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.

scholarly journals Altered Na/Ca exchange distribution in ventricular myocytes from failing hearts

2016 ◽  
Vol 310 (2) ◽  
pp. H262-H268 ◽  
Author(s):  
Hanne C. Gadeberg ◽  
Simon M. Bryant ◽  
Andrew F. James ◽  
Clive H. Orchard
Keyword(s):  
Heart Failure ◽  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Coronary Artery Ligation ◽  
Sham Operation ◽  
Ca Current ◽  
Artery Ligation ◽  
Whole Cell ◽  
Rat Hearts ◽  
T Tubules

In mammalian cardiac ventricular myocytes, Ca efflux via Na/Ca exchange (NCX) occurs predominantly at T tubules. Heart failure is associated with disrupted t-tubular structure, but its effect on t-tubular function is less clear. We therefore investigated t-tubular NCX activity in ventricular myocytes isolated from rat hearts ∼18 wk after coronary artery ligation (CAL) or corresponding sham operation (Sham). NCX current ( INCX) and l-type Ca current ( ICa) were recorded using the whole cell, voltage-clamp technique in intact and detubulated (DT) myocytes; intracellular free Ca concentration ([Ca]i) was monitored simultaneously using fluo-4. INCX was activated and measured during application of caffeine to release Ca from sarcoplasmic reticulum (SR). Whole cell INCX was not significantly different in Sham and CAL myocytes and occurred predominantly in the T tubules in Sham myocytes. CAL was associated with redistribution of INCX and ICa away from the T tubules to the cell surface and an increase in t-tubular INCX/ ICa density from 0.12 in Sham to 0.30 in CAL myocytes. The decrease in t-tubular INCX in CAL myocytes was accompanied by an increase in the fraction of Ca sequestered by SR. However, SR Ca content was not significantly different in Sham, Sham DT, and CAL myocytes but was significantly increased by DT of CAL myocytes. In Sham myocytes, there was hysteresis between INCX and [Ca]i, which was absent in DT Sham but present in CAL and DT CAL myocytes. These data suggest altered distribution of NCX in CAL myocytes.

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 Reduced density and altered regulation of rat atrial L-type Ca2+current in heart failure

2017 ◽  
Vol 312 (3) ◽  
pp. H384-H391 ◽  
Author(s):  
Richard C. Bond ◽  
Simon M. Bryant ◽  
Judy J. Watson ◽  
Jules C. Hancox ◽  
Clive H. Orchard ◽  
...  
Keyword(s):  
Heart Failure ◽  
Coronary Artery ◽  
Current Density ◽  
Ventricular Myocytes ◽  
Coronary Artery Ligation ◽  
Sham Operation ◽  
Atrial Myocytes ◽  
Artery Ligation ◽  
Whole Cell ◽  
Male Wistar Rats

Constitutive regulation by PKA has recently been shown to contribute to L-type Ca2+current ( ICaL) at the ventricular t-tubule in heart failure. Conversely, reduction in constitutive regulation by PKA has been proposed to underlie the downregulation of atrial ICaLin heart failure. The hypothesis that downregulation of atrial ICaLin heart failure involves reduced channel phosphorylation was examined. Anesthetized adult male Wistar rats underwent surgical coronary artery ligation (CAL, N=10) or equivalent sham-operation (Sham, N=12). Left atrial myocytes were isolated ~18 wk postsurgery and whole cell currents recorded (holding potential=-80 mV). ICaLactivated by depolarizing pulses to voltages from -40 to +50 mV were normalized to cell capacitance and current density-voltage relations plotted. CAL cell capacitances were ~1.67-fold greater than Sham ( P ≤ 0.0001). Maximal ICaLconductance ( Gmax) was downregulated more than 2-fold in CAL vs. Sham myocytes ( P < 0.0001). Norepinephrine (1 μmol/l) increased Gmax>50% more effectively in CAL than in Sham so that differences in ICaLdensity were abolished. Differences between CAL and Sham Gmaxwere not abolished by calyculin A (100 nmol/l), suggesting that increased protein dephosphorylation did not account for ICaLdownregulation. Treatment with either H-89 (10 μmol/l) or AIP (5 μmol/l) had no effect on basal currents in Sham or CAL myocytes, indicating that, in contrast to ventricular myocytes, neither PKA nor CaMKII regulated basal ICaL. Expression of the L-type α1C-subunit, protein phosphatases 1 and 2A, and inhibitor-1 proteins was unchanged. In conclusion, reduction in PKA-dependent regulation did not contribute to downregulation of atrial ICaLin heart failure.NEW & NOTEWORTHY Whole cell recording of L-type Ca2+currents in atrial myocytes from rat hearts subjected to coronary artery ligation compared with those from sham-operated controls reveals marked reduction in current density in heart failure without change in channel subunit expression and associated with altered phosphorylation independent of protein kinase A.

