Rationale Cardiovascular disease represents a global pandemic. The advent of and recent advances in mouse genomics, epigenomics, and transgenics offer ever-greater potential for powerful avenues of research. However, progress is often constrained by unique complexities associated with the isolation of viable myocytes from the adult mouse heart. Current protocols rely on retrograde aortic perfusion using specialized Langendorff apparatus, which poses considerable logistical and technical barriers to researchers and demands extensive training investment. Objective To identify and optimize a convenient, alternative approach, allowing the robust isolation and culture of adult mouse cardiac myocytes using only common surgical and laboratory equipment. Methods and Results Cardiac myocytes were isolated with yields comparable to those in published Langendorff-based methods, using direct needle perfusion of the LV ex vivo and without requirement for heparin injection. Isolated myocytes can be cultured antibiotic free, with retained organized contractile and mitochondrial morphology, transcriptional signatures, calcium handling, responses to hypoxia, neurohormonal stimulation, and electric pacing, and are amenable to patch clamp and adenoviral gene transfer techniques. Furthermore, the methodology permits concurrent isolation, separation, and coculture of myocyte and nonmyocyte cardiac populations. Conclusions We present a novel, simplified method, demonstrating concomitant isolation of viable cardiac myocytes and nonmyocytes from the same adult mouse heart. We anticipate that this new approach will expand and accelerate innovative research in the field of cardiac biology.

BackgroundThe left atrial posterior wall (LAPW) is potentially an important area for the development and maintenance of atrial fibrillation. We assessed whether there are regional electrical differences throughout the murine left atrial myocardium that could underlie regional differences in arrhythmia susceptibility.MethodsWe used high-resolution optical mapping and sharp microelectrode recordings to quantify regional differences in electrical activation and repolarisation within the intact, superfused murine left atrium and quantified regional ion channel mRNA expression by Taqman Low Density Array. We also performed selected cellular electrophysiology experiments to validate regional differences in ion channel function.ResultsSpontaneous ectopic activity was observed during sustained 1Hz pacing in 10/19 intact LA and this was abolished following resection of LAPW (0/19 resected LA, P<0.001). The source of the ectopic activity was the LAPW myocardium, distinct from the pulmonary vein sleeve and LAA, determined by optical mapping. Overall, LAPW action potentials (APs) were ca. 40% longer than the LAA and this region displayed more APD heterogeneity. mRNA expression of Kcna4, Kcnj3 and Kcnj5 was lower in the LAPW myocardium than in the LAA. Cardiomyocytes isolated from the LAPW had decreased Ito and a reduced IKACh current density at both positive and negative test potentials.ConclusionsThe murine LAPW myocardium has a different electrical phenotype and ion channel mRNA expression profile compared with other regions of the LA, and this is associated with increased ectopic activity. If similar regional electrical differences are present in the human LA, then the LAPW may be a potential future target for treatment of atrial fibrillation.

AngII infusion caused two-fold increase in ROS production of WT hearts (p<0.05) (but not p47 phox KO mice), which was inhibited significantly by diphenyleneiodonium (DPI, a flavoprotein inhibitor) or superoxide dismutase, significantly but slightly by NG-nitro-l-arginine methyl ester (L-NAME, a nitric oxide synthase inhibitor), but not by rotenone (mitochondrial respiratory chain inhibitor) or oxypurinol (xanthine oxidase inhibitor). Increased ROS production in WT AngII-infused hearts was accompanied by significant phosphorylation of ERK1/2. In conclusion, p47 phox and p47 phox signalling through ERK1/2 play an important role in AngII-induced cardiac hypertrophy. Introduction Hyperglycemia-induced ROS generation within mitochondria plays a major role in the development of diabetic complications. Mitochondria are one of the most important cell organelles in diabetes research because of its crucial role as a regulator of energy balance. The present study was aimed to evaluate the effect galangin, a flavonoid, on oxidative mitochondrial damage in in streptozotocin (STZ)-induced diabetic rats. Materials and methods Diabetes was induced by intraperitoneal administration of low dose of STZ (40 mg/kg body weight (BW)) into male albino Wistar rats. Galangin (8 mg/kg BW) or glibenclamide (600 mg/kg BW) was given orally daily once for 45 days to normal and STZ-induced diabetic rats. Results Diabetic rats showed a significant (p<0.05) increase in kidney and heart mitochondrial oxidant (Thiobarbituric acid reactive substance) levels and a significant decrease in enzymatic (superoxide dismutase, glutathione peroxidase) and nonenzymatic (reduced glutathione) antioxidants levels as compared to control rats. The activities of mitochondrial enzymes such as isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, and malate dehydrogenase and mitochondrial respiratory chain enzymes such as NADH dehydrogenase and Cytochrome c-oxidase were decreased significantly (p<0.05) in diabetic rats as compared to control rats. Administration of galangin to diabetic rats resulted in the following findings as compared to diabetic control rats: the oxidant levels decreased significantly (p<0.05); the enzymatic and non-enzymatic antioxidants levels increased significantly (p<0.05); and the function of mitochondrial enzymes and the mitochondrial respiratory chain enzymes increased significantly (p<0.05).Conclusion From the results, we conclude that galangin could maintain kidney and heart mitochondrial function in diabetic rats.

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