An improved method for the detection of changes in brain extracellular glutamate levels.
We developed a method for in vivo real-time monitoring of the concentration of extracellular glutamate ([Glu]e) in the brain under anoxic conditions. A dialysis electrode (Sycopel Int., UK) was employed as a sensing device to measure the concentration of glutamate by enzyme amperometry, and an electron mediator, ferrocene, was introduced into the electrode together with glutamate oxidase. The ferrocene was covalently conjugated with a high molecular weight molecule, bovine serum albumin, to avoid outward diffusion through the dialysis membrane. With this set-up, the amperometric response was independent of the pO2 around the electrode in vitro up to 400 microM glutamate. Using this method, we investigated the dynamics of [Glu]e in the rat striatum during anoxia. [Glu]e increased rapidly at 102+/-5.4s (n = 6) after the start of nitrogen inhalation. The increase continued for about 30 s, and then [Glu]e decreased. The peak value of delta[Glu]e was 141+/-37 micro M. [Glu]e subsequently underwent another gradual increase, reaching 213+/-69 microM at 15 min after the start of nitrogen inhalation. This distinct biphasic profile was reproducible. We conclude that this method is very useful for monitoring [Glu]e in the brain under low pO2 conditions.
Biomedical Research Team, Frontier Technology Research Institute, Tokyo Gas Co. Ltd., Yokohama, Japan.
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T
Kohno
S
S
Asai
Y
Y
Iribe
I
I
Hosoi
K
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Shibata
K
K
Ishikawa
D000818:Animals; D015374:Biosensing Techniques; D001921:Brain / Q000378:metabolism*; D002417:Cattle; D004058:Diffusion; D004567:Electrodes, Implanted; D004569:Electroencephalography; D005110:Extracellular Space / Q000737:chemistry / Q000378:metabolism*; D005296:Ferrous Compounds / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D000860:Hypoxia / Q000378:metabolism; D008297:Male; D000075163:Metallocenes; D017551:Microdialysis / Q000379:methods; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D012710:Serum Albumin, Bovine / Q000378:metabolism
D005296:Ferrous Compounds; D000075163:Metallocenes; D012710:Serum Albumin, Bovine; D018698:Glutamic Acid; C004998:ferrocene