Extracellular Glutamate During Focal Cerebral Ischaemia in Rats: Time Course and Calcium Dependency

Abstract

The time course of changes in extracellular glutamic acid levels and their Ca2+ dependency were studied in the rat striatum during focal cerebral ischaemia, using microdialysis. Ischaemia‐induced changes were compared with those produced by high K+‐evoked local depolarization. To optimize time resolution, glutamate was analysed continuously as the dialysate emerged from the microdialysis probe by either enzyme fluorimetry or biosensor. The Ca2+ dependency of glutamate changes was examined by perfusing the probe with Ca2+‐free medium. With normal artificial CSF, ischaemia produced a biphasic increase in extracellular glutamate, which started from the onset of ischaemia. During the first phase lasting ∼10 min, dialysate glutamate level increased from 5.8 ± 0.9 µM· min−1 to 35.8 ± 6.2 µM where it stabilized for ∼3 min. During the second phase dialysate glutamate increased progressively to its maximum (82 ± 8 µM), reached after 55 min of ischaemia, where it remained for as long as it was recorded (3 h). The overall changes in extracellular glutamate were similar when Ca2+ was omitted from the perfusion medium, except that the first phase was no longer detectable and, early in ischaemia, extracellular glutamate increased at a significantly slower rate than in the control group (2.2 ± 1 µM· min−1; p < 0.05). On the basis of these data, we propose that most of the glutamate released in the extracellular space in severe ischaemia is of metabolic origin, probably originating from both neurons and glia, and caused by altered glutamate uptake mechanisms. Comparison with high K+‐induced glutamate release did not suggest that glutamate “exocytosis,” early after middle cerebral artery occlusion, was markedly limited by deficient ATP levels.

Cellular mechanisms underlying acquired epilepsy: The calcium hypothesis of the induction and maintainance of epilepsy

“…However, these changes are much less severe in the ischemic penumbra (Lipton, 1999). Extracellular glutamate concentrations in penumbral tissue are elevated to a lesser extent than in the core (Wang et al, 2001), estimated at 30-50 µM in different studies (Shimada et al, 1990;Wahl et al, 1994;Morimoto et al, 1996).…”

“…In the ischemic core, extracellular glutamate has been estimated to increase from a physiological level of 5 µM to values ranging from 100 µM to 10 mM (Graham et al, 1990;Wahl et al, 1994;Rusakov & Kullmann, 1998), which leads to excessive glutamate receptor activation. Excessive activation of glutamate receptors further enhances the anoxic depolarization and Ca 2+ influx.…”

“…Under physiological conditions, extracellular glutamate has been measured in the range of 1-5 µM (Wahl et al, 1994). Although transporters exist to move glutamate into the brain across the blood-brain barrier, the vast majority of glutamate is synthesized de novo from glucose, glutamine, or aspartate (Laterra et al, 1999).…”

Glutamate release by an Na+ load and oxidative stress in nerve terminals: relevance to ischemia/reperfusion

“…Evidence for both vesicular (Drejer et al 1985;Hershkowitz et al 1993) and non-vesicular (Szatkowski et al 1990;Pocock and Nicholls 1998) glutamate release during ischemia have been provided. It appears that in the early phase of ischemia, glutamate is released by a Ca 2+ -dependent vesicular mechanism (Katayama et al 1991;Wahl et al 1994;Pocock and Nicholls 1998;Asai et al 2000), which is followed by a Ca 2+ -independent non-vesicular release (Sanchez-Prieto and Gonzalez 1988; Ikeda et al 1989;Szatkowski et al 1990;Kawakami et al 2001). Real time glutamate measurement in an in vitro model of ischemia showed that glutamate uptake is inhibited within a few minutes of the onset of ischemia (Jabaudou et al 2000).…”

A simple method for measuring intracellular activities of glutamine synthetase and glutaminase in glial cells

“…For example, in stroke and ischemia the GS activity is strongly reduced due to the drop in intracellular ATP levels but also via oxidative damage and proteolytic degradation (23,28). This may indirectly contribute to the dramatic increases in extracellular glutamate levels seen in the ischemic brain (4,13,44). At least some forms of human epilepsy and animal epilepsy models are associated with loss of the GS protein in hippocampus that may contribute to increased extracellular glutamate levels and hyperexcitability (12,15).…”