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1 We investigated the roles of nitric oxide (NO) and endothelin-1 (ET-1) in organ dysfunction in diabetic mice with normal genotype (wild-type, WT) or myocyte-specific overexpression of endothelial NO synthase (eNOS) (transgenic, TG) after chronic oral treatment with the endothelin-A (ETA) receptor antagonist atrasentan. 2 Mice were rendered diabetic by injection of 200 mg kg−1 streptozotocin (STZ). Experimental groups were: untreated WT diabetic (n=9), untreated TG diabetic (n=9), atrasentan-treated WT diabetic (n=9), atrasentan-treated TG diabetic (n=8) and the four corresponding nondiabetic groups (n=5). Atrasentan was administered orally via drinking water at 3 mg kg−1 per day over 28 days. All diabetic mice developed similar hyperglycaemia (27–30 mmol l−1). 3 Atrasentan treatment significantly improved left ventricular systolic and diastolic function in response to exogenous norepinephrine, but there were no differences between genotypes. 4 Atrasentan antagonized the diabetic impairments in endothelium-dependent coronary relaxation and thromboxane-receptor mediated aortic constriction. Further, it improved cardiac and renal oxidant status as evident from reduced tissue malondialdehyde levels. 5 Atrasentan reduced diabetic urine flow, proteinuria and plasma creatinine levels, but creatinine clearance was not significantly altered. 6 These results suggest that in experimental type 1 diabetes, blocking ETA receptors ameliorates myocardial, coronary and renal function and improves tissue oxidant status, whereas raising myocardial NO levels has neither beneficial nor deleterious effects on diabetic cardiomyopathy in this transgenic model. British Journal of Pharmacology (2006) 148, 671–681. doi:10.1038/sj.bjp.0706772

To examine the acute effects of sunitinib on inotropic function, intracellular Ca(2+) transients, myofilament Ca(2+) sensitivity and generation of reactive oxygen species (ROS) in human multicellular myocardium and isolated mouse cardiomyocytes. To search for microRNAs as suitable biomarkers for indicating toxic cardiac effects.After exposure to sunitinib (0.1-10 µg/mL) developed force, diastolic tension and kinetic variables were assessed in isolated human myocardium. Changes in myocyte sarcomere length, whole-cell calcium transients, myofilament force-Ca(2+) relationship, and ROS generation were examined in isolated ventricular mouse cardiomyocytes. Microarray and realtime-PCR were used to screen for differentially expressed microRNAs in cultured cardiomyocytes that were exposed for 24 h to sunitinib.We found that higher concentrations of sunitinib (1 and 10 µg/mL) decreased developed force at 30 minutes 76.9 + 2.8 and 54.5 + 6.3%, compared to 96.1 + 2.6% in controls (P0.01). Sunitinib exposure significantly decreased sarcomere shortening and Ca2+ transients. Myofilament Ca(2+) sensitivity was not altered, while ROS levels were significantly increased after exposure to the drug. MicroRNA expression patterns were not altered by sunitinib.Sunitinib elicits a dose-dependent negative inotropic effect in myocardium, accompanied by a decline in intracellular Ca(2+) and increased ROS generation. In clinical practice, these cardiotoxic effects should be considered in cases where cardiac concentrations of sunitinib could be increased.

