arrhythmia (mechanisms); calcium; ion channels or ion channel; structural heart disease; CaMKII
Understanding relationships between heart failure and arrhythmias, important causes of suffering and sudden death, remains an unmet goal for biomedical researchers and physicians. Evidence assembled over the last decade supports a view that activation of the multifunctional Ca2+ and calmodulin-dependent protein kinase II (CaMKII) favors myocardial dysfunction and cell membrane electrical instability. CaMKII activation follows increases in intracellular Ca2+ or oxidation, upstream signals with the capacity to transition CaMKII into a Ca2+ and calmodulin-independeant, constitutively active enzyme. Constitutively active CaMKII appears poised to participate in disease pathways by catalyzing the phosphorylation of classes of protein targets important for excitation-contraction coupling and cell survival, including ion channels and Ca2+ homeostatic proteins, and transcription factors that drive hypertrophic and inflammatory gene expression. This rich diversity of downstream targets helps to explain the potential for CaMKII to simultaneously affect mechanical and electrical properties of heart muscle cells. Proof of concept studies from a growing number of investigators show that CaMKII inhibition is beneficial for improving myocardial performance and reducing arrhythmias. Here we review the molecular physiology of CaMKII, discuss CaMKII actions at key cellular targets and results of animal models of myocardial hypertrophy, dysfunction and arrhythmias that suggest CaMKII inhibition may benefit myocardial function while reducing arrhythmias.
CaMKII; Arrhythmias; Heart Failure; Ion channels; Remodeling
Myocardial infarction is a major cause of morbidity and mortality in the developing and developed world. Although current interventions have been successful in prolonging life, they are inadequate because mortality is still high among MI patients. The multifunctional Ca2+/calmodulin dependent protein kinase (CaMKII) plays a key role in the structure and contractility of the myocardium. CaMKII activity is increased in MI hearts and CaMKII promotes cardiac hypertrophy and inflammation, processes consistently activated by myocardial injury. Hypertrophy and inflammation are also related to neurohumoral and redox signaling that uncouple CaMKII activation from Ca2+/calmodulin dependence. Thus, CaMKII may act as a nodal point for integrating hypertrophic and inflammatory signaling in myocardium.
CaMKII; Heart; Myocardium; Myocardial infarction; Heart attack; Inflammation; Hypertrophy; Toll-like receptors; Oxidative stress; ROS; AngII
The multifunctional Ca2+ and calmodulin-dependent protein kinase II (CaMKII) is now recognized to play a central role in pathological events in the cardiovascular system. CaMKII has diverse downstream targets that promote vascular disease, heart failure and arrhythmias, so improved understanding of CaMKII signaling has the potential to lead to new therapies for cardiovascular disease. CaMKII is a multimeric serine-threonine kinase that is initially activated by binding calcified calmodulin (Ca2+/CaM). Under conditions of sustained exposure to elevated Ca2+/CaM CaMKII transitions into a Ca2+/CaM-autonomous enzyme by two distinct but parallel processes. Autophosphorylation of threonine 287 in the CaMKII regulatory domain ‘traps’ CaMKII into an open configuration even after Ca2+/CaM unbinding. More recently, our group identified a pair of methionines (281/282) in the CaMKII regulatory domain that undergo a partially reversible oxidation which, like autophosphorylation, prevents CaMKII from inactivating after Ca2+/CaM unbinding. Here we review roles of CaMKII in cardiovascular disease with an eye to understanding how CaMKII may act as a transduction signal to connect pro-oxidant conditions into specific downstream pathological effects that are relevant to rare and common forms of cardiovascular disease.
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is directly linked to mutations in proteins (e.g., RyR2R4496C) responsible for intracellular Ca2+ homeostasis in the heart. However, the mechanism of Ca2+ release dysfunction underlying CPVT has only been investigated in isolated cells but not in the in situ undisrupted myocardium.
Methods and Results
We investigated in situ myocyte Ca2+ dynamics in intact Langendorff perfused hearts (ex vivo) from wildtype (WT) and RyR2R4496C+/− mice using laser scanning confocal microscopy. We found that myocytes from both WT and RyR2R4496C+/− hearts displayed uniform, synchronized Ca2+ transients. Ca2+ transients from beat to beat were comparable in amplitude with identical activation and decay kinetics in WT and RyR2R4496C+/− hearts, suggesting that excitation-contraction (EC) coupling between the sarcolemmal Ca2+ channels and mutated RyR2R4496C+/− channels remains intact under baseline resting conditions. Upon adrenergic stimulation, RyR2R4496C+/− hearts exhibited a high degree of Ca2+ release variability (CRV). The varied pattern of Ca2+ release was absent in single isolated myocytes, independent of cell cycle length, synchronized among neighboring myocytes, and correlated with CPVT. A similar pattern of action potential variability, which was synchronized among neighboring myocytes, was also revealed under adrenergic stress in intact hearts but not in isolated myocytes.
