Myocardial remodelling induced by repeated low doses of isoproterenol.(Report)

Introduction

Left ventricular hypertrophy is an independent adaptive response to hemodynamic overload (Wang et al. 2003; Diwan and Dorn 2007) and a risk factor for sudden cardiac death. Experimental myocardial hypertrophy without systemic hypertension can be induced by pressure-volume overload as a result of isoproterenol administration (Rakusan et al. 1965; Meszaros and Levai 1990). Isoproterenol (or isoprenaline) is a [beta]-agonist with strong inotropic and chronotropic actions mediated by preceptors. Its administration results in greater oxygen demand by the working myocardium. By means of peripheral vasodilatation, it reduces systemic blood pressure via the activation of [[beta].sub.2]-receptors in vascular smooth muscle (Kahn et al. 1969).

Cardiac hypertrophy is associated with abnormal electrical activity. Meszaros and Levai (1990), Meszaros and Pasztor (1995), and Meszaros et al. (2001) have demonstrated that the membranes of cardiomyocytes that hypertrophied after repeated isoproterenol administration (Isohypertrophic) showed characteristic changes in membrane capacitance and action potentials. Shipsey et al. (1997) observed action potential prolongation in Iso-hypertrophic cells associated with changes in T-wave morphology that may have implications for arrhythmogenesis. Meszaros and Pasztor (1995) observed a prolongation of the ST interval in Iso-pretreated rats. Prolonged QRS and QT interval durations can be considered markers of myocardial hypertrophy (Yan and Antzelevitch 1998; Gima and Rudy 2002).

Cardiac hypertrophy induced by the repeated administration of low doses of isoproterenol (0.3-5 mg/kg) (Lijnen et al. 2000; Meszaros and Levai 1990) can, in principle, occur by cell proliferation and (or) the enlargement of individual cells. Furthermore, hemodynamic overload may be accompanied by necrotic changes in the heart leading to mitochondrial swelling, the disruption of cristae and outer mitochondrial membranes (Meszaros and Levai 1990), and the proliferation of connective tissue with a consequent accumulation of collagen (Meszaros and Levai 1990; Meszaros and Pasztor 1995; Nagano et al. 1992; Zimmer 1997; Suzuki et al. 1998; Lijnen et al. 2000; Ocaranza et al. 2002; Goldspink et al. 2004; Zhang et al. 2007).

Until now, the study of cell ultrastructure in isoproterenol-induced hypertrophy has concentrated on profound changes in severely damaged cells, and as a result, data on ultrastructural changes during the early stages of developing hypertrophy in the myocardium are lacking. The present investigation was aimed at characterizing the changes in the cytoarchitecture of the cardiomyocytes in the surviving myocardium that have adapted to the developing hemodynamic overload induced by isoproterenol and relating this cytoarchitecture to changes in the electrical activity of the hypertrophic heart. Additionally, we have utilized our recently developed approach (Kralova et al. 2008) of applying the Sokolow-Lyon voltage index, previously used in clinical trials for the detection of human cardiac hypertrophy (Norman and Levy 1995; Baillard et al. 2000; Okin et al. 2003; Oikarinen et al. 2004), to catecholamine-induced hypertrophy in rats. Materials and methods

Animal treatment

Two- to 3-month-old male Wistar rats obtained from the Dobra Voda breeding station, Slovak Republic, were used for the experiments. All animals were allowed to acclimatize to the housing conditions with free access to food and tap water for at least 7 days, followed by 3-5 days of daily handling before the beginning of the study. Animals from the isoproterenol group (Iso, n = 7) were treated with a subcutaneous (s.c.) injection of isoproterenol (Fluka, Steinheim, Germany) dissolved in the vehicle (0.9% NaCl containing 0.05% ascorbic acid to prevent isoproterenol oxidation) at a dose of 5 mg/kg once daily for 7 days to evoke myocardial hypertrophy (Meszaros and Levai 1990; Meszaros and Pasztor 1995; Meszaros et al. 2001; Krenek et al. 2006). The dose of isoproterenol was adjusted daily according to the actual body weight. Animals in the control group (n = 7) were treated daily with an s.c. injection of the vehicle.

The investigation conformed to the Guide for the Care and Use of Laboratory Animals published in the Collection of Laws of the Slovak Republic (Z.z. SR) No. 289/2003 and was approved by the Ethics Committee of the Faculty of Pharmacy, Comenius University, and by the State Veterinary and Food Administration, Slovak Republic (protocol No. Ro-2222/06-221).

Blood pressure and ECG study

Blood pressure was measured in conscious animals before administration of isoproterenol and 24 h after the 7 days of premedication before the heart isolation. Thereafter, the animals were anesthetized by thiopental sodium (45 mg/kg, 5% solution intraperitoneally, Biochemie Gmbh, Kundl, Austria) to perform electrocardiography in animals as well as in isolated hearts. At the end of experiments the hearts were placed into ice-cold Krebs-Henseleit solution for several minutes to stop their beating. Hearts were blotted dry and weighed, and left ventricle (LV) dimensions were measured using a slide rule.

Two additional groups of animals (control n = 5, Iso n = 5) treated in parallel were used for morphological and histological assessment.

Blood pressure measurement

The mean arterial blood pressure was measured by the tail-cuff method in conscious animals prewarmed to 39[degrees]C in …