NO donors were found to reduce the rate of Ca2+ release from isolated skeletal muscle sarcoplasmic reticulum (SR) and open probability single ryanodine receptor channels (RyRCs) in planar lipid bilayers, these effects prevented by quencher hemoglobin reversed 2-mercaptoethanol. assessed homogenates was also reduced that generated situ L-arginine endogenous, nitro-L-arginine methylester-sensitive NO-synthase. The effect on RyRC might explain NO-induced depression contractile force striated...

Rapid, sequence-specific DNA detection is essential for applications in medical diagnostics and genetic screening. Electrical biosensors that use immobilized nucleic acids are especially promising these because of their potential miniaturization automation. Current methods based on sequencing by synthesis rely optical readouts; however, a direct electrical method this technique not available. We report here an approach enzymatically catalyzed induced surface charge perturbation. discovered...

Antidepressants inhibit many membrane receptors and ionic channels, including the L-type calcium channel. Here, we investigated inhibition of current (<i>I</i>_Ca) by antidepressants in enzymatically isolated rat ventricular myocytes using whole-cell patch clamp. The molecular mechanism was studied comparing voltage state dependence antidepressant <i>I</i>_Ca to respective properties antagonists, studying effect...

The local control concept of excitation–contraction coupling in the heart postulates that activity sarcoplasmic reticulum ryanodine receptor channels (RyR) is controlled by Ca 2+ entry through adjoining sarcolemmal single dihydropyridine (DHPRs). One unverified premise this hypothesis RyR must be fast enough to track brief (&lt;0.5 ms) elevations accompanying DHPR channel openings. To define kinetic limits effective trigger signals, we recorded cardiac RyRs lipid bilayers during rapid and...

The synergic effect of luminal Ca2+, cytosolic and adenosine triphosphate (ATP) on activation cardiac ryanodine receptor (RYR2) channels was examined in planar lipid bilayers. dose–response RYR2 gating activity to ATP characterized at a diastolic Ca2+ concentration 100 nM over range concentrations and, vice versa, 1 mM concentrations. Low level (1 mM) significantly increased the affinity channel for but without substantial channel. Higher levels (8–53 markedly amplified effects by...

Despite its importance and abundance of experimental data, the molecular mechanism RyR2 activation by calcium is poorly understood. Recent studies involving coexpression wild-type (WT) together with a mutant deficient in calcium-dependent (Li, P., S.R. Chen. 2001. J. Gen. Physiol. 118:33-44) revealed large variations sensitivity RyR tetramers their monomer composition. Together previous results on kinetics Ca (Zahradníková, A., I. Zahradník, Györke, S. Györke. 1999. 114:787-798), these data...

The principal role of calcium current in the triggering release cardiac myocytes is well recognized. mechanism how (I(Ca)) controls intensity not clear because stochastic nature voltage-dependent gating channels (DHPRs) and calcium-dependent ryanodine receptors (RyRs). To disclose relation between DHPR openings probability release, local activation by I(Ca) was investigated rat ventricular using patch-clamp confocal microscopy. Calcium spikes were activated temporally synchronized triggers,...

Phenylalkylamines, benzothiazepines, and dihydropyridines bind noncompetitively to the L-type calcium channel. The molecular mechanisms of this interaction were investigated in enzymatically isolated rat ventricular myocytes using whole-cell patch-clamp technique. When applied alone, felodipine, verapamil, diltiazem inhibited current with values inhibitory constant (<i>K</i>_B) 11, 246, 512 nM, respectively, whereas...

Cytosolic calcium concentration in resting cardiac myocytes locally fluctuates as a result of spontaneous microscopic Ca2+ releases or abruptly rises an external trigger. These processes, observed sparks, are fundamental for proper function muscle. In this study, we analyze how the characteristics and triggered sparks related to ryanodine receptor (RYR) gating. We show that frequency probability distribution release flux quanta correspond quantitatively predictions allosteric homotetrameric...

Human ryanodine receptor 2 (hRyR2) mediates calcium release from the sarcoplasmic reticulum, enabling cardiomyocyte contraction. The N-terminal region of hRyR2 (amino acids 1-606) is target >30 arrhythmogenic mutations and contains a binding site for phosphoprotein phosphatase 1. Here, solution crystal structures determined under near-physiological conditions, as well homology model region, are presented. N-terminus held together by unique network interactions among its three domains, A, B...

In mammalian cardiac myocytes, calcium released into the dyadic space rapidly inactivates current ( I Ca ). We used this 2+ release-dependent inactivation (RDI) of as a local probe sarcoplasmic reticulum release activation. whole cell patch-clamped rat ventricular entry induced by short prepulses from —50 mV to positive voltages caused suppression peak during test pulse. The negative correlation between and pulse indicated that RDI evoked prepulse affected only channels in those dyads which...

Abstract Cardiac excitation-contraction coupling relies on dyads, the intracellular calcium synapses of cardiac myocytes, where plasma membrane contacts sarcoplasmic reticulum and electrical excitation triggers release. The morphology dyads dynamics local release vary substantially. To better understand correspondence between structure functionality we estimated incidences structurally different kinetically sites tested their responsiveness to experimental myocardial injury in left...

The dyads of cardiac myocytes contain ryanodine receptors (RYRs) that generate calcium sparks upon activation. To test how geometric factors RYR distribution contribute to the formation sparks, which cannot be addressed experimentally, we performed in silico simulations on a large set models release sites (CRSs). Our covered observed range number, density, and spatial arrangement. function CRSs was modeled by openings, with an open probability dependent concentrations free Ca2+ Mg2+ ions,...

Mg 2+ , an important constituent of the intracellular milieu in cardiac myocytes, is known to inhibit ryanodine receptor (RyR) Ca release channels by competing with at cytosolic activation sites channel. However, significance this competition for local, dynamic -signaling processes thought govern excitation-contraction (EC) coupling remains largely unknown. In present study, stimuli different waveforms (i.e., sustained and brief) were generated photolysis caged compound nitrophenyl...

The local calcium release flux signals (calcium spikes) evoked by membrane depolarization were recorded at high temporal resolution (2000 lines s(-1)) in isolated ventricular myocytes of male rats, using combination scanning confocal microscopy and the patch-clamp technique. kinetic properties spikes investigated. time course spike activation could be described reliably a model with higher-order (n = 3) kinetics, but not first-order exponential process. A termination coupled to its was...

Wolframin (Wfs1) is a membrane protein of the sarco/endoplasmic reticulum. Wfs1 mutations are responsible for Wolfram syndrome, characterized by diabetic and neurological symptoms. Although expressed in cardiac muscle, its role this tissue not clear. We have effect invalidation on calcium signalling-related processes isolated ventricular myocytes exon5-Wfs1 deficient rats (Wfs1 e5/ e5) before onset overt disease. Calcium transients contraction were measured field-stimulated using confocal...

Intracellular calcium signals are studied by laser-scanning confocal fluorescence microscopy. The required spatio-temporal resolution makes description of difficult because the low signal-to-noise ratio. We designed a new procedure spike analysis based on their fitting with model. accuracy and precision were tested synthetic datasets generated either randomly varied parameters Gaussian noise constant amplitude, or various amplitudes. Statistical was used to evaluate performance algorithms....

The role of cardiac ryanodine receptor (RyR) gating in the initiation and propagation calcium waves was investigated using a mathematical model comprising stochastic description RyR deterministic diffusion sequestration. We used one-dimensional array equidistantly spaced clusters, representing confocal scanning line, to simulate formation sparks. Our provided an excellent dependence frequency diastolic sparks increased tendency for production after decrease cytosolic buffering. developed...