Our studies using in situ confocal imaging approach suggest that mutated RyR2s are functionally normal at rest but display a high degree of CRV upon intense adrenergic stimulation. CRV is a Ca2+ release abnormality resulting from electrical defects rather than the failure of the Ca2+ release response to action potentials in mutated ventricular myocytes. Our data provide important insights into Ca2+ release and electrical dysfunction in an established model of CPVT.
arrhythmia (mechanisms); calcium; catecholaminergic polymorphic ventricular tachycardia; sarcoplasmic reticulum; ryanodine receptors
Many signals have risen and fallen in the tide of investigation into mechanisms of myocardial hypertrophy and heart failure (HF). In our opinion, the multifunctional Ca and calmodulin-dependent protein kinase II (CaMKII) has emerged as a molecule to watch, in part because a solid body of accumulated data essentially satisfy Koch's postulates, showing that the CaMKII pathway is a core mechanism for promoting myocardial hypertrophy and heart failure. Multiple groups have now confirmed the following: (1) that CaMKII activity is increased in hypertrophied and failing myocardium from animal models and patients; (2) CaMKII overexpression causes myocardial hypertrophy and HF and (3) CaMKII inhibition (by drugs, inhibitory peptides and gene deletion) improves myocardial hypertrophy and HF. Patients with myocardial disease die in equal proportion from HF and arrhythmias, and a major therapeutic obstacle is that drugs designed to enhance myocardial contraction promote arrhythmias. In contrast, inhibiting the CaMKII pathway appears to reduce arrhythmias and improve myocardial responses to pathological stimuli. This brief paper will introduce the molecular physiology of CaMKII and discuss the impact of CaMKII on ion channels, Ca handling proteins and transcription in myocardium. This article is part of a Special Issue entitled “Key Signaling Molecules Special Issue”.
Calmodulin kinase II; Cell signaling; Oxidation; Hypertrophy; Heart failure; Arrhythmias
Electrical and structural remodeling during the progression of cardiovascular disease is associated with adverse outcomes subjecting affected patients to overt heart failure (HF) and/or sudden death. Dysfunction in integral membrane protein trafficking has long been linked with maladaptive electrical remodeling. However, little is known regarding the molecular identity or function of these intracellular targeting pathways in the heart. Eps15 homology domain-containing (EHD) gene products (EHD1-4) are polypeptides linked with endosomal trafficking, membrane protein recycling, and lipid homeostasis in a wide variety of cell types. EHD3 was recently established as a critical mediator of membrane protein trafficking in the heart. Here, we investigate the potential link between EHD3 function and heart disease. Using four different HF models including ischemic rat heart, pressure overloaded mouse heart, chronic pacing-induced canine heart, and non-ischemic failing human myocardium we provide the first evidence that EHD3 levels are consistently increased in HF. Notably, the expression of the Na/Ca exchanger (NCX1), targeted by EHD3 in heart is similarly elevated in HF. Finally, we identify a molecular pathway for EHD3 regulation in heart failure downstream of reactive oxygen species and angiotensin II signaling. Together, our new data identify EHD3 as a previously unrecognized component of the cardiac remodeling pathway.
EHD proteins; heart failure; cardiac remodeling; regulation; animal models
Patients with systolic left ventricular dysfunction die progressively from congestive heart failure or die suddenly from cardiac arrhythmias. Myocardial hypertrophy is an early event in most forms of heart failure, but the majority of patients with myocardial hypertrophy do not develop heart failure. Developing improved therapies for targeting the cell signaling pathways that enable this deadly transition from early myocardial insult to heart failure and sudden death is a key goal for improving public health. In this issue of the JCI, Ling and colleagues provide new evidence that activation of the multifunctional Ca2+/calmodulin–dependent kinase IIδ is a decisive step on the path to heart failure in mice (see the related article beginning on page 1230).
