King, Nicola; McGivan, John D; Griffiths, Elinor J; Halestrap, Andrew P; Suleiman, M-Saadeh

Author(s)

King, Nicola

McGivan, John D

Griffiths, Elinor J

Halestrap, Andrew P

Suleiman, M-Saadeh

Publication Date

2003

Abstract

Glutamate loading has been shown to protect single isolated perfused cardiomyocytes against metabolic inhibition and wash-off. The mechanism underpinning this protection is unknown. This study aimed to investigate whether reactive oxygen species (ROS) are generated by single isolated perfused cardiomyocytes and whether the protective effect of glutamate loading on cell metabolism is linked to ROS. Single rat cardiomyocytes were isolated with or without glutamate to stimulate glutamate loading. ROS production was measured using 5-(and-6)- chloromethyl-2', 7'-dichlorodihydrofluorescein diacetate in various stressful conditions including metabolic inhibition and wash-off with/without antimycin A or myxothiazol; simulated ischaemia (without cyanide) and glucose reintroduction; and H₂O₂ perfusion. Reduced glutathione (GSH) levels were measured in control and glutamate-loaded cells with/without exposure to H₂O₂. Finally, the effect of glutamate on glutathione reductase and glutathione peroxidase activity was measured. In every stressful condition studied, ROS production was significantly lower in glutamate-loaded cells compared to controls. This occurred regardless of whether ROS were produced intracellularly (e.g. from the respiratory chain inhibited with antimycin A) or via the extracellular precursor H₂O₂. Glutamate-loaded cells also maintained their morphological integrity at higher H₂O₂ concentrations than control cells. Furthermore, during H₂O₂ exposure GSH levels decreased in glutamate-loaded cells but stayed constant in control cells. Glutamate stimulated the activity of glutathione peroxidase in a concentration dependent fashion. These results provide new evidence to show that the cardioprotective effect of glutamate loading may be mediated through an enhanced ability to destroy ROS in the cell.

Citation

Journal of Molecular and Cellular Cardiology, v.35, p. 975-984

ISSN

1095-8584

0022-2828

Link

https://hdl.handle.net/1959.11/5142

Publisher

Academic Press

Title

Glutamate loading protects freshly isolated and perfused adult cardiomyocytes against intracellular ROS generation

Type of document

Journal Article

Entity Type

Publication

Author(s)	King, Nicola McGivan, John D Griffiths, Elinor J Halestrap, Andrew P Suleiman, M-Saadeh
Publication Date	2003
Abstract	Glutamate loading has been shown to protect single isolated perfused cardiomyocytes against metabolic inhibition and wash-off. The mechanism underpinning this protection is unknown. This study aimed to investigate whether reactive oxygen species (ROS) are generated by single isolated perfused cardiomyocytes and whether the protective effect of glutamate loading on cell metabolism is linked to ROS. Single rat cardiomyocytes were isolated with or without glutamate to stimulate glutamate loading. ROS production was measured using 5-(and-6)- chloromethyl-2', 7'-dichlorodihydrofluorescein diacetate in various stressful conditions including metabolic inhibition and wash-off with/without antimycin A or myxothiazol; simulated ischaemia (without cyanide) and glucose reintroduction; and H₂O₂ perfusion. Reduced glutathione (GSH) levels were measured in control and glutamate-loaded cells with/without exposure to H₂O₂. Finally, the effect of glutamate on glutathione reductase and glutathione peroxidase activity was measured. In every stressful condition studied, ROS production was significantly lower in glutamate-loaded cells compared to controls. This occurred regardless of whether ROS were produced intracellularly (e.g. from the respiratory chain inhibited with antimycin A) or via the extracellular precursor H₂O₂. Glutamate-loaded cells also maintained their morphological integrity at higher H₂O₂ concentrations than control cells. Furthermore, during H₂O₂ exposure GSH levels decreased in glutamate-loaded cells but stayed constant in control cells. Glutamate stimulated the activity of glutathione peroxidase in a concentration dependent fashion. These results provide new evidence to show that the cardioprotective effect of glutamate loading may be mediated through an enhanced ability to destroy ROS in the cell.
Citation	Journal of Molecular and Cellular Cardiology, v.35, p. 975-984
ISSN	1095-8584 0022-2828
Link	https://hdl.handle.net/1959.11/5142
Publisher	Academic Press
Title	Glutamate loading protects freshly isolated and perfused adult cardiomyocytes against intracellular ROS generation
Type of document	Journal Article
Entity Type	Publication

Glutamate loading protects freshly isolated and perfused adult cardiomyocytes against intracellular ROS generation

Files: