Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts

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Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts Toshiro Kajii a, Shigeki Kobayashi a, Sho Shiba a, Shohei Fujii a, Masaki Tamitani a, Michiaki Kohno a, Yoshihide Nakamura a, Takuma Nanno a, Takayoshi Kato a, Shinichi Okuda a, Hitoshi Uchinoumi a, Tetsuro Oda a, Takeshi Yamamoto b, Masafumi Yano a, * a

Department of Medicine and Clinical Science, Division of Cardiology, Yamaguchi University Graduate School of Medicine, 1-1-1 Minamikogushi, Ube, Yamaguchi, 755-8505, Japan b Faculty of Health Sciences, Yamaguchi University Graduate School of Medicine, Ube, Japan

a r t i c l e i n f o

a b s t r a c t

Article history: Received 20 September 2019 Accepted 8 October 2019 Available online xxx

Aberrant Ca2þ release from cardiac ryanodine receptors (RyR2) has been shown to be one of the most important causes of lethal arrhythmia in various types of failing hearts. We previously showed that dantrolene, a specific agent for the treatment of malignant hyperthermia, inhibits Ca2þ leakage from the RyR2 by correcting the defective inter-domain interaction between the N-terminal (1e619 amino acids) and central (2000e2500 amino acids) domains of the RyR2 and allosterically enhancing the binding affinity of calmodulin to the RyR2 in diseased hearts. In this study, we examined whether dantrolene inhibits this Ca2þ leakage, thereby preventing the pharmacologically inducible ventricular tachycardia in ventricular pressure-overloaded failing hearts. Ventricular tachycardia (VT) was easily induced after an injection of epinephrine in mice after 8 weeks of transverse aortic constriction-induced pressure-overload. Pretreatment with dantrolene almost completely inhibited the pharmacologically inducible VT. In the presence of dantrolene, the occurrence of both Ca2þ sparks and spontaneous Ca2þ transients was inhibited, which was associated with enhanced calmodulin binding affinity to the RyR2. These results suggest that dantrolene could be a new potent agent in the treatment of lethal arrhythmia in cases of acquired heart failure. © 2019 Elsevier Inc. All rights reserved.

Keywords: Ryanodine receptor Sarcoplasmic reticulum Calcium Calmodulin Heart failure Ventricular tachycardia

1. Introduction Cardiac hypertrophy is a major risk factor for various cardiovascular diseases and sudden cardiac death. Recently, Ca2þ leakage from cardiac ryanodine receptors (RyR2) has been shown to be involved in the development of cardiac hypertrophy and its progression towards heart failure [1,2], and the inhibition of Ca2þ leakage leads to attenuation of cardiac hypertrophy [3]. We previously showed that defective inter-domain interaction between the N-terminal (1e619 amino acids) and central (2000e2500 amino acids) domains of the RyR2, called domain unzipping, causes Ca2þ to leak out from the RyR2, resulting in heart failure and catecholaminergic polymorphic ventricular tachycardia (CPVT) [4e6].

* Corresponding author. E-mail address: [email protected] (M. Yano).

Furthermore, we previously reported that domain unzipping causes the Ca2þ channel to become leaky via the dissociation of calmodulin (CaM) from the RyR2 during heart failure [7,8] and CPVT [9]. In addition, we recently reported that the aberrant Ca2þ release from the RyR2 in pressure overload-induced failing hearts, which is mediated by the defective interaction of calmodulin (CaM) with the RyR2, plays a critical role in the pathogenesis of lethal arrhythmia, and further that enhancing the binding affinity of CaM to the RyR2 prevented aberrant Ca2þ release [10]. Additionally, the occurrence of arrhythmogenic substrates observed in CPVTassociated cardiomyocytes during delayed afterdepolarization and triggered activity has been shown to be inhibited by increasing the binding affinity of CaM to the RyR2 [11]. Dantrolene is a specific agent used for treating malignant hyperthermia (MH), which is a disease caused by a single-amino acid mutation of the skeletal RyR1. Dantrolene has also been shown to inhibit the leakage of Ca2þ from the diseased RyR2 during CPVT or