Significance: In heart failure (HF), contractile dysfunction and arrhythmias result from disturbed intracellular Ca handling. Activated stress kinases like cAMP-dependent protein kinase A (PKA), protein kinase C (PKC), and Ca/calmodulin-dependent protein kinase II (CaMKII), which are known to influence many Ca-regulatory proteins, are mechanistically involved. Recent Advances: Beside classical activation pathways, it is becoming increasingly evident that reactive oxygen species (ROS) can directly oxidize these kinases, leading to alternative activation. Since HF is associated with increased ROS generation, ROS-activated serine/threonine kinases may play a crucial role in the disturbance of cellular Ca homeostasis. Many of the previously described ROS effects on ion channels and transporters are possibly mediated by these stress kinases. For instance, ROS have been shown to oxidize and activate CaMKII, thereby increasing Na influx through voltage-gated Na channels, which can lead to intracellular Na accumulation and action potential prolongation. Consequently, Ca entry via activated NCX is favored, which together with ROS-induced dysfunction of the sarcoplasmic reticulum can lead to dramatic intracellular Ca accumulation, diminished contractility, and arrhythmias. Critical Issues: While low amounts of ROS may regulate kinase activity, excessive uncontrolled ROS production may lead to direct redox modification of Ca handling proteins. Therefore, depending on the source and amount of ROS generated, ROS could have very different effects on Ca-handling proteins. Future Directions: The discrimination between fine-tuned ROS signaling and unspecific ROS damage may be crucial for the understanding of heart failure development and important for the investigation of targeted treatment strategies. Antioxid. Redox Signal. 18, 1063–1077.
Diabetes increases oxidant stress and doubles the risk of dying after myocardial
infarction, but the mechanisms underlying increased mortality are unknown. Mice with
streptozotocin-induced diabetes developed profound heart rate slowing and doubled
mortality compared with controls after myocardial infarction. Oxidized
Ca2+/calmodulin-dependent protein kinase II (ox-CaMKII) was
significantly increased in pacemaker tissues from diabetic patients compared with
that in nondiabetic patients after myocardial infarction. Streptozotocin-treated mice
had increased pacemaker cell ox-CaMKII and apoptosis, which were further enhanced by
myocardial infarction. We developed a knockin mouse model of oxidation-resistant
CaMKIIδ (MM-VV), the isoform associated with cardiovascular disease.
Streptozotocin-treated MM-VV mice and WT mice infused with MitoTEMPO, a mitochondrial
targeted antioxidant, expressed significantly less ox-CaMKII, exhibited increased
pacemaker cell survival, maintained normal heart rates, and were resistant to
diabetes-attributable mortality after myocardial infarction. Our findings suggest
that activation of a mitochondrial/ox-CaMKII pathway contributes to increased sudden
death in diabetic patients after myocardial infarction.
Timothy Syndrome (TS) is a disease of excessive cellular Ca2+ entry and life-threatening arrhythmias due to a mutation in the primary cardiac L-type Ca2+ channel (CaV1.2). The TS mutation causes loss of normal voltage-dependent inactivation (VDI) of CaV1.2 current (ICa). During cellular Ca2+ overload the calmodulin-dependent protein kinase II (CaMKII) causes arrhythmias. We hypothesized that CaMKII is a part of the proarrhythmic mechanism in TS.
Methods and Results
We developed an adult rat ventricular myocyte model of TS (G406R) by lenti virus-mediated transfer of wild type (WT) and TS CaV1.2. The exogenous CaV1.2 contained a mutation (T1066Y) conferring dihydropyridine resistance, so we could silence endogenous CaV1.2 with nifedipine and maintain peak ICa at control levels in infected cells. TS CaV1.2 infected ventricular myocytes exhibited the signature VDI loss under Ca2+ buffering conditions, not permissive for CaMKII activation. In physiological Ca2+ solutions, TS CaV1.2 expressing ventricular myocytes exhibited increased CaMKII activity and a proarrhythmic phenotype that included action potential prolongation, increased ICa facilitation and afterdepolarizations. Intracellular dialysis of a CaMKII inhibitory peptide, but not a control peptide, reversed increases in ICa facilitation, normalized the action potential and prevented afterdepolarizations. We developed a revised mathematical model that accounts for CaMKII-dependent and CaMKII-independent effects of the TS mutation.
