Properties of taurine release in glucose-free media in hippocampal slices from developing and adult mice

Research Article Properties of Taurine Release in Glucose-Free Media in Hippocampal Slices from Developing and Adult Mice Simo S. Oja and Pirjo Saransaari Medical School, 33014 University of Tampere, Finland Correspondence should be addressed to Simo S. Oja; simo.o[email protected] Received 14 May 2015; Accepted 21 July 2015 Academic Editor: Hari Shanker Sharma Copyright © 2015 S. S. Oja and P. Saransaari. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The release of preloaded [3H]taurine from hippocampal slices from developing 7-day-old and young adult 3-month-old mice was studied in a superfusion system in the absence of glucose. These hypoglycemic conditions enhanced the release at both ages, the effect being markedly greater in developing mice. A depolarizing K+concentration accentuated the release, which indicates that it was partially mediated by exocytosis. The anion channel blockers were inhibitory, witnessing the contribution of ion channels. NOgenerating agents fomented the release as a sign of the participation of excitatory amino acid receptors. The other second messenger systems were apparently less efficient. The much greater taurine release could be a reason for the well-known greater tolerance of developingnervoustissuetolackofglucose. 1. Introduction Hypoglycemia is a metabolic condition common during development and may lead to severe neurological defects in human infants. However, the effects of hypoglycemia on the developing brain are still incompletely understood [1]. The brain derives most of its energy from the oxidation of glucose, but during development it also has an ability to utilize alternative energy substrates which may offer protection during hypoglycemia [2, 3]. On the other hand, poor reserves of high-energy phosphates and a high metabolic rate may predispose the developing brain to hypoglycemic injury [4, 5]. In the brain the hippocampus is the region most sensitive to lack of oxygen and glucose. Energy deprivation leads to neuronal cell death, caused primarily by excitotoxicity due to excessive glutamate release [6, 7]. Taurine (2-aminoethanesulfonic acid) is present at high concentrations in the brain. During ontogenic development its concentration even exceeds that of the main excitatory transmitter glutamate [8]. It increases membrane chloride conductance, causing hyperpolarization and inhibiting neuronal firing [9, 10]. Taurine also attenuates the excessive neuronal accumulation of Ca2+,whichpredisposescells to damage [11] and prevents or reduces the glutamateinduced elevation of intracellular Ca2+ [12] by inhibiting the glutamate-induced release of Ca2+ from the internal pools [13] and the glutamate-induced Ca2+ influx through L-, P/Q-, and N-types of voltage-gated Ca2+ channels [14]. Taurine thus regulates cytoplasmic and mitochondrial calcium homeostasis [15] and in this manner protects neural cells against the toxicity of excitatory amino acids in the hippocampus [16]. Cell-damaging conditions, including hypoglycemia, increase the release of taurine [17, 18] together with that of excitatory amino acid neurotransmitters. Our primary assumption is that taurine release in the absence of an adequate supply of glucose could protect neural cells from injury. In the present study we therefore examined the general properties of taurine releaseinthehippocampusinhypoglycemiaandhowthe releaseisaffectedbyionchannels,secondmessengersystems, and adenosine receptors. 2. Materials and Methods 2.1. Materials. Developing (7-day-old) and young adult (3month-old) NMRI mice of both sexes were used in the Hindawi Publishing Corporation Journal of Amino Acids Volume 2015, Article ID 254583, 7 pages http://dx.doi.org/10.1155/2015/254583 2Journal of Amino Acids experiments. All efforts were made to minimize both the suffering and the number of the animals used. The experiments conformed to the European Community Directive (86/609/EEC) for ethical use of experimental animals and were approved by the Committee of Tampere University for animal experiments. [3H]Taurine (specific radioactivity 1.15 PBq/mol) was obtained from Amersham International, Bristol, UK. The various effectors were purchased from the Tocris Bioscience (Bristol, UK) or Sigma Aldrich (St. Louis, MO). 