Neuroscience Letters, 141 (1992)61-64 61 © 1992 Elsevier Scientific Publishers Ireland Ltd. All rights reserved 0304-3940/92/$ 05.00 NSL 08727 Calcium dependent release of T-aminobutyric acid (GABA) from human cerebral cortex T.S. Haugstad, E. Hegstad and I.A. Langmoen Institute for Surgical Research and Department of Neurosurgery, Rikshospitalet, The National Hospital, University of Oslo, Oslo (Norway) (Received 13 January 1992; Revised version received 25 March 1992; Accepted 3 April 1992) Key words: Aminoacid; GABA; Human brain; Release; Neurotransmitter; Brain slice The release of the amino acids GABA, taurine, glycine, glutamine and leucine from human neocortex was investigated in vitro by utilizing brain tissue removed during 8 standard temporal lobectomiesfor epilepsy or tumor. Slices (0.5 mm thick) were cut from each biopsy and randomly placed in three different chambers. After 90 min preincubation, the three sets of slices were incubated for 60 s in wells containing, respectively,(A) regular ACSF (control), (B) ACSF with 50 mM K+ (to depolarize the cell membrane) and (C) ACSF with 50 mM K +, 0 mM Ca2. and 4 mM Mg 2÷ (depolarization during blocked synaptic transmission). The content of amino acids in the wells was determined by high-performanceliquid chroma- tography after pre-column derivatization of the amino acids with o-phthalaldehyde. Membrane depolarization (well B) increased the GABA release to 650% (620 pmol/mg) of control (well A, 95 pmol/mg). Blocking synaptic transmission (well C) reduced the evoked release by 50% (360 pmol/mg). The releaseof glycine, taurine, glutamine and leucine during membranedepolarization was not significantlydifferent from the control values. The data provide evidencefor a Ca2+-dependent release of GABA, supporting a possible role of this amino acid as a neurotransmitter in human neocortex. The human brain contains y-aminobutyric acid (GABA), glycine and taurine. The extracellular concen- trations are reflected in the cerebrospinal fluid (CSF) [9, 15, 16]. GABA has been implicated in the pathophysiol- ogy of epilepsy [17, 25, 33], and certain drugs such as barbiturates and benzodiazepines act by modifying GABA receptor mediated responses [7, 23] (see also refs. 8, 10 for reviews). GABA and glycine are established as inhibitory neu- rotransmitters in the central nervous system of lower mammals [for reviews, see refs. 26, 29-31]. In the present paper we report a caa+-dependent release of GABA from human neocortex. Biopsies were obtained during 8 temporal lobectomies performed due to glioma [34] or epilepsy [12]. Standard surgical procedures were strictly followed. The prepara- tion of the tissue was essentially as described previously [6, 22]. The tissue was cooled in oxygenated artificial cer- ebrospinal fluid (ACSF, see below) at 4°C for 1 min. 0.5-mm-thick slices were cut and transferred to a prein- cubation chamber containing ACSF with the following composition (in mM): NaC1 123, KC1 3.75, KH2PO 4 Correspondence: I.A. Langmoen, Deparment of Neurosurgery, Riks- hospitalet, N-0027 Oslo, Norway 1.25, MgC12 1, CaCI2 2, NaHCO3 26, glucose (dextro- rotated) 9, and was saturated by a gas mixture of 95% 02 and 5% CO2 making pH 7.4. After resting 90 min in this chamber, the slices were transferred to the test chambers. ACSF samples were cleaned and deproteinized by ul- tracentrifugation through a 5 × 103 Da cutoff filter (Nihon Millipore, Yonezawa, Japan), and stored at -70°C until analysis. The content of amino acids was determined by high-performance liquid chromatography (HPLC) (Perkin Elmer, Norwalk, CT, USA). In brief, the amino acids were pre-column derivatized with o- phthalaldehyde and separated on a reversed phase Hy- persil ODS column (3.1 x 200 mm) (Chrompak Interna- tional B.V., Middelburg, The Netherlands) at 42°C. Mobil phases were A: 0.06 M sodium acetate, pH 6.5, and B: acetonitrile/0.1 M sodium acetate/methanol (14/4/ 1). The gradient was non-linear from 6 to 36% of mobile phase B through 45 min. The derivatized amino acids were quantified by a high sensitivity fluorescence detec- tor (excitation wavelength 340 nm, emission wavelength 460 nm). The results were tested statistically by the Wilcoxon two-sample rank sum test and are presented as mean + S.E.M. (standard error of the mean). One set of experiments was performed in an inter- phase chamber, continuously perfused with ACSF at a rate of 40 ml/h and superfused by a gas mixture consist-