In TS the loss of VDI is an upstream initiating event for arrhythmia phenotypes that are ultimately dependent on CaMKII activation.
action potentials; calcium; ion channels; myocytes
Right ventricular failure (RVF) is the main cause of death in patients with pulmonary artery hypertension (PAH). Sildenafil, a phosphodiesterase type 5 (PDE5) inhibitor, was recently approved for treatment of PAH patients. However, the mechanisms underlying RV contractile malfunction and the benefits of sildenafil on RV function are not well understood. We aimed to investigate 1) the ultrastructural and excitation-contraction coupling alterations underlying PAH-induced RVF; 2) whether the ultrastructural changes are reversible; 3) the mechanisms underlying the therapeutic benefits of sildenafil in PAH-RVF. We used a single injection of monocrotaline (MCT) in Wistar rats to induce pulmonary vascular proliferation, which led to PAH and RVF. RV myocytes displayed severe T-tubule loss and disorganization as well as blunted and dys-synchronous SR Ca2+ release. Sildenafil prevented and reversed the MCT-induced PAH and LV filling impairment. Early intervention with sildenafil prevented RV hypertrophy and the development of RVF, T-tubule remodeling and Ca2+ handling dysfunction. While late treatment with sildenafil did not reverse RV hypertrophy in animals with established RVF, RV systolic function was improved. Furthermore, late intervention partially reversed both the impairment of myocyte T-tubule integrity and Ca2+ handling protein and SR Ca2+ release function in MCT-treated rats. In conclusion, PAH-induced increase in RV afterload causes severe T-tubule remodeling and Ca2+ handling dysfunction in RV myocytes, leading to RV contractile failure. Sildenafil prevents and partially reverses ultrastructural, molecular and functional remodeling of failing RV myocytes. Reversal of pathological T-tubule remodeling, although incomplete, is achievable without the regression of RV hypertrophy.
right ventricle failure; pulmonary artery hypertension; PDE5 inhibitor; calcium; t-tubule; excitation-contraction coupling
The cardiovascular system operates under demands ranging from conditions of rest to extreme stress. One mechanism of cardiac stress tolerance is action potential duration shortening driven by ATP-sensitive potassium (KATP) channels. KATP channel expression has a significant physiologic impact on action potential duration shortening and myocardial energy consumption in response to physiologic heart rate acceleration. However, the effect of reduced channel expression on action potential duration shortening in response to severe metabolic stress is yet to be established. Here, transgenic mice with myocardium-specific expression of a dominant negative KATP channel subunit were compared with littermate controls. Evaluation of KATP channel whole cell current and channel number/patch was assessed by patch clamp in isolated ventricular cardiomyocytes. Monophasic action potentials were monitored in retrogradely perfused, isolated hearts during the transition to hypoxic perfusate. An 80-85% reduction in cardiac KATP channel current density results in a similar magnitude, but significantly slower rate, of shortening of the ventricular action potential duration in response to severe hypoxia, despite no significant difference in coronary flow. Therefore, the number of functional cardiac sarcolemmal KATP channels is a critical determinant of the rate of adaptation of myocardial membrane excitability, with implications for optimization of cardiac energy consumption and consequent cardioprotection under conditions of severe metabolic stress.
ATP-sensitive potassium channel; K-ATP; heart; glyburide; monophasic action potential
Doxorubicin (DOX) is one of the most effective chemotherapeutic agents, but cardiotoxicity limits DOX therapy. Although the mechanisms are not entirely understood, reactive oxygen species (ROS) appear to be involved in DOX cardiotoxicity. Ca/calmodulin dependent protein kinase II (CaMKII) can be activated by ROS through oxidation and is known to contribute to myocardial dysfunction through Ca leakage from the sarcoplasmic reticulum (SR).
We hypothesized that CaMKII contributes to DOX-induced defects in intracellular Ca ([Ca]i) handling.
Cardiac myocytes were isolated from wild-type (WT) adult rat hearts and from mouse hearts lacking the predominant myocardial CaMKII isoform (CaMKIIδ−/−, KO) vs. WT. Isolated cardiomyocytes were investigated 30 min after DOX (10 µmol/L) superfusion, using epifluorescence and confocal microscopy. Intracellular ROS-generation ([ROS]i) and [Ca]i handling properties were assessed. In a subset of experiments, KN-93 or AIP (each 1 µmol/L) were used to inhibit CaMKII. Melatonin (Mel, 100 µmol/L) served as ROS-scavenger. Western blots were performed to determine the amount of CaMKII phosphorylation and oxidation.
