Reticulum Calcium Release Channel Research Articles

Can the Arrangement of RyR2 in Cardiac Muscle Be Predicted?

Members of the two families of endo/sarcoplasmic reticulum calcium release channels, ryanodine receptors (RyRs) and IP3 receptors, share the same general architecture. They consist of an ionic channel domain inserted into an internal cell membrane, coupled to a very large cytoplasmic domain through which the channels are controlled and then interact with each other. RyRs were originally identified and purified based on the high affinity for the plant alkaloid ryanodine, defined by the work of I.

Effects of 50 Hz sinusoidal magnetic field on Ca2+ release channel ryanodine receptor of sarcoplasmic reticulum vesicles

To investigate the effects of sinusoidal magnetic field on isolated sarcoplasmic reticulum (SR) calcium release channel (RyR1) function. With the Ca2+ dynamic spectrum and isotope labeled methods, the Ca2+ release and [(3)H]-Ryanodine binding, the initial rates of NADH oxidation and the production of superoxide of SR exposed to 50 Hz sinusoidal magnetic field (MF) were investigated respectively. 0.4 mT, 50 Hz sinusoidal MF exposure for 30 min increased SR Ca2+ release initial rate about 35% from (10.82 +/- 0.89) pmol.mg(-1) pro.s(-1) to (14.69 +/- 1.21) pmol.mg(-1) pro.s(-1); and the [(3)H]-Ryanodine binding by about 15% from (2.13 +/- 0.05) pmol/mg pro to (2.45 +/- 0.07) pmol/mg pro, which regulated by 1 mmol/L NADH with 1 mmol/L NAD+. Meanwhile MF upregulated the rate of NADH oxidation by about 22% from (0.88 +/- 0.11) x 10(-4) FI/s to (1.07 +/- 0.13) x 10(-4) FI/s and upregulated the production of superoxide by about 32% from (0.99 +/- 0.09) x 10(-5) FI/s to (1.31 +/- 0.06) x 10(-5) FI/s. 0.4 mT sinusoidal MF increases the activity of RyR1 within the low redox potential environment, and promotes NADH oxidase activity and superoxide production.

Calcium Handling in Human Embryonic Stem Cell-Derived Cardiomyocytes

The objective of the current study was to characterize calcium handling in developing human embryonic stem cell-derived cardiomyocytes (hESC-CMs). To this end, real-time polymerase chain reaction (PCR), immunocytochemistry, whole-cell voltage-clamp, and simultaneous patch-clamp/laser scanning confocal calcium imaging and surface membrane labeling with di-8-aminonaphthylethenylpridinium were used. Immunostaining studies in the hESC-CMs demonstrated the presence of the sarcoplasmic reticulum (SR) calcium release channels, ryanodine receptor-2, and inositol-1,4,5-trisphosphate (IP3) receptors. Store calcium function was manifested as action-potential-induced calcium transients. Time-to-target plots showed that these action-potential-initiated calcium transients traverse the width of the cell via a propagated wave of intracellular store calcium release. The hESC-CMs also exhibited local calcium events ("sparks") that were localized to the surface membrane. The presence of caffeine-sensitive intracellular calcium stores was manifested following application of focal, temporally limited puffs of caffeine in three different age groups: early-stage (with the initiation of beating), intermediate-stage (10 days post-beating [dpb]), and late-stage (30-40 dpb) hESC-CMs. Calcium store load gradually increased during in vitro maturation. Similarly, ryanodine application decreased the amplitude of the spontaneous calcium transients. Interestingly, the expression and function of an IP3-releasable calcium pool was also demonstrated in the hESC-CMs in experiments using caged-IP3 photolysis and antagonist application (2 microM 2-Aminoethoxydiphenyl borate). In summary, our study establishes the presence of a functional SR calcium store in early-stage hESC-CMs and shows a unique pattern of calcium handling in these cells. This study also stresses the importance of the functional characterization of hESC-CMs both for developmental studies and for the development of future myocardial cell replacement strategies. Disclosure of potential conflicts of interest is found at the end of this article.

