Aspartate and glutamate prevents isoproterenol-induced cardiac toxicity by alleviating oxidative stress in rats

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Abstract

The protective effect of aspartate and glutamate in isoproterenol induced myocardial infarction (MI) was investigated in experimental animals. Male albino wistar rats were pretreated with aspartate [100 mg (kg body weight)-1day-1] or glutamate [100 mg (kg body weight)-1day-1] intraperitoneally for a period of 7 days. Following amino acid treatment, MI was induced in rats by subcutaneous injection of isoproterenol [200 mg (kg body weight)-1day-1] for 2 days. After 24 h following the last injection, the animals were sacrificed and the biochemical analysis was carried out. The activities of cardiac marker enzymes (alanine transaminase, aspartate transaminase, lactate dehydrogenase and creatine phosphokinase) were increased significantly (P<0.05) in the serum of MI induced rats as compared to control rats. The levels of glutathione and mitochondrial ATP and the activities of antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase, glutathione transferase and glutathione reductase) were decreased whereas lipid peroxides increased significantly (P<0.05) in the heart of MI induced rats as compared to control rats. However, pretreatment with aspartate or glutamate to MI induced rats significantly (P<0.05) reduced the activities of cardiac marker enzymes and increased the activities of antioxidant enzymes as compared to MI induced rats. Aspartate or glutamate pretreatment also increased the levels of glutathione and mitochondrial ATP while decreased the level of lipid peroxides in the cardiac tissue. The overall effects of aspartate and glutamate in reducing the oxidative stress in MI induced rats are similar. There was no significant difference between the control rats and aspartate or glutamate treated control rats. The present study shows that aspartate and glutamate could reduce oxidative stress in MI induced rats.

Introduction

Despite the rapid advances that have been made in the treatment of coronary artery disease, myocardial infarction (MI) remains the major cause of death in the developed world and a major pathology issue worldwide (Adams, 2002). MI is characterized by necrosis of cardiac muscle induced by decreased supply of blood to a portion of the myocardium. A disparity between oxygen demand and supply leads to ischemic necrosis of heart muscle (DeBono and Boon, 1992). Free radicals are the key mediators associated with ischemia-reperfusion injury (Wattanapitayakul and Bauer, 2001). They may be formed by infiltration of white cells into ischemic myocardium or in the endothelial cell by the action of xanthine oxidase during the period of ischemia (McCord and Roy, 1982).

MI induced by isoproterenol (ISO), a synthetic catecholamine and β-adrenergic agonist, has been reported to show metabolic and morphological changes in the heart of experimental animals similar to those seen in human myocardial ischemia (Wexler, 1978). ISO, upon oxidation increases lipid peroxidation through enhanced free radical formation and causes severe stress in the myocardium resulting in infarct like necrosis of the heart muscles (Chattopadhyay et al., 2003; Rajadurai and Stanely Mainzen Prince, 2007). Experimental induction of myocardial infarction by isoproterenol in animal is a well established model to study the protective role of various cardio-protective agents.

Amino acids play a central role in myocardial energy metabolism, in coordination of mitochondrial and cytosolic biochemical processes (Barrio et al., 1982). It is well documented that exogenous amino acids are converted to intermediates of citrate cycle and by their subsequent anaerobic substrate-level phosphorylation protect ischemic myocardium (Sanborn et al., 1979; Lazar et al., 1980). Among the amino acids, aspartate and glutamate have frequently been considered to be anti-ischemic and shown to protect the heart from functional damages caused by global ischemia and reperfusion (Williams et al., 2001; Pisarenko et al., 2006). Exogenous glutamate and aspartate were reported to provide metabolic support for the ischemic heart (Pisarenko et al., 1995a; Pisarenko et al., 2006). The metabolic pathway is thought to be the aspartate–malate shuttle followed by citrate cycle which increased the production of succinate promoting ATP generation and protect ischemic myocardium (Oldroyd et al., 1990; Pisarenko et al., 1995a). It was speculated that glutamate could offer protection during oxidative stress through the formation of GSH from exogenous glutamate. Moreover, many of the citrate cycle intermediates has been shown to possess antioxidant properties in different in vitro and in vivo systems (Puntel et al., 2007).

A preliminary study reports the effect of aspartate and glutamate on myocardial infarction (Singh et al., 1989). However, to our knowledge no studies on the protective effect of these two amino acids during oxidative stress in MI had been performed. Hence this study was carried out to determine whether aspartate and glutamate could protect the myocardium against oxidative stress in MI induced rats. In the present work, we investigated the effect of aspartate and glutamate on cardiac marker enzymes, lipid peroxidation, glutathione, antioxidant enzymes and mitochondrial ATP in MI induced rats.

Section snippets

Chemicals

All fine chemicals including isoproterenol hydrochloride, sodium-l-aspartate, sodium-l-glutamate and epinephrine were purchased from Sigma (St. Louis, MO, USA). All other reagents and solvents used in this study were of the highest analytical grade, unless otherwise indicated.

Experimental design

Nearly 40 male albino rats of Wistar strain weighing approximately 150–200 g (4–5 months old) were obtained from the Laboratory Animal Maintenance Unit, Tamilnadu Animal Science and Veterinary University, Madavaram, India.

Results

Table 1 shows the effect of aspartate and glutamate on the activities of cardiac marker enzymes in serum of control and experimental rats. The activities of marker enzymes such as ALT, AST, LDH and CPK increased significantly (P<0.05) in the serum of MI induced rats as compared to control rats. The level of TBARS increased whereas the level of GSH decreased significantly (P<0.05) in the heart of MI induced rats when compared with control rats (Fig. 1). Table 2 depicts the activities of

Discussion

Heart can use different substrates for oxidation and normally the specific choice is largely determined by their availability. Glucose is the dominant fuel for heart but it can utilize free fatty acids and lactate as an energy source during fasting and exercise, respectively. During ischemia, anaerobic glycolysis can proceed as long as pyruvate and lactate are disposed of by cell. This process does not occur during severe ischemia and glycolysis ceases (Amark et al., 2006). In this context,

Acknowledgement

R. Sivakumar is grateful to acknowledge the financial assistance awarded by Council of Scientific and Industrial Research (CSIR), New Delhi, India.

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