Measurements of Intracellular Calcium Transient and Contractile Function in Isolated Cardiomyocytes Stress values were determined after stabilization of HR. The cardiac variables were recorded under baseline conditions and under β-adrenergic stimulation using isoproterenol (10 μg kg −1 min −1, continuous intravenous administration although the tail vein) with or without pretreatment with MK571 (intravenous administration of 30 mg/kg 6 min before through the tail vein) 7, 15 or the same volume of normal saline as a control. From the arterial blood pressure tracings, the HR and the maximum positive values of first derivative of arterial pressure (+dP/dt) were quantified. After stabilization, the arterial systolic, mean arterial pressure and diastolic pressures were recorded. Pressure transducer catheter (size 2F, Millar Micro-tip catheter transducer, model SPR-407 Millar Instruments, Inc., Houston, TX), was introduced into the right carotid artery and connected to a pressure transducer (Gould Electronic, Cleveland, OH). In vivo, at least 2 days after the echocardiographic assessment, the rats were anesthetized using pentobarbital (50 mg/kg intraperitonally) to measure arterial blood pressure. Then, the left ventricle was carefully dissected and frozen in liquid nitrogen for Western blotting experiments. The heart was removed during anesthesia after the measurements of arterial blood pressure and weighed, except for the hearts used for the isolation of cardiomyocytes, in order to shorten the time of ischemia. Animals were purchased from Janvier (Le Genest St Isle, France) and cared according to the Guiding Principles in the Care and Use of Animals in a labeled housing place (B-75-13-08) with food and water ad libitum. Wistar male rats have been studied in two groups: senescent rats (24 months old) and young adult rats (3 months old).
The project has been submitted to the relevant Animal Care Committee through the French Ministry of High Education and Research (Comité Régional d’Ethique en Expérimentation Animale Paris-comité 3, Paris, France). Experiments were conducted in an authorized laboratory under supervision of an authorized researcher (J.
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