Increase in the extracellular glutamate level during seizures and electrical stimulation determined using a high temporal resolution technique

BMC Neuroscience

Silvia López-Pérez¹  Jorge Ortega-Ibarra¹  Alberto Morales-Villagrán²  Kenia Pardo-Peña¹  Laura Medina-Ceja¹ 

[1] Laboratory of Neurophysiology and Neurochemistry, Department of Cellular and Molecular Biology, CUCBA, University of Guadalajara, Jalisco, Mexico;Laboratorio de Neurofisiología y Neuroquímica, Departamento de Biología Celular y Molecular, Centro Universitario de Ciencias Biológicas y Agropecuarias, Universidad de Guadalajara, Camino Ing. R. Padilla Sánchez 2100, Las Agujas, Nextipac, Zapopan, CP 45110, Jalisco, Mexico

关键词: Seizures;    Novel device;    Microdialysis;    Glutamate;    Bicuculline;    4-Aminopyridine;   

Others  :  1170565
DOI  :  10.1186/s12868-015-0147-5

 received in 2014-09-11, accepted in 2015-02-20,  发布年份 2015

【 摘 要 】

Background

Glutamate has been measured using different methods to determine its role under normal and pathological conditions. Although microdialysis coupled with HPLC is the preferred method to study glutamate, this technique exhibits poor temporal resolution and is time consuming. The concentration of glutamate in dialysis samples can be measured via glutamate oxidase using the Amplex Red method.

Methods

A new device has been designed and constructed to rapidly deposit dialysis samples onto a polycarbonate plate at Cartesian coordinates (every five seconds). The samples were added to an enzymatic reaction that generates hydrogen peroxide from glutamate, which was quantified using fluorescence detection. Fluorescence emission was induced by laser excitation, stimulating each spot automatically, in addition to controlling the humidity, temperature and incubation time of the enzymatic reaction.

Results

The measurement of standard glutamate concentrations was linear and could be performed in dialysis samples. This approach was used to determine the effect of the convulsant drugs bicuculline and 4-aminopyridine on the extracellular glutamate concentration. Seizure activity was associated with a considerable increase in glutamate that correlated with altered EEG patterns for both drugs.

Conclusions

These results indicate that this method is able to read samples with high temporal resolution, and it is easy to use compared with classical methods such as high-performance liquid chromatography, with the advantage that a large number of samples can be measured in a single experimental series. This method provides an alternative approach to determine the concentrations of neurotransmitters or other compounds that generate hydrogen peroxide as a reaction product.

【 授权许可】

   
2015 Medina-Ceja et al.; licensee BioMed Central.

【 图 表 】

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