Journal of Neuroscience Methods xxx (2004) xxx–xxx High temporal resolution for in vivo monitoring of neurotransmitters in awake epileptic rats using brain microdialysis and capillary electrophoresis with laser-induced fluorescence detection Sandrine Parrot a , Val´ erie Sauvinet a , V´ eronique Riban b , Antoine Depaulis b , Bernard Renaud a , Luc Denoroy a,∗ a Laboratoire de Neuropharmacologie et Neurochimie, INSERM U512, Institut F´ ed´ eratif des Neurosciences de Lyon (IFR 19), Universit´ e Claude Bernard—Lyon I, 8 Avenue Rockefeller, 69373 Lyon Cedex 08, France b Jeune Equipe MRNT “Contrˆ ole des r´ eseaux synchrones ´ epileptiques”, Universit´ e Joseph Fourier—Grenoble, 2280 Rue de la Piscine, 38400 St. Martin d’H` eres, France Received 30 October 2003; accepted 22 March 2004 Abstract A method for high temporal resolution monitoring of five neurotransmitters, dopamine (DA), noradrenaline (NA), gamma-aminobutyric acid (GABA), glutamate (Glu), l-aspartate (L-Asp), in freely-moving rats using microdialysis and capillary electrophoresis with laser- induced fluorescence detection (CE-LIFD) was developed. An on-line device, including microdialysis and derivatization with naphthalene- 2,3-dicarboxaldehyde, mixes the dialysate with derivatization reagents directly in the collection tube, i.e. with no reactor. Thereafter, collected derivatized samples are analyzed off-line with an automated CE system coupled to a LIFD using a 442 nm excitation. The sampling time was limited by the minimal volume required for the analysis by the automated CE system used: neurotransmitters could be determined in 667 nl dialysates (940 nl after derivatization), i.e. in samples collected every 20 s with a flow rate of 2 l/min. The detection limits at the dialysis probe were 3 × 10 -9 ,1 × 10 -9 , 1.9 × 10 -8 , 4.2 × 10 -7 , 2.1 × 10 -7 mol/l for DA, NA, GABA, Glu and L-Asp, respectively. The protocol was validated using in vitro/in vivo tests and the performances—repeatability, linearity, characteristics of the probes—were determined. Finally, the high temporal resolution allowed the simultaneous monitoring of these neurotransmitters in rats with genetic absence epilepsy and revealed, for the first time, increases in GABA concentrations concomitantly with the seizures, detected when our new microdialysis method was combined to electroencephalographic recordings. © 2004 Elsevier B.V. All rights reserved. Keywords: Amino acids; Catecholamines; High sampling rate microdialysis; Awake rats 1. Introduction Studying rapid neurochemical events requires techniques enabling measurements with a sub-minute time scale. So far, biosensors or in vivo electrochemistry have been used to monitor such rapid changes in extracellular levels of neu- rotransmitters (for review, Kehr, 1999). However, with such a method, only one compound can be monitored at once, ∗ Corresponding author. Tel.: +33 478 777 558; fax: +33 478 777 209. E-mail address: denoroy@sante.univ-lyon1.fr (L. Denoroy). thus, limiting the use of biosensors when one wants to study interactions between several neurotransmitters. In contrast, microdialysis allows the simultaneous monitoring of sev- eral molecules since it is coupled to a separation-based an- alytical technique. However, microdialysis has been previ- ously limited in many cases by its poor temporal resolution when combined with the conventional high performance liq- uid chromatography which requires relatively large volumes of samples and therefore a 10–30 min sampling duration. In order to analyze sub-microliter dialysates obtained with a sub-minute sampling, our group and others have successfully 0165-0270/$ – see front matter © 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.jneumeth.2004.03.025