Angiotensin II-Mediated Sinoatrial Node Disease in Mice.

Sinoatrial node (SAN) disease mechanisms are poorly understood, and therapeutic options are limited. Natriuretic peptide(s) (NP) are cardioprotective hormones whose effects can be mediated partly by the NP receptor C (NPR-C). We investigated the role of NPR-C in angiotensin II (Ang II)-mediated SAN disease in mice. Ang II caused SAN disease due to impaired electrical activity in SAN myocytes and increased SAN fibrosis. Strikingly, Ang II treatment in NPR-C-/- mice worsened SAN disease, whereas co-treatment of wild-type mice with Ang II and a selective NPR-C agonist (cANF) prevented SAN dysfunction. NPR-C may represent a new target to protect against the development of Ang II-induced SAN disease.