scholarly journals Interaction of the Joining Region in Junctophilin-2 With the L-Type Ca 2+ Channel Is Pivotal for Cardiac Dyad Assembly and Intracellular Ca 2+ Dynamics

2021 ◽  
Vol 128 (1) ◽  
pp. 92-114
Author(s):  
Polina Gross ◽  
Jaslyn Johnson ◽  
Carlos M. Romero ◽  
Deborah M. Eaton ◽  
Claire Poulet ◽  
...  
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Ventricular Myocytes ◽  
Close Association ◽  
Ryanodine Receptors ◽  
Left Ventricular ◽  
Induced Mutations ◽  
Adrenergic Stimulation ◽  
T Tubules ◽  
Junctional Sarcoplasmic Reticulum

Rationale: Ca 2+ -induced Ca 2+ release (CICR) in normal hearts requires close approximation of L-type calcium channels (LTCCs) within the transverse tubules (T-tubules) and RyR (ryanodine receptors) within the junctional sarcoplasmic reticulum. CICR is disrupted in cardiac hypertrophy and heart failure, which is associated with loss of T-tubules and disruption of cardiac dyads. In these conditions, LTCCs are redistributed from the T-tubules to disrupt CICR. The molecular mechanism responsible for LTCCs recruitment to and from the T-tubules is not well known. JPH (junctophilin) 2 enables close association between T-tubules and the junctional sarcoplasmic reticulum to ensure efficient CICR. JPH2 has a so-called joining region that is located near domains that interact with T-tubular plasma membrane, where LTCCs are housed. The idea that this joining region directly interacts with LTCCs and contributes to LTCC recruitment to T-tubules is unknown. Objective: To determine if the joining region in JPH2 recruits LTCCs to T-tubules through direct molecular interaction in cardiomyocytes to enable efficient CICR. Methods and Results: Modified abundance of JPH2 and redistribution of LTCC were studied in left ventricular hypertrophy in vivo and in cultured adult feline and rat ventricular myocytes. Protein-protein interaction studies showed that the joining region in JPH2 interacts with LTCC-α1C subunit and causes LTCCs distribution to the dyads, where they colocalize with RyRs. A JPH2 with induced mutations in the joining region (mut PG1 JPH2) caused T-tubule remodeling and dyad loss, showing that an interaction between LTCC and JPH2 is crucial for T-tubule stabilization. mut PG1 JPH2 caused asynchronous Ca 2+ -release with impaired excitation-contraction coupling after β-adrenergic stimulation. The disturbed Ca 2+ regulation in mut PG1 JPH2 overexpressing myocytes caused calcium/calmodulin-dependent kinase II activation and altered myocyte bioenergetics. Conclusions: The interaction between LTCC and the joining region in JPH2 facilitates dyad assembly and maintains normal CICR in cardiomyocytes.

scholarly journals Sarcolemmal distribution of ICa and INCX and Ca2+ autoregulation in mouse ventricular myocytes

2017 ◽  
Vol 313 (1) ◽  
pp. H190-H199 ◽  
Author(s):  
Hanne C. Gadeberg ◽  
Cherrie H. T. Kong ◽  
Simon M. Bryant ◽  
Andrew F. James ◽  
Clive H. Orchard
Keyword(s):  
Steady State ◽  
Sarcoplasmic Reticulum ◽  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Surface Membrane ◽  
Ryanodine Receptors ◽  
Cell Voltage ◽  
Intact Cells ◽  
T Tubules

The balance of Ca2+ influx and efflux regulates the Ca2+ load of cardiac myocytes, a process known as autoregulation. Previous work has shown that Ca2+ influx, via L-type Ca2+ current ( ICa), and efflux, via the Na+/Ca2+ exchanger (NCX), occur predominantly at t-tubules; however, the role of t-tubules in autoregulation is unknown. Therefore, we investigated the sarcolemmal distribution of ICa and NCX current ( INCX), and autoregulation, in mouse ventricular myocytes using whole cell voltage-clamp and simultaneous Ca2+ measurements in intact and detubulated (DT) cells. In contrast to the rat, INCX was located predominantly at the surface membrane, and the hysteresis between INCX and Ca2+ observed in intact myocytes was preserved after detubulation. Immunostaining showed both NCX and ryanodine receptors (RyRs) at the t-tubules and surface membrane, consistent with colocalization of NCX and RyRs at both sites. Unlike INCX, ICa was found predominantly in the t-tubules. Recovery of the Ca2+ transient amplitude to steady state (autoregulation) after application of 200 µM or 10 mM caffeine was slower in DT cells than in intact cells. However, during application of 200 µM caffeine to increase sarcoplasmic reticulum (SR) Ca2+ release, DT and intact cells recovered at the same rate. It appears likely that this asymmetric response to changes in SR Ca2+ release is a consequence of the distribution of ICa, which is reduced in DT cells and is required to refill the SR after depletion, and NCX, which is little affected by detubulation, remaining available to remove Ca2+ when SR Ca2+ release is increased. NEW & NOTEWORTHY This study shows that in contrast to the rat, mouse ventricular Na+/Ca2+ exchange current density is lower in the t-tubules than in the surface sarcolemma and Ca2+ current is predominantly located in the t-tubules. As a consequence, the t-tubules play a role in recovery (autoregulation) from reduced, but not increased, sarcoplasmic reticulum Ca2+ release.