DOX increased [ROS]i and led to significant diastolic [Ca]i overload in rat myocytes. This was associated with reduced [Ca]i transients, a 5.8-fold increased diastolic SR Ca leak and diminished SR Ca content. ROS-scavenging partially rescued Ca handling. Western blots revealed increased CaMKII phosphorylation, but not CaMKII oxidation after DOX. Pharmacological CaMKII inhibition attenuated diastolic [Ca]i overload after DOX superfusion and led to partially restored [Ca]i transients and SR Ca content, presumably due to reduced Ca spark frequency. In line with this concept, isoform-specific CaMKIIδ-KO attenuated diastolic [Ca]i overload and Ca spark frequency.
DOX exposure induces CaMKII-dependent SR Ca leakage, which partially contributes to impaired cellular [Ca]i homeostasis. Pharmacological and genetic CaMKII inhibition attenuated but did not completely abolish the effects of DOX on [Ca]i. In light of the clinical relevance of DOX, further investigations seem appropriate to determine if CaMKII inhibition could reduce DOX-induced cardiotoxicity.
Physical activity is one of the most important determinants of cardiac function. The ability of the heart to increase delivery of oxygen and metabolic fuels relies on an array of adaptive responses necessary to match bodily demand while avoiding exhaustion of cardiac resources. The ATP-sensitive potassium (KATP) channel has the unique ability to adjust cardiac membrane excitability in accordance with ATP and ADP levels, and up-regulation of its expression that occurs in response to exercise could represent a critical element of this adaption. However, the mechanism by which KATP channel expression changes result in a beneficial effect on cardiac excitability and function remains to be established. Here, we demonstrate that an exercise-induced rise in KATP channel expression enhanced the rate and magnitude of action potential shortening in response to heart rate acceleration. This adaptation in membrane excitability promoted significant reduction in cardiac energy consumption under escalating workloads. Genetic disruption of normal KATP channel pore function abolished the exercise-related changes in action potential duration adjustment and caused increased cardiac energy consumption. Thus, an expression-driven enhancement in the KATP channel-dependent membrane response to alterations in cardiac workload represents a previously unrecognized mechanism for adaptation to physical activity and a potential target for cardioprotection.
KATP; K-ATP; remodeling; oxygen consumption; heart rate; exercise
Approximately half of patients with heart failure die suddenly as a result of ventricular arrhythmias. Although abnormal Ca2+ release from the sarcoplasmic reticulum (SR) through ryanodine receptors (RyR2) has been linked to arrhythmogenesis, the molecular mechanisms triggering release of arrhythmogenic Ca2+ remain unknown. We tested the hypothesis that increased RyR2 phosphorylation by Ca2+/calmodulin-dependent protein kinase II (CaMKII) is both necessary and sufficient to promote lethal ventricular arrhythmias.
Methods and Results
Mice in which the S2814 CaMKII site on RyR2 is constitutively activated (S2814D) develop pathological SR Ca2+ release events resulting in reduced SR Ca2+ load on confocal microscopy. These Ca2+ release events are associated with increased RyR2 open probability in lipid bilayer preparations. At baseline, young S2814D mice have structurally and functionally normal hearts without arrhythmias; however, they develop sustained ventricular tachycardia and sudden cardiac death upon catecholaminergic provocation by caffeine/epinephrine or programmed electrical stimulation. Young S2814D mice have a significant predisposition to sudden arrhythmogenic death after transverse aortic constriction (TAC) surgery. Finally, genetic ablation of the CaMKII site on RyR2 (S2814A) protects mutant mice from pacing-induced arrhythmias versus wild type mice after TAC surgery.
Our results suggest that CaMKII phosphorylation of RyR2 Ca2+ release channels at S2814 plays an important role in arrhythmogenesis and sudden cardiac death in mice with heart failure.
Cardiac arrhythmias; Ca2+/calmodulin kinase II; heart failure; ryanodine receptor; sarcoplasmic reticulum
Dilated cardiomyopathy (DCM), characterized by dilatation and dysfunction of the left ventricle, is an important cause of heart failure. Many mutations in various genes, including cytoskeletal protein genes and contractile protein genes, have been identified in DCM patients, but the mechanisms of how such mutations lead to DCM remain unknown.
