Role Of Sarcoplasmic Reticulum Ca2 Research Articles

Role of Sarcoplasmic Reticulum Ca2+ Release in Hypoxia-induced Mitochondrial ROS Generation and Underlying Molecular Mechanisms in Pulmonary Artery Smooth Muscle Cells

Role of Sarcoplasmic Reticulum Ca2+ Release in Hypoxia-induced Mitochondrial ROS Generation and Underlying Molecular Mechanisms in Pulmonary Artery Smooth Muscle Cells

The role of luminal Ca regulation in Ca signaling refractoriness and cardiac arrhythmogenesis.

Györke et al. discuss the role of sarcoplasmic reticulum Ca2+ in cardiac refractoriness and pathological implications.

Differences in the force–interval relationship of isolated human myocardium with chronic coronary artery disease with and without type 2 diabetes mellitus and the role of sarcoplasmic reticulum Ca2+-ATPase

The dynamics of the force–interval relationship of the human myocardium in coronary artery disease (CAD) and CAD with concomitant diabetes mellitus was studied, and its dependence on the level of sarcoplasmic reticulum (SR) Ca2+-ATPase expression was evaluated. The study was performed on myocardial biopsy material obtained during coronary bypass operation using cardiopulmonary techniques. Patients with chronic CAD and patients with CAD associated with type II diabetes mellitus were enrolled in the study. It was found that CAD patients with and without diabetes mellitus with similar clinical parameters had either negative or positive dynamics of the force–interval relationship. The positive force–interval relationship was associated with a “high level” of Ca2+-ATPase, while the negative force–interval relationship was associated with a “low level” of this protein. In CAD associated with diabetes mellitus of short duration, the positive dynamics of the force–interval relationship is more pronounced and corresponds to a higher level of SR Ca2+-ATPase expression than in CAD alone.

Optical Mapping of Intra-Sarcoplasmic Reticulum Ca2+ and Transmembrane Potential in the Langendorff-perfused Rabbit Heart.

Sarcoplasmic reticulum (SR) Ca2+ handling plays a key role in normal excitation-contraction coupling and aberrant SR Ca2+ handling is known to play a significant role in certain types of arrhythmia. Because arrhythmias are spatially distinct, emergent phenomena, they must be investigated at the tissue level. However, methods for directly probing SR Ca2+ in the intact heart remain limited. This article describes the protocol for dual optical mapping of transmembrane potential (Vm) and free intra-SR [Ca2+] ([Ca2+]SR) in the Langendorff-perfused rabbit heart. This approach takes advantage of the low-affinity Ca2+ indicator Fluo-5N, which has minimal fluorescence in the cytosol where intracellular [Ca2+] ([Ca2+]i) is relatively low but exhibits significant fluorescence in the SR lumen where [Ca2+]SR is in the millimolar range. In addition to revealing SR Ca2+ characteristics spatially across the epicardial surface of the heart, this approach has the distinct advantage of simultaneous monitoring of Vm, allowing for investigations into the bidirectional relationship between Vm and SR Ca2+ and the role of SR Ca2+ in arrhythmogenic phenomena.

Human skeletal muscle glycogen utilization in exhaustive exercise: role of subcellular localization and fibre type

Although glycogen is known to be heterogeneously distributed within skeletal muscle cells, there is presently little information available about the role of fibre types, utilization and resynthesis during and after exercise with respect to glycogen localization. Here, we tested the hypothesis that utilization of glycogen with different subcellular localizations during exhaustive arm and leg exercise differs and examined the influence of fibre type and carbohydrate availability on its subsequent resynthesis. When 10 elite endurance athletes (22 ± 1 years, VO2 max = 68 ± 5 ml kg-1 min-1, mean ± SD) performed one hour of exhaustive arm and leg exercise, transmission electron microscopy revealed more pronounced depletion of intramyofibrillar than of intermyofibrillar and subsarcolemmal glycogen. This phenomenon was the same for type I and II fibres, although at rest prior to exercise, the former contained more intramyofibrillar and subsarcolemmal glycogen than the latter. In highly glycogen-depleted fibres, the remaining small intermyofibrillar and subsarcolemmal glycogen particles were often found to cluster in groupings. In the recovery period, when the athletes received either a carbohydrate-rich meal or only water the impaired resynthesis of glycogen with water alone was associated primarily with intramyofibrillar glycogen. In conclusion, after prolonged high-intensity exercise the depletion of glycogen is dependent on subcellular localization. In addition, the localization of glycogen appears to be influenced by fibre type prior to exercise, as well as carbohydrate availability during the subsequent period of recovery. These findings provide insight into the significance of fibre type-specific compartmentalization of glycogen metabolism in skeletal muscle during exercise and subsequent recovery. .