scholarly journals MCU Overexpression Rescues Inotropy and Reverses Heart Failure by Reducing SR Ca 2+ Leak

2021 ◽  
Author(s):  
Ting Liu ◽  
Ni Yang ◽  
Agnieszka Sidor ◽  
Brian O'Rourke
Keyword(s):  
Oxidative Stress ◽  
Heart Failure ◽  
Cardiac Function ◽  
Ventricular Myocytes ◽  
Interstitial Fibrosis ◽  
Left Ventricular ◽  
Viral Gene ◽  
Adrenergic Stimulation ◽  
Guinea Pig Model

Rationale: In heart failure (HF), impaired sarcoplasmic reticulum (SR) Ca 2+ release and cytosolic Na + overload depress mitochondrial Ca 2+ (mCa 2+ ) signaling, resulting in a diminished ability to maintain matrix NAD(P)H redox potential, leading to increased oxidative stress when workload increases. Enhancing mCa 2+ can reverse this defect but could potentially increase the likelihood of mitochondrial Ca 2+ overload. Objective: To determine if moderate mitochondrial Ca 2+ uniporter (MCU) overexpression has beneficial or detrimental effects on the development of HF and incident arrythmias in a guinea pig model (ACi) of HF and sudden cardiac death. Methods and Results: In vivo viral gene transfer was used to increase MCU levels by ~30% in ACi hearts. Left ventricular myocytes from hearts with MCU overexpression (ACi+MCU) displayed enhanced mCa 2+ uptake, decreased oxidative stress, and increased β‐adrenergic- and frequency-dependent augmentation of Ca 2+ transients and contractions, compared to myocytes from ACi hearts. MCU overexpression decreased SR Ca 2+ leak in the ACi group and mitigated the elevated ryanodine receptor disulfide crosslinks in HF. β‐adrenergic responses were blunted in isolated perfused ACi hearts and these deficiencies were normalized in ACI+MCU hearts. To examine the in vivo effects of MCU overexpression, ACi hearts were transduced with the MCU virus 2 3w after aortic constriction, at the onset of cardiac decompensation. Two weeks later, cardiac function worsened in the untreated ACi group (fractional shortening: 39{plus minus}1% at 2w and 32{plus minus}1% at 4w), whereas MCU overexpression significantly improved cardiac function (36{plus minus}1% at 2w and 42{plus minus}2% at 4w). MCU overexpression in the decompensating ACi heart also attenuated pulmonary edema and interstitial fibrosis and prevented triggered arrhythmias. Conclusions: Moderate MCU overexpression in failing hearts enhances contractility and responses to β-adrenergic stimulation in isolated myocytes and perfused hearts by inhibiting mitochondrial oxidative stress-induced SR Ca 2+ leak. MCU overexpression also reversed HF and inhibited ectopic ventricular arrhythmias.

Decrease in the density of t-tubular L-type Ca2+ channel currents in failing ventricular myocytes

2011 ◽  
Vol 300 (3) ◽  
pp. H978-H988 ◽  
Author(s):  
Miwa Horiuchi-Hirose ◽  
Toshihide Kashihara ◽  
Tsutomu Nakada ◽  
Nagomi Kurebayashi ◽  
Hisashi Shimojo ◽  
...  
Keyword(s):  
Heart Failure ◽  
Sarcoplasmic Reticulum ◽  
Current Density ◽  
Ventricular Myocytes ◽  
Osmotic Shock ◽  
Adrenergic Stimulation ◽  
Whole Cell ◽  
Subunit Expression ◽  
Dependent Protein ◽  
T Tubules