Chronic receptor-mediated activation of Gi/o proteins alters basal t-tubular and sarcolemmal L-type Ca2+ channel activity through phosphatases in heart failure

2012 ◽  
Vol 302 (8) ◽  
pp. H1645-H1654 ◽  
Author(s):  
Toshihide Kashihara ◽  
Tsutomu Nakada ◽  
Hisashi Shimojo ◽  
Miwa Horiuchi-Hirose ◽  
Simmon Gomi ◽  
...  
Keyword(s):  
Heart Failure ◽  
Current Density ◽  
Ventricular Myocytes ◽  
Heterotrimeric G Proteins ◽  
Channel Activity ◽  
Open Probability ◽  
Current Densities ◽  
Fractional Shortening ◽  
Pp2a Inhibitor

L-type Ca2+ channels (LTCCs) play an essential role in the excitation-contraction coupling of ventricular myocytes. We previously found that t-tubular (TT) LTCC current density was halved by the activation of protein phosphatase (PP)1 and/or PP2A, whereas surface sarcolemmal (SS) LTCC current density was increased by the inhibition of PP1 and/or PP2A activity in failing ventricular myocytes of mice chronically treated with isoproterenol (ISO mice). In the present study, we examined the possible involvement of inhibitory heterotrimeric G proteins (Gi/o) in these abnormalities by chronically administrating pertussis toxin (PTX) to ISO mice (ISO + PTX mice). Compared with ISO mice, ISO + PTX mice exhibited significantly higher fractional shortening of the left ventricle. The expression level of Gαi2 proteins was not altered by the treatment of mice with ISO and/or PTX. ISO + PTX myocytes had normal TT and SS LTCC current densities because they had higher and lower availability and/or open probability of TT and SS LTCCs than ISO myocytes, respectively. A selective PKA inhibitor, H-89, did not affect LTCC current densities in ISO + PTX myocytes. A selective PP2A inhibitor, fostriecin, did not affect SS or TT current density in control or ISO + PTX myocytes but significantly increased TT but not SS LTCC current density in ISO myocytes. These results indicate that chronic receptor-mediated activation of Gi/o in vivo decreases basal TT LTCC activity by activating PP2A and increases basal SS LTCC activity by inhibiting PP1 without modulating PKA in heart failure.

Steady-state twitch Ca2+ fluxes and cytosolic Ca2+ buffering in rabbit ventricular myocytes

1996 ◽  
Vol 270 (1) ◽  
pp. C192-C199 ◽  
Author(s):  
L. M. Delbridge ◽  
J. W. Bassani ◽  
D. M. Bers
Keyword(s):  
Steady State ◽  
Sarcoplasmic Reticulum ◽  
Voltage Clamp ◽  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Whole Cell ◽  
Rabbit Ventricular Myocytes ◽  
The Mean ◽  
State Contraction ◽  
Caffeine Contractures

Intracellular Ca2+ ([Ca2+]i) transients and transsarcolemmal Ca2+ currents were measured in indo 1-loaded isolated rabbit ventricular myocytes during whole cell voltage clamp to quantitate the components of cytosolic Ca2+ influx and to describe the dynamic aspects of cytosolic Ca2+ buffering during steady-state contraction (0.5 Hz, 22 degrees C). Sarcolemmal Ca2+ influx was directly measured from the integrated Ca2+ current (Ica) recorded during the clamp (158 +/- 10 attomoles; amol). Sarcoplasmic reticulum (SR) Ca2+ content was determined from the integrated electrogenic Na+/Ca2+ exchange current (Ix) induced during rapid application and sustained exposure of cells to caffeine to elicit the release of the SR Ca2+ load (1,208 +/- 170 amol). The mean steady-state SR Ca2+ load was calculated to be 87 +/- 13 microM (mumol/l nonmitochondrial cytosolic volume). Ca2+ influx via Ica represented approximately 14% of the stored SR Ca2+ and 23% of the total cytosolic Ca2+ flux during a twitch (47 +/- 6 microM). Comparison of electrophysiologically measured Ca2+ fluxes with Ca2+ transients yields apparent buffering values of 60 for caffeine contractures and 110 for twitches (delta Ca2+ total/delta Ca2+ free). This is consistent with the occurrence of "active" buffering of cytosolic Ca2+ by SR Ca2+ uptake during the twitch.

scholarly journals Ultrastructural uncoupling between T-tubules and sarcoplasmic reticulum in human heart failure

