Ryanodine receptor studies using genetically engineered mice

AbstractRyanodine receptors (RyR) regulate intracellular Ca2+ release in many cell types and have been implicated in a number of inherited human diseases. Over the past 15years genetically engineered mouse models have been developed to elucidate the role that RyRs play in physiology and pathophysiology. To date these models have implicated RyRs in fundamental biological processes including excitation–contraction coupling and long term plasticity as well as diseases including malignant hyperthermia, cardiac arrhythmias, heart failure, and seizures. In this review we summarize the RyR mouse models and how they have enhanced our understanding of the RyR channels and their roles in cellular physiology and disease

Analysis of Cardiac Ion Channels to Understand Arrhythmias Which Lead to Sudden Cardiac Death

There are 300,000-400,000 fatalities attributed to sudden cardiac death every year in the U.S. due to a lack of sufficient research on mechanisms causing arrhythmias1. Malfunctions with the ion channels in the heart may lead to lethal arrhythmias. The purpose of this work is to study ion channels and evaluate malfunctions relative to normally functioning hearts. Plasmid insertion in E. coli assayed whether functional ion channels reach the membrane, and confocal fluorescent microscopy was used to illuminate cellular functionality. In addition, genetic analysis was used to determine the extent of hereditary factors in sudden cardiac death. Genes that encode for the voltage-gated sodium, potassium, and calcium ion channels were analyzed at the genetic level using isolated DNA samples and traditional Sanger sequencing methods to identify mutations that may be responsible for sudden cardiac death syndromes. For example, Long QT syndrome, Short QT syndrome, and Brugada syndrome are caused by mutations in these ion channels. Once these mutations are identified, genetic engineering techniques can be used in the generation of new heart cells from the stem cells found in somatic tissue. Generation of such heart cells is important because it could lead to the development of personalized treatment for degenerative diseases such as heart failure in the future. Rubart, M. et al., Mechanisms of Sudden Cardiac Death, 2005. J. clin. invest. 115(9):2305-2315

Leaky ryanodine receptors in β-sarcoglycan deficient mice: a potential common defect in muscular dystrophy

Disruption of the sarcolemma-associated dystrophin-glycoprotein complex underlies multiple forms of muscular dystrophy, including Duchenne muscular dystrophy and sarcoglycanopathies. A hallmark of these disorders is muscle weakness. In a murine model of Duchenne muscular dystrophy, mdx mice, cysteine-nitrosylation of the calcium release channel/ryanodine receptor type 1 (RyR1) on the skeletal muscle sarcoplasmic reticulum causes depletion of the stabilizing subunit calstabin1 (FKBP12) from the RyR1 macromolecular complex. This results in a sarcoplasmic reticular calcium leak via defective RyR1 channels. This pathological intracellular calcium leak contributes to reduced calcium release and decreased muscle force production. It is unknown whether RyR1 dysfunction occurs also in other muscular dystrophies. To test this we used a murine model of Limb-Girdle muscular dystrophy, deficient in β-sarcoglycan (Sgcb−/−). Skeletal muscle RyR1 from Sgcb−/− deficient mice were oxidized, nitrosylated, and depleted of the stabilizing subunit calstabin1, which was associated with increased open probability of the RyR1 channels. Sgcb−/− deficient mice exhibited decreased muscle specific force and calcium transients, and displayed reduced exercise capacity. Treating Sgcb−/− mice with the RyR stabilizing compound S107 improved muscle specific force, calcium transients, and exercise capacity. We have previously reported similar findings in mdx mice, a murine model of Duchenne muscular dystrophy. Our data suggest that leaky RyR1 channels may underlie multiple forms of muscular dystrophy linked to mutations in genes encoding components of the dystrophin-glycoprotein complex. A common underlying abnormality in calcium handling indicates that pharmacological targeting of dysfunctional RyR1 could be a novel therapeutic approach to improve muscle function in Limb-Girdle and Duchenne muscular dystrophies