NEURONAL AND GLIAL HANDLING OF GLUTAMATE AND GLUTAMINE DURING HYPOOSMOTIC STRESS - A BIOCHEMICAL AND QUANTITATIVE IMMUNOCYTOCHEMICAL ANALYSIS USING THE RAT CEREBELLUM AS A MODEL

Biochemical and immunocytochemical analyses were performed to resolve how glutamate and glutamine are handled in rat cerebellar cortex in ac ute hypoosmotic stress. Rats were subjected to a 15-20% reduction in p lasma osmolality by intraperitoneal injection of distilled water and t hen perfusion fixed after 4 or 8 h survival. Some rats in the latter g roup had their plasma isoosmolality restored by injections of hyperton ic saline 4 h prior to perfusion. Water loading caused a pronounced in crease in the tissue level of glutamine and an equimolar decrease in t he level of glutamate after 4 h survival. The increase in glutamine wa s transient, as judged by analyses at 8 h survival. Light microscopic immunocytochemistry revealed a pronounced enhancement of the glutamine immunolabelling of glial cells (Golgi epithelial cells and astrocytes ), including their perivascular end feet, and quantitative immunogold analyses at the electron microscopic level showed that this enhancemen t reflected a 50% increase in the intracellular concentration of fixed glutamine. Since water loading was associated with glial swelling thi s change corresponded to a several-fold increase in the glial content of glutamine. There was a modest reduction in the overall staining int ensity for glutamate. The biochemical and immunocytochemical changes w ere reversed upon restoration of plasma osmolality by hypertonic salin e. These findings suggest that hypoosmotic stress causes an increased conversion of glutamate to glutamine in glial cells and that the latte r amino acid is subsequently lost from the tissue. The flux of glutama te carbon skeletons through the glutamine synthetase pathway in glia, prior to an efflux to the systemic circulation, may explain how glutam ate, an excitatory transmitter and potential toxin, can be used as an organic osmolyte in brain tissue.