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Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071

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heart failure by shifting the interaction mode between the N-terminal and central domains from unzipping to zipping, and hence allosterically increasing the binding affinity of CaM to the RyR2 [6e9]. Based on these previous findings, herein, we evaluated the effects of dantrolene on inducible VT and aberrant Ca2þ release from the RyR2 in a mouse model of chronic left ventricular pressure overload induced by transverse aortic constriction (TAC). 2. Materials and methods 2.1. Mice This study conformed to the Guide for the Care and Use of Laboratory Animals published by the U.S. National Institutes of Health (NIH Publication No. 85-23, revised 1996). The care of the animals and the protocols used were in accordance with the guidelines laid down by the Animal Ethics Committee of the Yamaguchi University School of Medicine. Mice were anesthetized in an induction chamber with 2% isoflurane mixed with 0.5e1.0 L/min of 100% O2, as described previously [6]. The transverse aorta was identified and then was tied with a 27½-gauge blunt needle using 7-0 silk to yield a constriction 0.4 mm in diameter to achieve TAC. 2.2. Surface electrocardiogram (ECG) ECG was monitored using ECG telemetry (Data Sciences International, St. Paul, MN), as described previously [12]. A surface ECG was performed at baseline and for 30 min after an intraperitoneal injection of epinephrine (2 mg/kg of body weight). Three mice were treated with dantrolene i.p. (DAN, 40 mg/kg/day) once a day for 3 days before receiving the injection of epinephrine. 2.3. Isolation of cardiac cardiomyocytes Cardiomyocytes were isolated from the hearts of the treated mice, as described previously [6]. Briefly, after the mice were anesthetized and sacrificed, the left ventricle (LV) myocardium was minced with scissors in a fresh buffer containing collagenase, and then the rod-shaped adult mice cardiomyocytes were prepared by retrograde perfusion of the heart with 95% O2 into which 5% CO2 was bubbled. The isolated mice cardiomyocytes were transferred to laminin-coated glass culture dishes and incubated at 37  C in a 5% CO2 and 95% O2 atmosphere. 2.4. Analysis of Ca2þ sparks and sarcoplasmic reticulum (SR) Ca2þ content The frequency of occurrence of Ca2þ sparks was measured using a laser-scanning confocal microscope system (LSM-510, Carl Zeiss), as described previously [6]. Briefly, intact cardiomyocytes were loaded with fluo-4 AM (20 mM; Molecular Probes, OR, USA) for 20 min at room temperature. The microscope system was used in line-scan mode, wherein a single cardiomyocyte was scanned repeatedly (520.8 Hz) along a line parallel to the longitudinal axis, avoiding nuclei. To monitor Ca2þ sparks, cardiomyocytes were stimulated until the Ca2þ transient frequency reached a steady state, and then stimulation was stopped and Ca2þ sparks were recorded during the subsequent ~10 s of rest. Data were analyzed with SparkMaster, an automated analysis program that allows for rapid and reliable spark analysis [6]. To assess SR Ca2þ content, caffeine (10 mM) was rapidly perfused into the cardiomyocytes to discharge their SR-loaded Ca2þ, as described previously [6].

2.5. Monitoring of Ca2þ transients in cardiomyocytes Isolated ventricular myocytes were incubated with 20 mM fluo-4 acetoxymethyl ester for 20 min at room temperature and washed twice with Tyrode’s solution. All experiments were conducted at 28  C. Measurements of intracellular Ca2þ in cells stimulated by a field electric stimulator (IonOptix, MA, USA) were performed with fluorescent digital microscopy (BZ9000, Keyence, Japan). The relative occurrence of spontaneous Ca2þ releases upon cessation of stimulation at frequencies of 1, 2, 3, 4, and 5 Hz was then measured in intact cardiomyocytes [6]. 2.6. Determination of the binding affinity of endogenous CaM to the RyR2 in intact cardiomyocytes Isolated cardiomyocytes were plated on glass-based dishes, fixed with 4% paraformaldehyde in phosphate-buffered saline (PBS) for 5 min, washed three times with PBS, and permeabilized in 0.5% Triton X-100 and 1% bovine serum albumin (BSA) for 20 min, as previously described [12]. These cardiomyocytes were then incubated overnight at 4  C with a monoclonal mouse anti-CaM antibody (EP799Y, Abcam) and anti-RyR antibody (Sigma-Aldrich, C333) in 1% BSA and 0.5% Triton X-100, followed by labeling each of these with an Alexa488-conjugated goat anti-rabbit secondary antibody (Molecular Probes) and Alexa633-conjugated goat antimouse secondary antibody (Molecular Probes), respectively. Cardiomyocytes were then washed three times with PBS. For colocalization of the anti-CaM antibody (Alexa488; green) and antiRyR antibody (Alexa633; red), images were taken at 633 nm and 488 nm excitation in conjunction with an LP 650 filter and a BP 505e530 nm, respectively. Both secondary antibodies showed no detectable fluorescence patterns, and did not cross-react with either primary antibody. In the co-localization analysis, fluorescent images were taken sequentially to prevent spectral cross-talk. 2.7. Determination of the binding affinity of exogenous CaM to the RyR2 in saponin-permeabilized cardiomyocytes CaM fluorescently labeled (F-CaM) with HiLyte Fluor 647 (AnaSpec Inc., CA, USA) was introduced into saponin-permeabilized cardiomyocytes, as described previously [10]. Then, the localized distribution of CaM was determined by making densitometric measurements of F-CaM fluorescence. Briefly, the fluorescently labeled cardiomyocytes were laser-scanned with the confocal microscope (LSM-510, Carl Zeiss) (with numerical aperture ¼ 1.3, excitation at 633 nm, and emission at 640 nm). The sarcomererelated periodical increase in the HiLyte Fluor 647 fluorescence intensity from baseline was integrated with respect to the selected longitudinal distance (~25 mm), and then the measured value was divided by the distance. 2.8. Statistical analyses Unpaired t-tests were used for statistical comparisons of data obtained during the two different situations, whereas one-way ANOVA with a post hoc Tukey’s test was used for statistical comparison of more than two groups. All data are expressed as mean ± SEM. A probability value of less than 0.05 was considered statistically significant. 3. Results 3.1. Dantrolene inhibits pharmacologically inducible VT The application of TAC for 8 weeks increased the size of the left

Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071

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ventricular chamber and decreased the extent of its systolic shortening (Fig. 1A). Dilated LV end-diastolic dimension (LVDD), dilated LV end-systolic dimension (LVDS) and decreased LV fractional shortening (FS) were confirmed in TAC-treated mice as compared with sham-operated mice(FS: 33.6 ± 1.6% in Sham vs

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15.08 ± 2.1% in TAC, P < 0.001; LVDD: 3.00 ± 0.08 mm in Sham vs 4.85 ± 0.22 mm in TAC, P < 0.001; LVDS: 2.00 ± 0.04 mm in Sham vs 3.88 ± 0.14 mm in TAC, P < 0.001) (Fig. 1B) The intraperitoneal injection of epinephrine induced sustained or incessant VT in TACtreated mice. (Fig. 1C and D). In contrast, VT was not

Fig. 1. Echocardiograms and surface ecocardiograms (ECGs) of sham- and TAC-treated mice. (A) Representative echocardiograms of sham- and TAC-treated mice. (B) Summarized data obtained from echocardiograms. (C) Representative ECG recordings of sham- and TAC-treated mice after an intraperitoneal injection of epinephrine (2 mg/kg body weight) Dantrolene (DAN) was intraperitoneally injected to TAC-treated mice once a day for 3 days before receiving the injection of epinephrine. The experiments were performed twice in each mouse. (D).Durations of VT and numbers of VT episodes observed during the 5 min period, without (D-) or with (Dþ) pretreatment of dantrolene, in TAC-treated mice. Data are presented as the results of six experiments from 3 TAC-treated mice. TAC ¼ transverse aortic constriction.

Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071

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A Sham

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Fig. 2. Acute effect of dantrolene on Ca2þ sparks and SR Ca2þ content in TAC-treated cardiomyocytes. (A) Representative line-scan images of Ca2þ sparks in sham- and TACtreated cardiomyocytes, in the presence or absence of dantrolene (1 mM). Summary of (B) spark frequency (SpF) and (C) SR Ca2þ content in 11 cells from sham mice (n ¼ 3) and in 10e12 cells from TAC-treated mice (n ¼ 3), in the presence or absence of dantrolene (1 mM). Data are shown as mean ± SEM values.

Fig. 3. Acute effect of dantrolene on the occurrence of spontaneous Ca2þ transients upon cessation of stimulation at 1, 2, 3, 4, and 5 Hz in TAC-treated cardiomyocytes. (A) Representative records of intracellular Ca2þ transients during pacing in sham- and TAC-treated cardiomyocytes, in the presence or absence of dantrolene (1 mM). (B) Summarized percent (%) occurrence of spontaneous Ca2þ transient (SCaT) events in sham- and TAC-treated cardiomyocytes, in the presence or absence of dantrolene (1 mM). Data represent the mean ± SEM values of 230 cells from sham- (n ¼ 3) and 117e192 cells from TAC-treated mice (n ¼ 3).

Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071

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pharmacologically induced in dantrolene-pretreated TAC mice (Fig. 1C and D). 3.2. Dantrolene inhibits Ca2þ sparks and spontaneous Ca2þ transients in pressure-overloaded failing cardiomyocytes To clarify whether the aforementioned anti-arrhythmic effect of dantrolene is attributable to its inhibitory effect on aberrant Ca2þ

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release from the RyR2, we examined the effect of dantrolene on the occurrence of spontaneous Ca2þ release events (i.e., spontaneous Ca2þ sparks and transients). The baseline Ca2þ spark frequency (SpF) increased significantly in cardiomyocytes from TAC-treated mice (Fig. 2). However, in the presence of dantrolene (1 mM), the SpF decreased significantly. The SR Ca2þ content markedly decreased in cardiomyocytes from TAC-treated mice, whereas it increased in the presence of dantrolene (1 mM), suggesting that the

Fig. 4. Acute effect of dantrolene on characteristics of CaM binding to the RyR2 in sham- and TAC-treated cardiomyocytes. (A) Representative images of endogenous CaM detected using anti-CaM antibody and co-localized with RyR2. Left: CaM (green); right: RyR2 (red). (B) Summarized fluorescence signal of endogenous CaM (divided by that of the RyR2), expressed as ratios relative to control values. Data are shown as the mean ± SEM values of 15 cells from sham-treated mice (n ¼ 3) and 14e15 cells from TAC-treated mice (n ¼ 3). (C) Characteristics of binding by the exogenously introduced HiLyte fluorescently labeled CaM (HiLyte-CaM) with the RyR2 in sham- and TAC-treated saponin-permeabilized cardiomyocytes {- or þ DAN (1 mM)}. (D) Summarized binding curves. HiLyte-CaM fluorescence was measured and expressed as the ratio of the observed fluorescence to its maximum value. Binding curves were derived using Hill equipment. Data are shown as mean ± SEM values. Binding curves were obtained from 9 to 15 cells from sham-treated mice (n ¼ 3) and 9e15 cells from TAC-treated mice (n ¼ 3). Values of dissociation constant (Kd) are compared in the bar graph insets.

Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071

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aberrant Ca2þ release from the RyR2 was partly involved in the decreased SR Ca2þ content observed. Spontaneous Ca2þ transients were virtually absent in cardiomyocytes from sham-treated mice (Fig. 3). In contrast, spontaneous Ca2þ transients occurred in cardiomyocytes from TAC-treated mice when the pacing rate increased from 1 to 5 Hz (Fig. 3). However, these spontaneous Ca2þ transients disappeared in the presence of dantrolene (Fig. 3). 3.3. Dantrolene restores normal CaM binding affinity to the RyR2 in pressure-overloaded failing cardiomyocytes We previously showed that the decreased binding affinity of CaM to the RyR2 in TAC-treated cardiomyocytes is the cause of aberrant Ca2þ release events from the RyR2 [10]. Herein, we further examined whether dantrolene ameliorates the decreased binding of CaM to the RyR2. Confirming our previous observations [9,10], endogenous CaM co-localized well with the RyR2 on the Zline(Fig. 4A). In resting cardiac ventricular myocytes, only ~1% of the total CaM is freely diffusible [13] and the bound CaM is highly concentrated along the Z-line, where more than 90% of the CaM is bound to the RyR2 [14]. Therefore, most of the CaM bound to the Zline can be presumed to be bound to the RyR2. The amount of endogenous CaM bound to the Z-line decreased in TAC-treated cardiomyocytes, whereas its levels were restored to baseline in the presence of dantrolene (Fig. 4B). To further examine the effect of dantrolene on the kinetics of CaM binding to the RyR2 in TAC-treated cardiomyocytes, we assessed the amount of CaM binding to different sub-cellular fractions by introducing CaM, fluorescently labeled with HiLyte Fluor 647 (F-CaM), into saponin-permeabilized cardiomyocytes (Fig. 4C). Using this method, we previously confirmed that the fluorescence signals of exogenously introduced F-CaM and of the immuno-staining of RyR2 showed a periodical pattern associated with that of the sarcomere, and, importantly, the staining patterns of both CaM and RyR2 matched each other very well. As shown in Fig. 4D, the concentration dependence of CaM binding to the Z-line was more greatly shifted towards higher concentrations in TACtreated cardiomyocytes than in sham-treated cardiomyocytes, whereas it was restored in the presence of dantrolene (1 mM).

changes domain unzipping to zipping), thereby inhibiting Ca2þ leakage through RyR2 [6,18,19]. Recently, several studies suggested that defective interactions of CaM with the RyR2 are a pathogenic factor in various heart diseases. For instance, according to Meissner et al. [22], knock-in mice containing RyR2 with a mutated CaM-binding domain that was unable to bind with CaM developed heart hypertrophy and early death. We have also shown that the binding affinity of CaM to the RyR2 in the failing RyR2 from pacing-induced canine failing hearts or TAC-induced LV pressure-overloaded mouse hearts was significantly lower than that of the normal RyR2 [7,8,10]. Further, betaadrenergic stimulation of CPVT-associated RyR2 reduced the binding affinity of CaM to the RyR2 considerably, while this treatment caused no appreciable change in the RyR2 in wild-type mice [9]. In contrast, enhancing the binding affinity of CaM to the RyR2 by applying three-amino acid substitutions to the N-terminal of CaM (GSH-CaM) inhibited the aberrant Ca2þ release in pacinginduced canine failing hearts [8] and CPVT-associated mouse hearts [11]. These findings strongly suggest that the defective interdomain interaction between the N-terminal domain and the central domain of the RyR2 is allosterically coupled with the reduction of its CaM binding affinity, thereby causing the observed aberrant Ca2þ release. However, this also shows that dantrolene can reverse these abnormal processes and prevent lethal arrhythmia. One of the most important aspects of this study is that dantrolene was found to be as effective in (acquired) chronic LV pressure-overloaded hearts as in (hereditary) CPVT-associated hearts through stabilization of the RyR2 and prevention of lethal arrhythmia. For the clinical use of dantrolene, it is quite advantageous that it exerts acute anti-arrhythmic effects with no detrimental effect on cardiac function, and it actually rather substantially improves the contractile function in failing hearts [17,19]. These beneficial effects are clearly different from those of other anti-arrhythmic drugs, such as beta-blockers or amiodarone. In conclusion, dantrolene, a stabilizer of the RyR2, inhibited aberrant Ca2þ release, including the occurrence of spontaneous Ca2þ sparks and transients, and thereby prevented the damaging effects of pharmacologically triggered, sustained VT in chronically LV pressure overload-induced failing hearts.