Differences in Sarcoplasmic Reticulum Ca(superscript 2+) Leak Characteristics between Systolic and Diastolic Heart Failure Rabbit Models

Diastolic heart failure (DHF) and systolic heart failure (SHF) are two clinical subsets of chronic heart failure (CHF). Sarcoplasmic reticulum (SR) Ca²⁺ leak has been measured in SHF and might contribute to contractile dysfunction and arrhythmogenesis. However, no study has investigated a similar phenomenon in DHF. Thus, we established DHF and SHF rabbit models and compared the differences in Ca²⁺ leak between these models. New Zealand white rabbits were randomly divided into three groups (n = 8 in each group): sham operation (SO) group, DHF group and SHF group. Cardiac functions were determined by echocardiography and hemodynamic assays. The SR Ca²⁺ leak was measured with a calcium-imaging device and the expression and activities of related proteins were evaluated with Western blots and autophosphorylation. In the DHF group, there was significantly increased ventricular wall thickness and stiffness, reduced diastolic function, and total amount of FK506 binding protein 12.6 (FKBP12.6), increased expression and activity of protein kinase A (PKA) and phosphorylation site (P2809) in the ryanodine receptor (RyR2), but no prominent Ca²⁺ leak. In the SHF group, there was significantly increased ventricular cavity size, reduced systolic function, increased SR Ca²⁺ leak, reduced total amount of FKBP12.6 and FKBP12.6-RyR2 association, increased expression and activity of PKA and Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) and their RyR2 phosphorylation sites with unchanged P2030. Our results suggest that a prominent SR Ca²⁺ leak was not observed in the DHF model, which may provide a new idea for the reasons in preserved systolic function, and CaMKII possibly plays a more important role in SR Ca²⁺ leak.

Ca2+spark-dependent and -independent sarcoplasmic reticulum Ca2+leak in normal and failing rabbit ventricular myocytes

Sarcoplasmic reticulum (SR) Ca²(+) leak is an important component of cardiac Ca²(+) signalling. Together with the SR Ca²(+)-ATPase (SERCA)-mediated Ca²(+) uptake, diastolic Ca²(+) leak determines SR Ca²(+) load and, therefore, the amplitude of Ca²(+) transients that initiate contraction. Spontaneous Ca²(+) sparks are thought to play a major role in SR Ca²(+) leak. In this study, we determined the quantitative contribution of sparks to SR Ca²(+) leak and tested the hypothesis that non-spark mediated Ca²(+) release also contributes to SR Ca²(+) leak. We simultaneously measured spark properties and intra-SR free Ca²(+) ([Ca²(+)](SR)) after complete inhibition of SERCA with thapsigargin in permeabilized rabbit ventricular myocytes. When [Ca²(+)](SR) declined to 279 ± 10 μm, spark activity ceased completely; however SR Ca²(+) leak continued, albeit at a slower rate. Analysis of sparks and [Ca²(+)](SR) revealed, that SR Ca²(+) leak increased as a function of [Ca²(+)](SR), with a particularly steep increase at higher [Ca²(+)](SR) ( >600 μm) where sparks become a major pathway of SR Ca²(+) leak. At low [Ca²(+)](SR) (< 300 μm), however, Ca²(+) leak occurred mostly as non-spark-mediated leak. Sensitization of ryanodine receptors (RyRs) with low doses of caffeine increased spark frequency and SR Ca²(+) leak. Complete inhibition of RyR abolished sparks and significantly decreased SR Ca²(+) leak, but did not prevent it entirely, suggesting the existence of RyR-independent Ca²(+) leak. Finally, we found that RyR-mediated Ca²(+) leak was enhanced in myocytes from failing rabbit hearts. These results show that RyRs are the main, but not sole contributor to SR Ca²(+) leak. RyR-mediated leak occurs in part as Ca²(+) sparks, but there is clearly RyR-mediated but Ca²(+) sparks independent leak.

