https://ogma.newcastle.edu.au/vital/access/ /manager/Index ${session.getAttribute("locale")} 5 https://ogma.newcastle.edu.au/vital/access/ /manager/Repository/uon:34969 d = 121 ± 14 nM. Ex-vivo phosphorylation/dephosphorylation experiments suggested that the divergent CaM regulation of healthy and failing human RyR2 was caused by differences in RyR2 phosphorylation by protein kinase A and Ca-CaM-dependent kinase II. Ca²⁺-spark measurements in murine cardiomyocytes harbouring RyR2 phosphomimetic or phosphoablated mutants at S2814 and S2808 suggest that phosphorylation of residues corresponding to either human RyR2-S2808 or S2814 is both necessary and sufficient for RyR2 regulation by CaM. Our results challenge the current concept that CaM universally functions as a canonical inhibitor of RyR2 across species. Rather, CaM's biological action on human RyR2 appears to be more nuanced, with inhibitory activity only on phosphorylated RyR2 channels, which occurs during exercise or in patients with heart failure.]]> Wed 24 Jun 2020 11:42:42 AEST ]]> https://ogma.newcastle.edu.au/vital/access/ /manager/Repository/uon:18980 i = 10 µM at 100 nM Ca²⁺) that was similar to RyR2 from rat and sheep obtained under the same experimental conditions. However, in the presence of 0.1 mM Ca²⁺, RyR2s from human were 3.5-fold less sensitive to cytoplasmic Mg²⁺ inhibition than those from sheep and rat. The Kₐ values for luminal Ca²⁺ activation were similar in the three species (35 µM for human, 12 µM for sheep, and 10 µM for rat). From the relationship between open probability and luminal [Ca²⁺], the peak open probability for the human RyR2 was approximately the same as that for sheep, and both were ~10-fold greater than that for rat RyR2. Human RyR2 also showed the same sensitivity to luminal Mg²⁺ as that from sheep, whereas rat RyR2 was 10-fold more sensitive. In all species, modulation of RyR2 gating by luminal Ca²⁺ and Mg²⁺ only occurred when cytoplasmic [Ca²⁺] was <3 µM. The activation response of RyR2 to luminal and cytoplasmic Ca²⁺ was strongly dependent on the Mg²⁺ concentration. Addition of physiological levels (1 mM) of Mg²⁺ raised the Kₐ for cytoplasmic Ca²⁺ to 30 µM (human and sheep) or 90 µM (rat) and raised the Kₐ for luminal Ca²⁺ to ~1 mM in all species. This is the first report of the regulation by Ca²⁺ and Mg²⁺ of native RyR2 receptor activity from healthy human hearts.]]> Sat 24 Mar 2018 07:58:52 AEDT ]]> https://ogma.newcastle.edu.au/vital/access/ /manager/Repository/uon:46291 2+-activated K⁺ channel, K_Ca1.1, and lies within a linkage interval for atrial fibrillation (AF). Insights into the cardiac functions of K_Ca1.1 are limited, and KCNMA1 has not been investigated as an AF candidate gene. Methods; The KCNMA1 gene was sequenced in 118 patients with familial AF. The role of K_Ca1.1 in normal cardiac structure and function was evaluated in humans, mice, zebrafish, and fly. A novel KCNMA1 variant was functionally characterized.Results: A complex KCNMA1 variant was identified in 1 kindred with AF. To evaluate potential disease mechanisms, we first evaluated the distribution of K_Ca1.1 in normal hearts using immunostaining and immunogold electron microscopy. K_Ca1.1 was seen throughout the atria and ventricles in humans and mice, with strong expression in the sinus node. In an ex vivo murine sinoatrial node preparation, addition of the K_Ca1.1 antagonist, paxilline, blunted the increase in beating rate induced by adrenergic receptor stimulation. Knockdown of the K_Ca1.1 ortholog, kcnma1b, in zebrafish embryos resulted in sinus bradycardia with dilatation and reduced contraction of the atrium and ventricle. Genetic inactivation of the Drosophila K_Ca1.1 ortholog, slo, systemically or in adult stages, also slowed the heartbeat and produced fibrillatory cardiac contractions. Electrophysiological characterization of slo-deficient flies revealed bursts of action potentials, reflecting increased events of fibrillatory arrhythmias. Flies with cardiac-specific overexpression of the human KCNMA1 mutant also showed increased heart period and bursts of action potentials, similar to the K_Ca1.1 loss-of-function models. Conclusions: Our data point to a highly conserved role of K_Ca1.1 in sinus node function in humans, mice, zebrafish, and fly and suggest that K_Ca1.1 loss of function may predispose to AF.]]> Mon 14 Nov 2022 16:44:58 AEDT ]]>