4. Discussion Declaration of competing interest Sustained VT, which frequently occurs in various kinds of diseased hearts, is known to be a major cause of sudden cardiac death. In the clinical setting, the effective pharmacological agents available for use in preventing such lethal arrhythmia in failing hearts are so far limited to beta-blockers and amiodarone [15]. However, there are a number of cases in which refractory VT has been observed, despite the use of beta-blockers and amiodarone. Therefore, a novel approach for the treatment of VT is desirable. It has been widely recognized that the aberrant release of Ca2þ from the RyR2 triggers delayed afterdepolarization and triggered activity, leading to lethal arrhythmia [16]. We previously reported that pre-treatment with dantrolene, a specific agent for treating MH, inhibited the aberrant Ca2þ release induced by protein kinase A (PKA) phosphorylation, and thereby prevented VT induced by either epinephrine or exercise, and significantly improved exercise tolerance in R2474S/þCPVT-associated knock-in mice [17]. The mechanism by which dantrolene prevents inducible VT is most likely through its stabilization of the diseased RyR2, on the basis of the findings that, in either RyR1 or RyR2, dantrolene specifically binds to domains with the same amino acid sequences (Leu590Cys609 in RyR1 or Leu600-Cys619 in RyR2) [18e21]. Further, dantrolene corrects the defective inter-domain interaction between the N-terminal and central domains of these receptor proteins (i.e.,

All authors confirm that there is no financial conflict of interest regarding this study. Acknowledgment This work was supported by grants-in-aid for scientific research from JSPS KAKENHI: JP26293189 to MY, JP15K09085 to TY, JP15K09142 to SK. We wish to acknowledge Yoko Okamoto, Satomi Tateda for their technical assistance. References [1] H. Ling, T. Zhang, L. Pereira, C.K. Means, H. Cheng, Y. Gu, N.D. Dalton, K.L. Peterson, J. Chen, D. Bers, J.H. Brown, Requirement for Ca2þ/calmodulindependent kinase II in the transition from pressure overload-induced cardiac hypertrophy to heart failure in mice, J. Clin. Investig. 119 (2009) 1230e1240. [2] T.H. Fischer, J. Herting, T. Tirilomis, A. Renner, S. Neef, K. Toischer, € rster, J.D. Schmitto, J. Gummert, F.A. Scho €ndube, D. Ellenberger, A. Fo G. Hasenfuss, L.S. Maier, S. Sossalla, Ca2þ/calmodulin-dependent protein kinase II and protein kinase A differentially regulate sarcoplasmic reticulum Ca2þ leak in human cardiac pathology, Circulation 128 (2013) 970e981. [3] S. Sedej, A. Schmidt, M. Denegri, S. Walther, M. Matovina, G. Arnstein, E.M. Gutschi, Windhager, S. Ljubojevi c, S. Negri, F.R. Heinzel, E. Bisping,

Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071

T. Kajii et al. / Biochemical and Biophysical Research Communications xxx (xxxx) xxx

[4]

[5]

[6]

[7]

[8]

[9]

[10]

[11]

[12]