Methods and Results
We established the mouse model of DCM by expressing a mutated cardiac α-actin gene, which has been reported in patients with DCM, in the heart (mActin-Tg). mActin-Tg mice showed gradual dilatation and dysfunction of the left ventricle, resulting in death by heart failure. The number of apoptotic cardiomyocytes and protein levels of p53 were increased in the hearts of mActin-Tg mice. Overexpression of Bcl-2 or downregulation of p53 decreased the number of apoptotic cardiomyocytes and improved cardiac function. This mouse model showed a decrease in myofilament calcium sensitivity and activation of calcium/calmodulin-dependent kinase IIδ (CaMKIIδ). The inhibition of CaMKIIδ prevented the increase in p53 and apoptotic cardiomyocytes and ameliorated cardiac function.
CaMKIIδ plays a critical role in the development of heart failure in part by accumulation of p53 and induction of cardiomyocyte apoptosis in the DCM mouse model.
apoptosis; CaMKII; cardiomyopathy; heart failure; genes; p53
The transverse tubule (t-tubule) system is the ultrastructural substrate for excitation-contraction coupling in ventricular myocytes; t-tubule disorganization and loss are linked to decreased contractility in end stage heart failure (HF).
To examine 1) if pathological t-tubule remodeling occurs early in compensated hypertrophy and, if so, how it evolves during the transition from hypertrophy to HF; 2) the role of junctophilin-2 in t-tubule remodeling.
Methods and Results
We investigated t-tubule remodeling in relation to ventricular function during HF progression using state-of-the-art confocal imaging of t-tubules in intact hearts, using a thoracic aortic banding (TAB) rat HF model. We developed a quantitative t-tubule power (TTpower) index to represent the integrity of t-tubule structure. We found that discrete local loss and global reorganization of the t-tubule system (leftward shift of TTpower histogram) started early in compensated hypertrophy in left ventricular (LV) myocytes, prior to LV dysfunction, as detected by echocardiography. With progression from compensated hypertrophy to early and late HF, t-tubule remodeling spread from the LV to the right ventricle (RV), and TTpower histograms of both ventricles gradually shifted leftward. The mean LV TTpower showed a strong correlation with ejection fraction and heart weight to body weight ratio. Over the progression to HF we observed a gradual reduction in the expression of a junctophilin protein (JP-2) implicated in the formation of t-tubule / sarcoplasmic reticulum junctions. Furthermore, we found that JP-2 knockdown by gene silencing reduced t-tubule structure integrity in cultured adult ventricular myocytes.
T-tubule remodeling in response to TAB stress begins prior to echocardiographically detectable LV dysfunction and progresses over the development of overt structural heart disease. LV t-tubule remodeling is closely associated with the severity of cardiac hypertrophy and predicts LV function. Thus, t-tubule remodeling may constitute a key mechanism underlying the transition from compensated hypertrophy to HF.
t-tubule; myocardial remodeling; hypertrophy; heart failure; confocal microscopy
Increasing evidence suggests that cardiac pacemaking is the result of two sinoatrial node (SAN) cell mechanisms: a ‘voltage clock’ and a Ca2+ dependent process, or ‘Ca2+ clock.’ The voltage clock initiates action potentials (APs) by SAN cell membrane potential depolarization from inward currents, of which the pacemaker current (If) is thought to be particularly important. A Ca2+ dependent process triggers APs when sarcoplasmic reticulum (SR) Ca2+ release activates inward current carried by the forward mode of the electrogenic Na+/Ca2+ exchanger (NCX). However, these mechanisms have mostly been defined in rodents or rabbits, but are unexplored in single SAN cells from larger animals. Here, we used patch-clamp and confocal microscope techniques to explore the roles of the voltage and Ca2+ clock mechanisms in canine SAN pacemaker cells. We found that ZD7288, a selective If antagonist, significantly reduced basal automaticity and induced irregular, arrhythmia-like activity in canine SAN cells. In addition, ZD7288 impaired but did not eliminate the SAN cell rate acceleration by isoproterenol. In contrast, ryanodine significantly reduced the SAN cell acceleration by isoproterenol, while ryanodine reduction of basal automaticity was modest (∼14%) and did not reach statistical significance. Importantly, pretreatment with ryanodine eliminated SR Ca2+ release, but did not affect basal or isoproterenol-enhanced If. Taken together, these results indicate that voltage and Ca2+ dependent automaticity mechanisms coexist in canine SAN cells, and suggest If and SR Ca2+ release cooperate to determine baseline and catecholamine-dependent automaticity in isolated dog SAN cells.
sinoatrial node cells; action potentials; funny current; sarcoplasmic reticulum; pacemaker