In some forms of cardiac hypertrophy and failure, the gain of Ca2+-induced Ca2+ release [CICR; i.e., the amount of Ca2+ released from the sarcoplasmic reticulum normalized to Ca2+ influx through L-type Ca2+ channels (LTCCs)] decreases despite the normal whole cell LTCC current density, ryanodine receptor number, and sarcoplasmic reticulum Ca2+ content. This decrease in CICR gain has been proposed to arise from a change in dyad architecture or derangement of the t-tubular (TT) structure. However, the activity of surface sarcolemmal LTCCs has been reported to increase despite the unaltered whole cell LTCC current density in failing human ventricular myocytes, indicating that the “decreased CICR gain” may reflect a decrease in the TT LTCC current density in heart failure. Thus, we analyzed LTCC currents of failing ventricular myocytes of mice chronically treated with isoproterenol (Iso). Although Iso-treated mice exhibited intact t-tubules and normal LTCC subunit expression, acute occlusion of t-tubules of isolated ventricular myocytes with osmotic shock (detubulation) revealed that the TT LTCC current density was halved in Iso-treated versus control myocytes. Pharmacological analysis indicated that kinases other than PKA or Ca2+/calmodulin-dependent protein kinase II insufficiently activated, whereas protein phosphatase 1/2A excessively suppressed, TT LTCCs in Iso-treated versus control myocytes. These results indicate that excessive β-adrenergic stimulation causes the decrease in TT LTCC current density by altering the regulation of TT LTCCs by protein kinases and phosphatases in heart failure. This phenomenon might underlie the decreased CICR gain in heart failure.

β-blockade abolishes the augmented cardiac tPA release induced by transactivation of heterodimerised bradykinin receptor-2 and β2-adrenergic receptor in vivo

2014 ◽  
Vol 112 (11) ◽  
pp. 951-959 ◽  
Author(s):  
Morten Eriksen ◽  
Arnfinn Ilebekk ◽  
Alessandro Cataliotti ◽  
Cathrine Rein Carlson ◽  
Torstein Lyberg ◽  
...  
Keyword(s):  
Adrenergic Receptor ◽  
Sympathetic Nerves ◽  
Ventricular Myocardium ◽  
Left Ventricular ◽  
Left Ventricular Myocardium ◽  
Adrenergic Stimulation ◽  
Β2 Adrenergic Receptor ◽  
Β Blocker

SummaryBradykinin (BK) receptor-2 (B2R) and β2-adrenergic receptor (β2AR) have been shown to form heterodimers in vitro. However, in vivo proofs of the functional effects of B2R-β2AR heterodimerisation are missing. Both BK and adrenergic stimulation are known inducers of tPA release. Our goal was to demonstrate the existence of B2R-β2AR heterodimerisation in myocardium and to define its functional effect on cardiac release of tPA in vivo. We further investigated the effects of a non-selective β-blocker on this receptor interplay. To investigate functional effects of B2R-β2AR heterodimerisation (i. e. BK transactivation of β2AR) in vivo, we induced serial electrical stimulation of cardiac sympathetic nerves (SS) in normal pigs that underwent concomitant BK infusion. Both SS and BK alone induced increases in cardiac tPA release. Importantly, despite B2R desensitisation, simultaneous BK infusion and SS (BK+SS) was characterised by 2.3 ± 0.3-fold enhanced tPA release compared to SS alone. When β-blockade (propranolol) was introduced prior to BK+SS, tPA release was inhibited. A persistent B2R-β2AR heterodimer was confirmed in BK-stimulated and nonstimulated left ventricular myocardium by immunoprecipitation studies and under non-reducing gel conditions. All together, these results strongly suggest BK transactivation of β2AR leading to enhanced β2AR-mediated release of tPA. Importantly, non-selective β-blockade inhibits both SS-induced release of tPA and the functional effects of B2R-β2AR heterodimerisation in vivo, which may have important clinical implications.

Age and hypertrophy alter the contribution of sarcoplasmic reticulum and Na+/Ca2+ exchange to Ca2+ removal in rat left ventricular myocytes

2007 ◽  
Vol 42 (3) ◽  
pp. 582-589 ◽  
Author(s):  
Mark R. Fowler ◽  
James R. Naz ◽  
Mark D. Graham ◽  
Clive H. Orchard ◽  
Simon M. Harrison
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Ventricular Myocytes ◽  
Left Ventricular

scholarly journals Role of type 2A phosphatase regulatory subunit B56α in regulating cardiac responses to β-adrenergic stimulation in vivo

2018 ◽  
Vol 115 (3) ◽  
pp. 519-529 ◽  
Author(s):  
Sarah-Lena Puhl ◽  
Kate L Weeks ◽  
Alican Güran ◽  
Antonella Ranieri ◽  
Peter Boknik ◽  
...  
Keyword(s):  
Systolic Dysfunction ◽  
Heart Tissue ◽  
Left Ventricular ◽  
Regulatory Subunit ◽  
Adrenergic Stimulation ◽  
Hypertrophic Response ◽  
Cardiac Responses ◽  
Cardiac Phenotype ◽  
And Function