€mper, B. Pieske, Subclinical abM.A. Vos, C. Napolitano, S.G. Priori, J. Kockska normalities in sarcoplasmic reticulum Ca2þ release promote eccentric myocardial remodeling and pump failure death in response to pressure overload, J. Am. Coll. Cardiol. 63 (2014) 1569e1579. T. Oda, M. Yano, T. Yamamoto, T. Tokuhisa, S. Okuda, M. Do, T. Ohkusa, Y. Ikeda, S. Kobayashi, N. Ikemoto, M. Matsuzaki, Defective regulation of interdomain interactions within the ryanodine receptor plays a key role in the pathogenesis of heart failure, Circulation 111 (2005) 3400e3410. T. Yamamoto, M. Yano, X. Xu, H. Uchinoumi, H. Tateishi, M. Mochizuki, T. Oda, S. Kobayashi, N. Ikemoto, M. Matsuzaki, Identification of target domains of the cardiac ryanodine receptor to correct channel disorder in failing hearts, Circulation 117 (2008) 762e772. H. Uchinoumi, M. Yano, T. Suetomi, M. Ono, X. Xu, H. Tateishi, T. Oda, S. Okuda, M. Doi, S. Kobayashi, T. Yamamoto, Y. Ikeda, T. Ohkusa, N. Ikemoto, M. Matsuzaki, Catecholaminergic polymorphic ventricular tachycardia is caused by mutation-linked defective conformational regulation of the ryanodine receptor, Circ. Res. 106 (2010) 1413e1424. M. Ono, M. Yano, A. Hino, T. Suetomi, X. Xu, T. Susa, H. Uchinoumi, H. Tateishi, T. Oda, S. Okuda, M. Doi, S. Kobayashi, T. Yamamoto, N. Koseki, H. Kyushiki, N. Ikemoto, M. Matsuzaki, Dissociation of calmodulin from cardiac ryanodine receptor causes aberrant Ca2þ release in heart failure, Cardiovasc. Res. 87 (2010) 609e617. A. Hino, M. Yano, T. Kato, M. Fukuda, T. Suetomi, M. Ono, W. Murakami, T. Susa, S. Okuda, M. Doi, S. Kobayashi, T. Yamamoto, N. Koseki, H. Kyushiki, N. Ikemoto, M. Matsuzaki, Enhanced binding of calmodulin to the ryanodine receptor corrects contractile dysfunction in failing hearts, Cardiovasc. Res. 96 (2012) 433e443. X. Xu, M. Yano, H. Uchinoumi, A. Hino, T. Suetomi, M. Ono, H. Tateishi, T. Oda, S. Okuda, M. Doi, S. Kobayashi, T. Yamamoto, Y. Ikeda, N. Ikemoto, M. Matsuzaki, Defective calmodulin binding to the cardiac ryanodine receptor plays a key role in CPVT-associated channel dysfunction, Biochem. Biophys. Res. Commun. 394 (2010) 660e666. T. Kato, T. Yamamoto, Y. Nakamura, T. Nanno, G. Fukui, Y. Sufu, Y. Hamada, T. Maeda, S. Nishimura, H. Ishiguchi, W. Murakami, M. Fukuda, X. Xu, A. Hino, M. Ono, T. Oda, S. Okuda, S. Kobayashi, N. Koseki, H. Kyushiki, M. Yano, Correction of impaired calmodulin binding to RyR2 as a novel therapy for lethal arrhythmia in the pressure-overloaded heart failure, Heart Rhythm 14 (2017) 120e127. M. Fukuda, T. Yamamoto, S. Nishimura, T. Kato, W. Murakami, A. Hino, M. Ono, H. Tateishi, T. Oda, S. Okuda, S. Kobayashi, N. Koseki, H. Kyushiki, M. Yano, Enhanced binding of calmodulin to RyR2 corrects arrhythmogenic channel disorder in CPVT-associated myocytes, Biochem. Biophys. Res. Commun. 448 (2014) 1e7. Y. Nakamura, T. Yamamoto, S. Kobayashi, M. Tamitani, Y. Hamada, G. Fukui,

[13] [14]

[15]

[16] [17]

[18]

[19]

[20]

[21]

[22]