1. We investigated the roles of nitric oxide (NO) and endothelin-1 (ET-1) in organ dysfunction in diabetic mice with normal genotype (wild-type, WT) or myocyte-specific overexpression of endothelial NO synthase (eNOS) (transgenic, TG) after chronic oral treatment with the endothelin-A (ETA) receptor antagonist atrasentan. 2. Mice were rendered diabetic by injection of 200 mg kg-1 streptozotocin (STZ). Experimental groups were: untreated WT diabetic (n=9), untreated TG diabetic (n=9), atrasentan-treated WT diabetic (n=9), atrasentan-treated TG diabetic (n=8) and the four corresponding nondiabetic groups (n=5). Atrasentan was administered orally via drinking water at 3 mg kg-1 per day over 28 days. All diabetic mice developed similar hyperglycaemia (27-30 mmol l-1). 3. Atrasentan treatment significantly improved left ventricular systolic and diastolic function in response to exogenous norepinephrine, but there were no differences between genotypes. 4. Atrasentan antagonized the diabetic impairments in endothelium-dependent coronary relaxation and thromboxane-receptor mediated aortic constriction. Further, it improved cardiac and renal oxidant status as evident from reduced tissue malondialdehyde levels. 5. Atrasentan reduced diabetic urine flow, proteinuria and plasma creatinine levels, but creatinine clearance was not significantly altered. 6. These results suggest that in experimental type 1 diabetes, blocking ETA receptors ameliorates myocardial, coronary and renal function and improves tissue oxidant status, whereas raising myocardial NO levels has neither beneficial nor deleterious effects on diabetic cardiomyopathy in this transgenic model.

Recent evidence suggests a key role of transient receptor potential (TRP) channels in cardiac pathophysiology with TRPC3 as one potential key player in cardiac remodeling. TRPC3 is typically up-regulated by hypertrophic stimuli and may be involved in distorted Ca2+ signaling that drives pathological remodeling. As TRPC proteins generate non-selective cation conductances, we hypothesized that these channels may not only govern Ca2+-mediated gene expression but exert in addition a severe impact on basic electrical properties and excitability of the myocardium. Utilizing the patch clamp technique we characterized membrane currents and electrical properties of cardiomyocytes in response to enhanced TRPC3 expression in the murine HL-1 model. Stimulation of TRPC3-overexpressing HL-1 cells with endothelin-1 [100 nM] (ET-1) as a Gqi-PLC-activator gave rise to a conductance with features distinctly different from the properties described for TRPC3 conductance in expression systems. In HL-1 cells, the TRPC3 over-expression-induced conductance in physiological solutions reversed at about −50 mV, displayed profound outward rectification and was suppressed by the TRPC3-inhibitor Pyr3 [10 µM] (ethyl-1-(4-(2,3,3-trichloroacrylamide)phenyl)-5-(trifluoromethyl)-1H-pyrazole-4-carboxylate). As a result of this conductance, action potential duration was effectively shortened by ET-1 in HL-1 myocytes over-expressing the TRPC3, while this effect was minute in wild-type myocytes. Moreover, TRPC3 over-expression enabled significant depolarizing effects of ET-1 along with action potential shortening and reduction of refractory period. Our results suggests that increased expression of TRPC3 in cardiomyocytes may significantly contribute to electrical remodeling in hypertrophic hearts, generating changes in action potential morphology that are likely to promote arrhythmias.

Recent evidence suggests involvement of transient receptor potential (TRP)-related cation channels in cardiac physiology and pathophysiology, with TRPC3 as one potential key player in cardiac hypertrophy. It has been suggested that TRPC3 is upregulated in hypertrophy development and contributes to Ca2+ signals that govern pathological remodelling. As TRPC proteins form nonselective cation channels, we hypothesized that these molecules determine not only Ca2+-mediated gene expression but also excitability and basic electrical properties of the myocardium in pathological states. We utilized the patch clamp technique to characterize the electrical remodelling associated with enhanced TRPC3 expression in cardiac myocytes. The murine HL-1 atrial cell model, which was found to express both TRPC3 and TRPC6 at low levels along with typical signalling partners of TRPC proteins such as Na+/Ca2+-exchanger-1 and CaV1.2, was modified to overexpress a functional YFP-TRPC3 fusion protein. TRPC3 over-expressing cells displayed substantially altered electrical properties even in the absence of TRPC-activating stimuli, as evident from a significant shortening of basal action potential duration and impaired frequency-dependent adaptation of action potential shape. Action potential-clamp experiments as well as mathematical modelling attempts were performed to delineate the consequences of TRPC3 overexpression and to analyze the mechanisms of TRPC3-mediated electrical remodelling in the heart. Our results suggest a pivotal role of TRPC channels in cardiac electrical remodelling associated with maladaptive hypertrophy.