Cardiomyopathies and sudden cardiac death caused by RyR2 mutations: Are the channels the beginning and the end?☆

See article by Milting et al. [3] (pages 496–505) in this issue. A basic goal of medicine is to cure diseases. Acquired diseases may sometimes be prevented by various methods of exercising, by the “healthy way” of living. Inherited diseases are different; their effective treatment (provided that there is any) requires the recognition of the disease in a sufficiently early stage of life. One – and probably the most promising – way to do so is to establish the link between disorders and genetic alterations. This can be achieved by mapping the mutations corresponding to a given disease, and in the past decade a large number of mutations affecting molecules involved in calcium handling of skeletal and cardiac muscles, including the ryanodine receptor, have been identified [1]. Regarding the cardiac sarcoplasmic reticulum (SR) calcium release channel (RyR2), one of the first published mutations was presented in the pioneering work of Laitinen et al. [2]. In that study, three different RyR2 mutations (P2328S, Q4201R, and V4653F) were associated with familiar polymorphic ventricular tachycardia (FPVT), and each of them was shown to fully co-segregate with the characteristic arrhythmic phenotype. The importance of this kind of knowledge cannot be overestimated, since it makes it possible to recognize a hidden inherited cardiac disorder that exhibits no visible sign of myocardial damage while threatening with close to 30% mortality due to sudden cardiac death (SCD) upon heavy exercise. The article by Milting et al. in this issue of Cardiovascular Research reports two new single nucleotide polymorphisms (G1885E and G1886S) … * Corresponding author. Tel.: +36 52 416634; fax: +36 52 432289. Email address: nanasi{at}phys.dote.hu

The analysis of ultrastructure and gene expression of sarco/endoplasmic reticulum calcium handling proteins in alloxan-induced diabetic rat myocardium

Objective — The change of mechanical properties of isolated diabetic rat papillary muscle, myocardial ultrastructure and gene expression of sarco/endoplasic reticulum calcium handling proteins in alloxan-induced diabetic rat heart was investigated.Methods — Diabetes was induced in male Sprague-Dawley rats (180~220 g) by administering a single tail-vein injection (40 mg/kg) of alloxan. The control group was injected with normal saline. At the end of 2, 4 and 6 weeks after the induction of diabetes, the right ventricular papillary muscles were isolated and perfused with oxygen saturated Tyrode solution for assessment of the contractile function. Light and electron microscopic analysis was used to analyse the myocardial ultrastructure in rats six weeks after induction. Gene expression of calcium handling proteins was measured by reverse transcription-polymerase chain reaction and Western blot.Results — In the diabetic rats, +dT/dtmax and –dT/dtmax were decreased (P < 0.01), while 1/2 diastolic intervals were longer than control at the end of 4 and 6 weeks (P < 0.01). Electron microscopic analysis of the myocardium revealed a spectrum of subcellular remodelling which was characterized by damaged myofibrils and mitochondria and dilated swollen sarcoplasmic reticulum. The levels of both mRNA and protein of phospholamban were significantly increased, whereas 1, 4, 5-trisphophate inositol receptor type 2, ryanodine receptor type 2 mRNAs were significantly decreased compared with that in the age-matched 6 weeks control rats. In contrast, levels of both mRNA and protein of sarco/endoplasic reticulum Ca2+-ATPase were not affected.Conclusions — Contractile dysfunction of papillary muscles and subcellular remodelling exist in alloxan-induced diabetic rat myocardium. Up-regulation of phospholamban and down-regulation of sarco/endoplasic reticulum calcium release channel may be the molecular mechanism.

Cardiac Type 2 Inositol 1,4,5-Trisphosphate Receptor: INTERACTION AND MODULATION BY CALCIUM/CALMODULIN-DEPENDENT PROTEIN KINASE II

The type 2 inositol 1,4,5-trisphosphate receptor (InsP(3)R2) was identified previously as the predominant isoform in cardiac ventricular myocytes. Here we reported the subcellular localization of InsP(3)R2 to the cardiomyocyte nuclear envelope (NE). The other major known endo/sarcoplasmic reticulum calcium-release channel (ryanodine receptor) was not localized to the NE, indicating functional segregation of these channels and possibly a unique role for InsP(3)R2 in regulating nuclear calcium dynamics. Immunoprecipitation experiments revealed that the NE InsP(3)R2 associates with Ca(2+)/calmodulin-dependent protein kinase IIdelta (CaMKIIdelta), the major isoform expressed in cardiac myocytes. Recombinant InsP(3)R2 and CaMKIIdelta(B) also co-immunoprecipitated after co-expression in COS-1 cells. Additionally, the amino-terminal 1078 amino acids of the InsP(3)R2 were sufficient for interaction with CaMKIIdelta(B) and associated upon mixing following separate expression. CaMKII can also phosphorylate InsP(3)R2, as demonstrated by (32)P labeling. Incorporation of CaMKII-treated InsP(3)R2 into planar lipid bilayers revealed that InsP(3)-mediated channel open probability is significantly reduced ( approximately 11 times) by phosphorylation via CaMKII. We concluded that the InsP(3)R2 and CaMKIIdelta likely represent two central components of a multiprotein signaling complex, and this raises the possibility that calcium release via InsP(3)R2 in the myocyte NE may activate local CaMKII signaling, which may feedback on InsP(3)R2 function.