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X. Xu, S. Nishimura, T. Kato, H. Uchinomi, T. Oda, S. Okuda, M. Yano, Ryanodine receptor-bound calmodulin is essential to protect against catecholaminergic polymorphic ventricular tachycardia, JCI Insight 4 (2019) e126112. X. Wu, D.M. Bers, Free and bound intracellular calmodulin measurements in cardiac myocytes, Cell Calcium 41 (2007) 353e364. Y. Yang, T. Guo, T. Oda, A. Chakraborty, L. Chen, H. Uchinoumi, A.A. Knowlton, B.R. Fruen, R.L. Cornea, G. Meissner, D.M. Bers, Cardiac myocyte Z-line calmodulin is mainly RyR2-bound, and reduction is arrhythmogenic and occurs in heart failure, Circ. Res. 114 (2014) 295e306. P. Ponikowski, A.A. Voors, S.D. Anker, H. Bueno, J.G. Cleland, A.J. Coats, V. Falk, J.R. Gonz alez-Juanatey, V.P. Harjola, E.A. Jankowska, M. Jessup, C. Linde, P. Nihoyannopoulos, J.T. Parissis, B. Pieske, J.P. Riley, G.M. Rosano, L.M. Ruilope, F. Ruschitzka, F.H. Rutten, P. van der Meer, Authors/Task Force Members, Document Reviewers, 2016 ESC Guidelines for the diagnosis and treatment of acute and chronic heart failure: the Task Force for the diagnosis and treatment of acute and chronic heart failure of the European Society of Cardiology (ESC). Developed with the special contribution of the Heart Failure Association (HFA) of the ESC, Eur. J. Heart Fail. 18 (2016) 891e975. S. Wagner, L.S. Maier, D.M. Bers, Role of sodium and calcium dysregulation in tachyarrhythmias in sudden cardiac death, Circ. Res. 116 (2015) 1956e1970. S. Kobayashi S, M. Yano, H. Uchinoumi, T. Suetomi, T. Susa T, M. Ono, X. Xu, H. Tateishi, T. Oda, S. Okuda, M. Doi, T. Yamamoto, M. Matsuzaki, Dantrolene, a therapeutic agent for malignant hyperthermia, inhibits catecholaminergic polymorphic ventricular tachycardia in a RyR2(R2474S/þ) knock-in mouse model, Circ. J. 74 (2010) 2579e2584. S. Kobayashi S, M.L. Bannister, J.P. Gangopadhyay, T. Hamada, J. Parness, N. Ikemoto, Dantrolene stabilizes domain interactions within the ryanodine receptor, J. Biol. Chem. 280 (2005) 6580e6587. S. Kobayashi, M. Yano, T. Suetomi, M. Ono, H. Tateishi, M. Mochizuki, X. Xu, H. Uchinoumi, S. Okuda, T. Yamamoto, N. Koseki, H. Kyushiki, N. Ikemoto, M. Matsuzaki, Dantrolene, a therapeutic agent for malignant hyperthermia, markedly improves the function of failing cardiomyocytes by stabilizing interdomain interactions within the ryanodine receptor, J. Am. Coll. Cardiol. 53 (2009) 1993e2005. K. Paul-Pletzer, T. Yamamoto, M.B. Bhat, J. Ma, N. Ikemoto, L.S. Jimenez, H. Morimoto, P.G. Williams, J. Parness, Identification of a dantrolene-binding sequence on the skeletal muscle ryanodine receptor, J. Biol. Chem. 277 (2002) 34918e34923. K. Paul-Pletzer, T. Yamamoto, N. Ikemoto, L.S. Jimenez, H. Morimoto, P.G. Williams, J. Ma, J. Parness, Probing a putative dantrolene-binding site on the cardiac ryanodine receptor, Biochem. J. 387 (2005) 905e909. N. Yamaguchi, N. Takahashi, L. Xu, O. Smithies, G. Meissner G, Early cardiac hypertrophy in mice with impaired calmodulin regulation of cardiac muscle Ca release channel, J. Clin. Investig. 117 (2007) 1344e1353.

Please cite this article as: T. Kajii et al., Dantrolene prevents ventricular tachycardia by stabilizing the ryanodine receptor in pressure- overload induced failing hearts, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.10.071