2013 ◽  
Vol 98 (2) ◽  
pp. 269-276 ◽  
Author(s):  
H.-B. Zhang ◽  
R.-C. Li ◽  
M. Xu ◽  
S.-M. Xu ◽  
Y.-S. Lai ◽  
...  
Keyword(s):  
Heart Failure ◽  
Sarcoplasmic Reticulum ◽  
Human Heart ◽  
Human Heart Failure ◽  
T Tubules

scholarly journals The calcium stored in the sarcoplasmic reticulum acts as a safety mechanism in rainbow trout heart

2014 ◽  
Vol 307 (12) ◽  
pp. R1493-R1501 ◽  
Author(s):  
Caroline Cros ◽  
Laurent Sallé ◽  
Daniel E. Warren ◽  
Holly A. Shiels ◽  
Fabien Brette
Keyword(s):  
Rainbow Trout ◽  
Sarcoplasmic Reticulum ◽  
Ventricular Myocytes ◽  
Adrenergic Stimulation ◽  
Relative Contribution ◽  
Safety Mechanism ◽  
Rapid Changes ◽  
Intracellular Ca ◽  
As Stress

Cardiomyocyte contraction depends on rapid changes in intracellular Ca2+. In mammals, Ca2+ influx as L-type Ca2+ current ( ICa) triggers the release of Ca2+ from sarcoplasmic reticulum (SR) and Ca2+-induced Ca2+ release (CICR) is critical for excitation-contraction coupling. In fish, the relative contribution of external and internal Ca2+ is unclear. Here, we characterized the role of ICa to trigger SR Ca2+ release in rainbow trout ventricular myocytes using ICa regulation by Ca2+ as an index of CICR. ICa was recorded with a slow (EGTA) or fast (BAPTA) Ca2+ chelator in control and isoproterenol conditions. In the absence of β-adrenergic stimulation, the rate of ICa inactivation was not significantly different in EGTA and BAPTA (27.1 ± 1.8 vs. 30.3 ± 2.4 ms), whereas with isoproterenol (1 μM), inactivation was significantly faster with EGTA (11.6 ± 1.7 vs. 27.3 ± 1.6 ms). When barium was the charge carrier, inactivation was significantly slower in both conditions (61.9 ± 6.1 vs. 68.0 ± 8.7 ms, control, isoproterenol). Quantification revealed that without isoproterenol, only 39% of ICa inactivation was due to Ca2+, while with isoproterenol, inactivation was Ca2+-dependent (∼65%) and highly reliant on SR Ca2+ (∼46%). Thus, SR Ca2+ is not released in basal conditions, and ICa is the main trigger of contraction, whereas during a stress response, SR Ca2+ is an important source of cytosolic Ca2+. This was not attributed to differences in SR Ca2+ load because caffeine-induced transients were not different in both conditions. Therefore, Ca2+ stored in SR of trout cardiomyocytes may act as a safety mechanism, allowing greater contraction when higher contractility is required, such as stress or exercise.

CaM kinase augments cardiac L-type Ca2+ current: a cellular mechanism for long Q-T arrhythmias

1999 ◽  
Vol 276 (6) ◽  
pp. H2168-H2178 ◽  
Author(s):  
Yuejin Wu ◽  
Leigh B. MacMillan ◽  
R. Blair McNeill ◽  
Roger J. Colbran ◽  
Mark E. Anderson
Keyword(s):  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Cam Kinase ◽  
Early Afterdepolarizations ◽  
Ultrastructural Studies ◽  
Intracellular Ca ◽  
Dependent Protein ◽  
T Tubules ◽  
Rabbit Ventricular Myocytes

Early afterdepolarizations (EAD) caused by L-type Ca2+ current ( I Ca,L) are thought to initiate long Q-T arrhythmias, but the role of intracellular Ca2+ in these arrhythmias is controversial. Rabbit ventricular myocytes were stimulated with a prolonged EAD-containing action potential-clamp waveform to investigate the role of Ca2+/calmodulin-dependent protein kinase II (CaM kinase) in I Ca,L during repolarization. I Ca,L was initially augmented, and augmentation was dependent on Ca2+ from the sarcoplasmic reticulum because the augmentation was prevented by ryanodine or thapsigargin. I Ca,Laugmentation was also dependent on CaM kinase, because it was prevented by dialysis with the inhibitor peptide AC3-I and reconstituted by exogenous constitutively active CaM kinase when Ba2+ was substituted for bath Ca2+. Ultrastructural studies confirmed that endogenous CaM kinase, L-type Ca2+ channels, and ryanodine receptors colocalized near T tubules. EAD induction was significantly reduced in current-clamped cells dialyzed with AC3-I (4/15) compared with cells dialyzed with an inactive control peptide (11/15, P = 0.013). These findings support the hypothesis that EADs are facilitated by CaM kinase.

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