Severe food restriction induces myocardial dysfunction related to SERCA2 activity

Previous studies have shown that food restriction promotes myocardial dysfunction in rats. However, the molecular mechanisms that are responsible are unclear. We investigated the role of sarcoplasmic reticulum Ca2+-ATPase (SERCA2) on myocardial performance in food-restricted rats. Male Wistar-Kyoto rats, 60 days old, were fed a control or restricted diet (daily energy intake reduced to 50% of the control) for 90 days. Expression of Serca2a, phospholamban (PLB), Na+/Ca2+ exchanger (NCX), and thyroid hormone receptor (TRalpha1, TRbeta1) mRNA was determined by quantitative PCR. SERCA2 activity was measured by using 20 micromol/L cyclopiazonic acid (CPA) in a left ventricular papillary muscle preparation during isometric contraction in basal conditions and during post-rest contraction. Serum concentrations of thyroxine (T4) and thyrotropin (TSH) were also determined. The 50%-restricted diet reduced body and ventricular weight and serum T4 and TSH levels. The interaction of CPA and food restriction reduced peak developed tension and maximum rate of tension decline (-dT/dt), but increased the resting tension intensity response during post-rest contraction. PLB and NCX mRNA were upregulated and TRalpha1 mRNA was downregulated by food restriction. These results suggest that food restriction promotes myocardial dysfunction related to impairment of sarcoplasmic reticulum Ca2+ uptake as a result of a hypothyroid state.

Junctin – the quiet achiever

Recent advances in understanding skeletal and cardiac muscle function have evolved with recognition of the active role played by the intracellular sarcoplasmic reticulum (SR) Ca2+ store in contraction. The key proteins in this store are the Ca2+ binding protein calsequestrin (CSQ), the ryanodine receptor (RyR) Ca2+ release channel and triadin and junctin (Beard et al. 2004). The CSQ–triadin–junctin–RyR complex (Fig. 1) in the SR lumen forms a ‘Ca2+ transduction machine’ that is central to EC coupling and to normal muscle development. Other proteins in the lumen of the SR, including the histidine rich calcium binding protein (HRC) (Suk et al. 1999), JP-45 (Anderson et al. 2003) and SRP-27 (Bleunven et al. 2008), must also contribute to control of SR intraluminal Ca2+ load, but the precise nature of their role remains undetermined. JP-45 in particular is ideally placed to communicate store load to the excitation–contraction (EC) coupling process as it binds to both CSQ and the dihydropyridine receptor (DHPR) in the surface/transverse tubule membrane. The importance of the luminal proteins has been underlined by the recent discovery that changes in Ca2+ signalling due to mutations in CSQ or to lack of its expression can result in sudden cardiac death (Viatchenko-Karpinski et al. 2004). Furthermore, studies in animal models show that changes in CSQ, junctin, triadin and HRC expression can lead to defective Ca2+ signalling. The review by Pritchard & Kranius (2009) examines the roles of junctin and HRC in Ca2+ cycling and their potential significance in heart failure. This commentary focuses on protein-protein interactions between CSQ, triadin, junctin and RyR proteins that may underlie their roles in Ca2+ cycling. Figure 1 Illustration of the possible interactions between triadin, junctin, CSQ1 and RyR1 in the lumen of the SR

Upregulation Of Cam Kinase IIδ Modulates Spontaneous Ca2+ Wave Properties In A Rabbit Model Of Heart Failure

