Imaging of L-glutamate fluxes in mouse brain slices based on an enzyme-based membrane combined with a difference-image analysis

Ayumi Hirano, Nobukatsu Moridera, Mai Akashi, Minoru Saito, Masao Sugawarat

Research output: Contribution to journalArticlepeer-review

18 Citations (Scopus)

Abstract

A time-resolved imaging method for visualizing L-glutamate release in mammalian brain slices is proposed by using an enzyme membrane combined with a difference-image analysis. The enzyme membrane is composed of L-glutamate oxidase and horseradish peroxidase incorporated into a bovine serum albumin matrix. L-Glutamate triggers an enzyme-coupling reaction to convert a redox substrate (DA-64) to Bindschedler's Green, which gives a green color signal. The difference-image analysis is based on calculating slopes of a signal versus time (t) plot in the time range from (t - 40 s) to (t + 40 s) for visualizing L-glutamate release in terms of its flux (in mol Min-1 cm-2). The method was applied to a time-resolved imaging of hippocampal distribution of ischemia-induced L-glutamate release in mouse brain slices. The image of L-glutamate distribution showed that the level and time courses of L-glutamate fluxes were neuronal region-dependent. The maximum flux of L-glutamate at CA1 was observed at 7.7 min after ischemia. The flux at 7.7 min increased in the order of CA1 ≈ CA3 > DG. The time course of the L-glutamate flux in the CA1 region was biphasic and that in the DG region was modestly biphasic. In the CA3 region, such biphasic release of L-glutamate was not seen. The ischemia-induced L-glutamate flux was accelerated when Mg2+ was omitted from an extracellular solution. The present enzyme membrane-based approach provides a useful method for visualizing distribution of L-glutamate release in the brain slices during ischemia.

Original languageEnglish
Pages (from-to)3775-3783
Number of pages9
JournalAnalytical Chemistry
Volume75
Issue number15
DOIs
Publication statusPublished - 2003 Aug 1
Externally publishedYes

ASJC Scopus subject areas

  • Analytical Chemistry

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