Early biochemical and histological alterations in rat corticoencephalic cell cultures following metabolic damage and treatment with modulators of mitochondrial ATP-sensitive potassium channels

• Published in: Neurochemistry international Vol. 43; no. 6; pp. 563 - 571
• Main Authors: Reinhardt, R, Manaenko, A, Guenther, A, Franke, H, Dickel, T, Garcia de Arriba, S, Muench, G, Schneider, D, Wagner, A, Illes, P
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 01.11.2003; Elsevier
• Subjects: Animals; ATP-Binding Cassette Transporters; Biological and medical sciences; Cell Hypoxia; Cells, Cultured; Cerebral Cortex - cytology; Cerebral Cortex - drug effects; Cerebral Cortex - metabolism; Corticoencephalic cell cultures; Decanoic Acids - pharmacology; Diazoxide - pharmacology; Female; Fundamental and applied biological sciences. Psychology; HPLC; Hydroxy Acids - pharmacology; Ischemia; KATP Channels; Kinetics; Mitochondria - drug effects; Mitochondria - metabolism; Neurons - cytology; Neurons - drug effects; Neurons - metabolism; Potassium channels; Potassium Channels - drug effects; Potassium Channels - physiology; Potassium Channels, Inwardly Rectifying; Rats; Rats, Wistar; Ribonucleotides - metabolism; Potassium channels; Ischemia; HPLC; Corticoencephalic cell cultures
• ISSN: 0197-0186; 1872-9754
• DOI: 10.1016/S0197-0186(03)00053-6

The present study was aimed at characterizing alterations of the nucleotide content and morphological state of rat corticoencephalic cell cultures subjected to metabolic damage and treatment with modulators of mitochondrial ATP-dependent potassium channels (mitoK ATP). In a first series of experiments, in vitro ischemic changes of the contents of purine and pyrimidine nucleoside diphosphates and triphosphates were measured by high performance liquid chromatography (HPLC) and the corresponding histological alterations were determined by celestine blue/acid fuchsin staining. As an ischemic stimulus, incubation with a glucose-free medium saturated with argon was used. Ischemia decreased the levels of adenosine, guanine and uridine triphosphate (ATP, GTP, UTP) and increased the levels of the respective dinucleotides ADP and UDP, whereas the GDP content was not changed. Both 5-hydroxydecanoate (5-HD) and diazoxide failed to alter the contents of nucleoside diphosphates and triphosphates, when applied under normoxic conditions. 5-HD (30 μM) prevented the ischemia-induced changes of nucleotide and nucleoside levels. Diazoxide (300 μM), either alone or in combination with 5-hydroxydecanoate (30 μM) was ineffective. Pyruvate (5 mM) partially reversed the effects of ischemia or ischemia plus 2-deoxyglucose (20 mM) in the incubation medium. Diazoxide (300 μM) and 5-HD (30 μM) had no effect in the presence of pyruvate (5 mM) and 2-deoxyglucose (20 mM). Staining the cells with celestine blue/acid fuchsin in order to classify them as intact, reversibly or profoundly injured, revealed a protective effect of 5-HD. When compared with 5-HD, diazoxide, pyruvate and 2-deoxyglucose had similar but less pronounced effects. In conclusion, these results suggest a protective role of 5-hydroxydecanoate on early corticoencephalic nucleotide and cell viability alterations during ischemia.