Abstract Aims B56α is a protein phosphatase 2A (PP2A) regulatory subunit that is highly expressed in the heart. We previously reported that cardiomyocyte B56α localizes to myofilaments under resting conditions and translocates to the cytosol in response to acute β-adrenergic receptor (β-AR) stimulation. Given the importance of reversible protein phosphorylation in modulating cardiac function during sympathetic stimulation, we hypothesized that loss of B56α in mice with targeted disruption of the gene encoding B56α (Ppp2r5a) would impact on cardiac responses to β-AR stimulation in vivo. Methods and results Cardiac phenotype of mice heterozygous (HET) or homozygous (HOM) for the disrupted Ppp2r5a allele and wild type (WT) littermates was characterized under basal conditions and following acute β-AR stimulation with dobutamine (DOB; 0.75 mg/kg i.p.) or sustained β-AR stimulation by 2-week infusion of isoproterenol (ISO; 30 mg/kg/day s.c.). Left ventricular (LV) wall thicknesses, chamber dimensions and function were assessed by echocardiography, and heart tissue collected for gravimetric, histological, and biochemical analyses. Western blot analysis revealed partial and complete loss of B56α protein in hearts from HET and HOM mice, respectively, and no changes in the expression of other PP2A regulatory, catalytic or scaffolding subunits. PP2A catalytic activity was reduced in hearts of both HET and HOM mice. There were no differences in the basal cardiac phenotype between genotypes. Acute DOB stimulation induced the expected inotropic response in WT and HET mice, which was attenuated in HOM mice. In contrast, DOB-induced increases in heart rate were unaffected by B56α deficiency. In WT mice, ISO infusion increased LV wall thicknesses, cardiomyocyte area and ventricular mass, without LV dilation, systolic dysfunction, collagen deposition or foetal gene expression. The hypertrophic response to ISO was blunted in mice deficient for B56α. Conclusion These findings identify B56α as a potential regulator of cardiac structure and function during β-AR stimulation.

Abstract 631 ADAM12 Is An Important Regulator Of Myocyte Apoptosis Induced By β-adrenergic Receptor Stimulation

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Hang Xi ◽  
Khadija Rafiq ◽  
Marie Hanscom ◽  
Rachid Seqqat ◽  
Nikolay L Malinin ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Left Ventricular ◽  
Surface Proteins ◽  
Neonatal Rat ◽  
Human Heart Failure ◽  
Activation Mechanisms ◽  
Myocyte Apoptosis ◽  
Protease Deficient

ADAM (A Disintegrin And Metalloprotease)12 is a member of a family of cell surface proteins with protease and cell-binding activities. Recent work showed ADAM12 up-regulation in human heart failure. However, the activation mechanisms of ADAM12 in the heart are obscure. We hypothesized that β-adrenergic receptors (AR) stimulation regulates ADAM12 activation in neonatal rat ventricular myocytes (NRVMs) in-vitro and after injection of isoproterenol (ISO) in-vivo. Wistar rats received a single injection of ISO (5 mg/kg) and were sacrificed 6, 24 and 72 hrs later. In comparison with controls, left ventricular function was impaired in rats 24 hrs after ISO injection and started to improve at 72 hrs. The fraction of myocytes undergoing apoptosis peaked 24 hrs after ISO injection and declined thereafter. ADAM12 protein was reduced in hearts from ISO treated animals at 6 hrs, pointing to a possible increase in ADAM12 proteolytic activity. However, both ADAM12 expression and activation were significantly up-regulated at 24 and 72 hrs after ISO injection. We therefore assessed whether ADAM12 activation was involved in myocyte apoptosis secondary to excess exposure of catecholamine. Acute stimulation with ISO (10 μM, 30 min to 3 hrs) induced accumulation of ADAM12 N-terminal cleavage product in conditioned medium, demonstrating activation of the ADAM metalloprotease activity. However, chronic stimulation with ISO for 24 hrs and 48 hrs significantly increased both ADAM12 expression and secretion. This ISO-induced ADAM12 expression/activation was mediated through β 1 -AR stimulation and was dependent on intracellular calcium elevation and protein kinase C activation. Adenoviral expression of an ADAM12 protease-deficient mutant (ADAM12DeltaMP) blocks β-AR-induced myocyte apoptosis, while transduction of NRVMs with adenovirus harboring ADAM12 significantly increased myocyte apoptosis. These data suggest that ADAM12 is a regulator of myocyte apoptosis induced by β-AR in NRVMs and may play an important autocrine role in mediating the effects of β-AR on myocardial remodeling.

Abstract 18111: Flecainide Exerts its Antiarrhythmic Action in CPVT Through Blockade of Neuronal Na+ channel-mediated Arrhythmogenic Diastolic Ca2+ Release

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Przemyslaw Radwanski ◽  
Hsiang-Ting Ho ◽  
Björn Knollmann ◽  
Andriy Belevych ◽  
Sándor Györke
Keyword(s):  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Cellular Level ◽  
Antiarrhythmic Effect ◽  
Na Channel ◽  
Antiarrhythmic Action ◽  
Pharmacological Interventions ◽  
Cellular Excitability

Background: Flecaininde is an effective antiarrhythmic in management of CPVT. Its antiarrhythmic action has been attributed to direct effect on RyR2 and reduced cellular excitability through the inhibition of cardiac-type Na + channels. Recently we demonstrated that neuronal Na + channels (nNa v s) colocalize with the ryanodine receptors (RyR2) Ca 2+ release channels on the sarcoplasmic reticulum. Here we explore a novel mechanism that may contribute to the antiarrhythmic effect of flecainide, mainly uncoupling of aberrant Na + /Ca 2+ signaling through nNa v inhibition. Methods: To study the effects of flecainide on Ca 2+ signaling we used a murine model of cardiac calsequestrin-associated CPVT. We performed confocal microscopy in intact isolated ventricular myocytes to assess Ca 2+ handling and recorded late Na + current (I Na ) during various pharmacological interventions. Surface electrocardiograms were performed during catecholamine challenge to monitor arrhythmic activity in vivo . Results: During catecholamine stimulation with isoproterenol (Iso; 100 nM) disruption of the cross-talk between nNa v s and RyR2 by nNa v blockade with 100nM tetrodotoxin (TTX) and riluzole (10μM) as well as flecainide (2.5μM) reduced Iso-promoted late I Na and DCR in isolated intact CPVT cardiomyocytes. To further examine the role of nNa v -mediated late I Na in genesis of DCR we augmented nNa v channel activity with β-Pompilidotoxin (β-PMTX, 40μM). Effects of β-PMTX in CPVT cardiomyocytes were reversed by nNa v blockade with TTX and riluzole as well as flecainide. This reduction in late I Na and DCR frequency with riluzole and flecainide in the presence of β-PMTX on cellular level translated to decreased ventricular arrhythmias in CPVT mice. Conclusion: These data suggest that disruption of nNa v -mediated late I Na can prevent arrhythmogenic DCR in CPVT. Importantly, the antiarrhythmic effects of flecainide can be attributed, at least in part, to its nNa v blocking properties.

Inotropic sensitivity to beta-adrenergic stimulation in early sepsis

1988 ◽  
Vol 255 (4) ◽  
pp. H699-H703 ◽  
Author(s):  
L. W. Smith ◽  
K. H. McDonough
Keyword(s):  
Contractile Function ◽  
Plasma Catecholamines ◽  
Left Ventricular ◽  
Adrenergic Stimulation ◽  
Maximal Increase ◽  
Beta Adrenergic ◽  
Arterial Blood ◽  
Myocardial Contractile ◽  
Early Sepsis

In early sepsis, maintenance of in vivo cardiovascular performance is at least partly dependent on sympathetic support to hearts with intrinsic contractile defects. Yet prolonged sympathetic stimulation, as occurs in sepsis, would be expected to alter the heart's ability to respond to this stimulation. We have investigated myocardial inotropic sensitivity to beta-adrenergic stimulation in a model of sepsis in which animals, at the time studied, exhibited bacteremia, normal arterial blood pressure and cardiac output, elevated heart rate, and elevated plasma catecholamines. Intrinsic myocardial contractile function, as assessed by the maximal rate of left ventricular pressure development (LV dP/dtmax) in an isovolumically contracting heart preparation, was significantly depressed in septic animals. To determine whether hearts from septic animals could respond normally to beta-adrenergic stimulation, we studied inotropic response to a bolus of isoproterenol in these isolated hearts. With maximal isoproterenol stimulation, hearts from septic animals were able to attain the same dP/dtmax as were hearts from control animals. With lower levels of isoproterenol, there was also no difference in inotropic indexes between the two groups when response was expressed as a percent of the maximal increase in dP/dtmax achieved with isoproterenol. These results suggest that in early sepsis, despite intrinsic myocardial contractile dysfunction, the ability of the heart to modulate its inotropic state in response in beta-adrenergic stimulation is intact.

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