kn-93 and Myocardial-Infarction

Topics: Animals; Autophagy; Beclin-1; Benzylamines; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Death; Heart Function Tests; Humans; In Vitro Techniques; Myocardial Infarction; Myocardial Reperfusion Injury; Myocytes, Cardiac; Phosphorylation; Protein Kinase Inhibitors; Rats; RNA, Small Interfering; Sulfonamides

Diastolic spontaneous Ca(2+) waves (DCWs) are recognized as important contributors to triggered arrhythmias. DCWs are thought to arise when [Ca(2+)] in sarcoplasmic reticulum ([Ca(2+)](SR)) reaches a certain threshold level, which might be reduced in cardiac disease as a consequence of sensitization of ryanodine receptors (RyR2s) to luminal Ca(2+).. We investigated the mechanisms of DCW generation in myocytes from normal and diseased hearts, using a canine model of post-myocardial infarction ventricular fibrillation (VF).. The frequency of DCWs, recorded during periodic pacing in the presence of a β-adrenergic receptor agonist isoproterenol, was significantly higher in VF myocytes than in normal controls. Rather than occurring immediately on reaching a final [Ca(2+)](SR), DCWs arose with a distinct time delay after attaining steady [Ca(2+)](SR) in both experimental groups. Although the rate of [Ca(2+)](SR) recovery after the SR Ca(2+) release was similar between the groups, in VF myocytes the latency to DCWs was shorter, and the [Ca(2+)](SR) at DCW initiation was lower. The restitution of depolarization-induced Ca(2+) transients, assessed by a 2-pulse protocol, was significantly faster in VF myocytes than in controls. The VF-related alterations in myocyte Ca(2+) cycling were mimicked by the RyR2 agonist, caffeine. The reducing agent, mercaptopropionylglycine, or the CaMKII inhibitor, KN93, decreased DCW frequency and normalized restitution of Ca(2+) release in VF myocytes.. The attainment of a certain threshold [Ca(2+)](SR) is not sufficient for the generation of DCWs. Postrelease Ca(2+) signaling refractoriness critically influences the occurrence of DCWs. Shortened Ca(2+) signaling refractoriness due to RyR2 phosphorylation and oxidation is responsible for the increased rate of DCWs observed in VF myocytes and could provide a substrate for synchronization of arrhythmogenic events at the tissue level in hearts prone to VF.

Topics: Adrenergic beta-Agonists; Animals; Benzylamines; Caffeine; Calcium Channel Agonists; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cardiac Pacing, Artificial; Death, Sudden, Cardiac; Disease Models, Animal; Dogs; Excitation Contraction Coupling; Female; Isoproterenol; Male; Myocardial Infarction; Myocytes, Cardiac; Oxidation-Reduction; Phosphorylation; Protein Kinase Inhibitors; Reaction Time; Reducing Agents; Ryanodine Receptor Calcium Release Channel; Sarcoplasmic Reticulum; Sulfonamides; Time Factors; Tiopronin; Ventricular Fibrillation

Oestrogen confers cardioprotection by down-regulating the beta(1)-adrenoceptor and suppressing the expression and activity of protein kinase A. We hypothesized that oestrogen may also protect the heart by suppressing Ca(2+)/calmodulin-dependent protein kinase II (CaMKII), another signalling messenger activated by the beta(1)-adrenoceptor, that enhances apoptosis.. We first determined the expression of CaMKII in the heart from sham and ovariectomized rats with and without oestrogen replacement. We then determined the effects of CaMKII inhibition (KN93, 2.5 micromolxL(-1)) in the presence or absence of 10(-7) molxL(-1) isoprenaline, a non-selective beta-adrenoceptor agonist. We also determined the percentage apoptosis in myocytes from rats in each group with or without beta-adrenoceptor stimulation.. Both CaMKIIdelta and phosphorylated CaMKII were up-regulated in the hearts from ovariectomized rats, and they were restored to normal by oestrogen replacement. The infarct size and lactate dehydrogenase release were significantly greater after ovariectomy. Similarly, cardiac contractility, the amplitude of the electrically induced intracellular Ca(2+) transient and the number of apoptotic cells were also greater in ovariectomized rats upon ischaemia/reperfusion in the presence or absence of isoprenaline. Most importantly, the responses to ischaemic insult in ovariectomized rats were reversed not only by oestrogen replacement, but by blockade of CaMKII with KN93.. Oestrogen confers cardioprotection at least partly by suppressing CaMKIIdelta. This effect of oestrogen on CaMKII is independent of the beta-adrenoceptor and occurs in addition to down-regulation of the receptor.

Topics: Adrenergic beta-Agonists; Animals; Apoptosis; Benzylamines; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cells, Cultured; Disease Models, Animal; Down-Regulation; Drug Implants; Estrogens; Female; Isoproterenol; Myocardial Contraction; Myocardial Infarction; Myocardial Reperfusion Injury; Myocytes, Cardiac; Ovariectomy; Phosphorylation; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, beta; Recovery of Function; Sulfonamides

The cardiac ryanodine receptor (RyR2)/calcium release channel on the sarcoplasmic reticulum is required for muscle excitation-contraction coupling. Using site-directed mutagenesis, we identified the specific Ca2+/calmodulin-dependent protein kinase II (CaMKII) phosphorylation site on recombinant RyR2, distinct from the site for protein kinase A (PKA) that mediates the "fight-or-flight" stress response. CaMKII phosphorylation increased RyR2 Ca2+ sensitivity and open probability. CaMKII was activated at increased heart rates, which may contribute to enhanced Ca2+-induced Ca2+ release. Moreover, rate-dependent CaMKII phosphorylation of RyR2 was defective in heart failure. CaMKII-mediated phosphorylation of RyR2 may contribute to the enhanced contractility observed at higher heart rates. The full text of this article is available online at http://circres.ahajournals.org.

Topics: Adrenergic beta-Agonists; Amino Acid Sequence; Animals; Benzylamines; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Cardiac Pacing, Artificial; Cyclic AMP-Dependent Protein Kinases; Enzyme Inhibitors; Heart Failure; Heart Rate; Humans; Isoproterenol; Molecular Sequence Data; Mutagenesis, Site-Directed; Myocardial Infarction; Myocardium; Phosphorylation; Phosphoserine; Protein Processing, Post-Translational; Rabbits; Rats; Rats, Sprague-Dawley; Recombinant Fusion Proteins; Ryanodine Receptor Calcium Release Channel; Sequence Alignment; Sequence Homology, Amino Acid; Sulfonamides; Tacrolimus Binding Proteins; Ultrasonography