In ventricular cardiomyocytes, CaMKIIδ (calcium/calmodulin-dependent protein kinase IIδ) is known to modulate Ca2+-handling proteins of the sarcoplasmic reticulum (SR). Dysregulation of Ca2+-handling is evident in cardiomyopathy and may be mediated by changes in CaMKIIδ expression and/or activity. The present study assesses CaMKIIδ expression and activity in rabbit left ventricular(LV) whole homogenates one week after coronary artery ligation(CAL). Changes in CaMKIIδ are translated to changes in SR function in isolated LV cardiomyocytes.Quantitative immunoblotting of CaMKIIδ protein revealed expression was increased by ∼2-fold in CAL (0.096±0.01(sham)vs.0.214±0.042(CAL) normalised mean ratio, (n=7). Similarly, CaMKIIδ activity was increased by ∼1.5-fold in CAL (0.168±0.022 (sham) vs. 0.247±0.028 (CAL) pmolPO4−inc/min/μg protein, n=7). In isolated permeabilised cardiomyocytes, spontaneous Ca2+ waves were studied to assess changes in function associated with upregulation of CaMKIIδ. This was accomplished by measuring changes in Ca2+ wave properties in the presence and absence of AIP (autocamtide-2-related inhibitory peptide). Cells were incubated with a modest concentration (300nM) of AIP for >30 min before use; these were then loaded with fluo-3 and fluorescence was monitored by confocal linescan microscopy with subsequent conversion to Ca2+. In cells from sham animals, no changes in Ca2+ waves were observed in the presence of AIP. In cells from LVD animals AIP caused Ca2+ waves to be reduced in frequency (-20.8±3.4%), increased minimum Ca2+ (+17.4±5.0%) as well as increased rate of decline (+15.8±2.0%). These changes are consistent with CaMKIIδ playing an increased role in SR Ca2+ handling following CAL. Since CaMKIIδ expression and activity are both significantly increased in this model, increased SR Ca2+ handling could occur via CaMKII-mediated effects on SERCA activity producing increased SR accumulation of Ca2+. This would result in an elevated sensitivity to β-adrenergic stimulation that could be arrhythmogenic.

Contribution of sarcoplasmic reticular calcium to smooth muscle contractile activation: gestational dependence in isolated rat uterus.

1. The contribution of Ca2+ released from the sarcoplasmic reticulum (SR) to smooth muscle contractile activation remains poorly understood. By simultaneously monitoring cytosolic [Ca2+] ([Ca2+]i) and force in isolated rat uterine smooth muscle, we report the influence of SR Ca2+ release on contractility during conditions (a) of altered SR Ca2+ homeostasis and (b) where the only source of activating Ca2+ was derived from the SR. 2. In myometria of non-pregnant rats, ryanodine (1-50 microM), a modulator of calcium-induced calcium release (CICR), had no effect on the spontaneous [Ca2+]i or force transients. However, depletion of SR Ca2+ by inhibiting the SR Ca2+-ATPase (with cyclopiazonic acid (CPA), 20 microM) resulted in an enhancement of spontaneous [Ca2+]i and force transients. 3. In myometria of pregnant rats, although ryanodine had no effect in 40% of tissues studied it produced a small but significant enhancement of the integrated spontaneous [Ca2+]i and force transient in 60% of cases. The potentiating effects of CPA were enhanced in myometria of pregnant rats compared with non-pregnant rats, often resulting in maintained [Ca2+]i increases and contraction. 4. In zero external Ca2+, agonist-induced SR Ca2+ release resulted in transient increases in [Ca2+]i and force. The magnitude of these agonist-induced [Ca2+]i and force changes were significantly enhanced in myometria of pregnant rats. No evidence for agonist-induced Ca2+-independent force production was observed. 5. These results indicate that CICR plays little role in SR Ca2+ release from the myometrium, and that there are gestational-dependent alterations in the ability of SR Ca2+ mobilization to contribute to contractile activation. The implications of these findings for the co-ordination of myometrial [Ca2+]i signalling and contractility are discussed.

P-415 - Comparison of the role of sarcoplasmic reticulum Ca2+ pump of esophageal striated musculature of SHRSP and WKY

P-415 - Comparison of the role of sarcoplasmic reticulum Ca2+ pump of esophageal striated musculature of SHRSP and WKY