Real-time detection of L-glutamate released from C6 glioma cells using a modified enzyme-luminescence method.
There is an increasing interest in new strategies to detect neurotransmitters released from nerve cells in real time for brain science, drug assessment, and so on. Previously we reported real-time monitoring of dopamine release from nerve model cells by enzyme-catalyzed luminescence measurement with tyramine oxidase and peroxidase. In the present study, the system was modified with glutamate oxidase instead of tyramine oxidase to detect L-glutamate sensitively ( approximately 10 nM) and rapidly with high temporal resolution (<1 s). We applied this modified method successfully to perform real-time monitoring of L-glutamate release from brain model cell (C6 glioma cell) using a luminescence plate reader upon stimulation with high concentration of KCl (>10 mM) or 5-hydroxytryptamine (>1 microM). The measurement solution was not toxic and therefore the L-glutamate release from the cell was measured by the second stimulation after exchanging the measurement solution. We conclude that the developed monitoring system is suitable for real-time detection of dynamic L-glutamate release from nerve cells in vitro and will be suitable for application in assessment of drugs acting on the nervous system.
Zakir Hossain
S M
SM
Department of Advanced Nano and Biosciences, Graduate School of Innovative Life Science, University of Toyama, 3190 Gofuku, Toyama 930-8555. Japan. zhossain@ucalgary.ca
Shinohara
Hiroaki
H
Wang
Feifei
F
Kitano
Hiromi
H
D000818:Animals; D015374:Biosensing Techniques; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D005910:Glioma / Q000378:metabolism*; D018698:Glutamic Acid / Q000032:analysis* / Q000378:metabolism; D049449:Luminescence; D010088:Oxidoreductases / Q000378:metabolism*; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D012680:Sensitivity and Specificity; D012701:Serotonin / Q000494:pharmacology; D018490:Serotonin Agents / Q000494:pharmacology; D013268:Stimulation, Chemical; D013997:Time Factors
D018490:Serotonin Agents; D012701:Serotonin; D018698:Glutamic Acid; D011189:Potassium Chloride; D010088:Oxidoreductases