Highly sensitive on-line measurement of glutamate released from rat brain neurons in vivo and in vitro

• Published in: Japanese Journal of Pharmacology Vol. 71; no. suppl.1; p. 189
• Main Authors: Torimitsu, Keiichi, Niwa, Osamu, Kawana, Akio, Osborne, Pefer, Yamamoto, Katsunobu
• Format: Journal Article
• Language: English; Japanese
• Published: The Japanese Pharmacological Society 1996
• ISSN: 0021-5198
• DOI: 10.1016/S0021-5198(19)36996-3

A novel measurement system of glutamate has been developed. The system includes glutamate oxidase immobilized enzyme column which could generate hydrogen peroxide and leads to the electrochemical detectors of glassy carbon. The detectors are coated with Os-polyvinyl pyridine mediator containing horse radish peroxidase. On-line measurement had been carried sampling the medium by using micro-dialysis tube of CMA (1 mm in length for in vitro, 3 mm for in vivo). Adult and E18 embryonic rat cortex were used for the measurement in viva and in vitro. Dissociated cells were prepared by papain digestion and were cultured for 2 weeks in the DMEM based medium containing the heat-inactivated FBS and the heat-inactivated horse serum. Experiments have been carried in serum-free conditions either in viva or in vitro to detect glutamate release from the nerve cells and tissues. About 10^-5 - 10^-6 M glutamate release has been detected after addition of drugs, such as potassium stimulation or anesthetic drugs. Using this system, glutamate release could be detected in the range of 10^-8 to 10^-4 M in the linear response. As this system allows us continuous measurement for more than 8 hrs and the sensors were stable for more than a week in the conditions of 4℃, the system would be extremely useful in the pharmacological study either in viva or in vitro conditions.