2.2. Release Experiments. Coronal slices 0.4 mm thick weighing 15–20 mg were manually prepared from the mouse hippocampus with a tissue slicer of Stadie-Riggs type. The slices were immediately immersed in 5 mL of oxygenated medium and incubated with 0.01 mM [3H]taurine (50 MBq/L) at 37∘C for 30 min under agitation. The standard Krebs-RingerHepes medium contained (in mmol/L) NaCl 127, KCl 5, CaCl2 0.8, MgSO41.3, Na2HPO41.3, N-2-hydroxyethylpiperazineN󸀠-2-ethanesulphonic acid (Hepes) 15, NaOH 11, and Dglucose10(pH7.4).Thesliceswerethentransferredinto 0.25 mL cups and superfused with the above medium at a rate of 0.25 mL/min for 50 min in a system in which freely floating shaken slices were kept under a continuous flow of oxygen in order to preserve their viability [19]. Hypoglycemic conditions were induced by omitting glucose from the superfusion media. Potassium stimulation was applied from 30 to 50 min with 50 mM K+. In our experimental setup this K+concentration has yielded the best and most reproducible responses in GABA and taurine release [19]. This high K+concentration may cause release not only from neurons but from glial cells as well [20]. However, taurine release is typically slow at onset and prolonged and high K+concentrations above those prevailing in vivo should be used in in vitro experiments [21]. The different effectors were added to the medium at the onset of superfusions, as explained in the table legends. The superfusion medium was pooled during the first 20min, whereafter 2min fractions (0.5 mL) were collected directly into small scintillation vials with a fraction collector. After superfusion the slices were weighed, homogenized in ice-cold 5% (w/v) trichloroacetic acid solution, and centrifuged, and the clear supernatants were used for scintillation counting. The effluent samples weresubjectedtothesameanalyses. 2.3. Estimation of Efflux Rate Constants. Desaturation curves of labeled taurine from the slices were plotted as a function of time on the basis of the radioactivities remaining in theslicesaftersuperfusionandrecoveredinthecollected superfusate fractions [19]. During superfusion the release of labeled taurine originates initially from the extracellular spaces in slices. This source is gradually exhausted during thefirst20minandthereleasesubsequentlyoccursfromthe intracellular pools. The efflux rate constants of taurine for the time intervals of 20 to 30 min (𝑘1,initialreleasephase)and 34–50 min (𝑘2,laterreleasephase)werecomputedasnegative slopesfortheregressionlinesofthelogarithmofradioactivity remaining in the slices versus superfusion time. There were no differences between the results from male and female mice and the results from both sexes were therefore combined. 2.4. Statistical Analysis. The significance of the results was analyzed with two-way analysis of variance (ANOVA) using SPSS statistics, version 17.0, computer program. The analyses were done by grouping the results according to the nature of theeffectorsstudied,potassiumstimulation,chloridechannel blockers, NO-generating agents, adenosine agonists, and second messengers. When significant effects were detected, the post hoc Bonferroni test was applied to bring out the differences between the sample means. They were considered significant when the calculated 𝑝values were less than 0.05 or 0.01. 3. Results Hypoglycemia significantly enhanced taurine release in both age groups. In adult 3-month-old mice the fractional release rate constant in the presence of glucose in medium was for the superfusion period of 34–50 min (1.54±0.04)×10−3 (𝑛=18) andintheabsenceofglucose(1.93 ± 0.18)×10−3,𝑛=11, significantly different at a level of p= 0.042. The hypoglycemia effect was more marked in 7-day-old mice, the corresponding constants being (0.38±0.20)×10−3,𝑛=11,and(1.20±0.12) ×10−3,𝑛=11,p= 0.000, respectively. Stimulation by 50 mM K+enhancedthereleaseintheabsenceofglucose(df =3, 𝐹 = 23.007,𝑝 = 0.000). The enhancements were 60 per cent and 63 per cent in adult (𝑝 = 0.000) and developing (𝑝= 0.001) mice, respectively (see Table 1). The anion channel blockers generally inhibited the release (df =7,𝐹 = 13.562,𝑝 = 0.000). Of them, DIDS significantly inhibited the K+-stimulatedtaurinereleaseinbothadult (𝑝 = 0.013) and developing (𝑝 = 0.044) mice, being in adultmicealsoeffectiveontheunstimulatedrelease(𝑝= 0.047) (Table 1). SITS, the blocker of chloride transport, was even more effective in all experimental situations (in adult mice, unstimulated release 𝑝 = 0.11,stimulatedrelease, 𝑝 = 0.035, and in developing mice the corresponding data 𝑝 = 0.016 and 𝑝 = 0.017). Another transport inhibitor 9-AC was not effective. All nitric oxide generators, SNAP, SNP, and hydroxylamine, were strong stimulators in taurine release in both adult (df =7,𝐹 = 32.190,𝑝 = 0.000)and developing (df =7,𝐹 = 13.732,𝑝 = 0.000) mice and in both theunstimulatedandK +-stimulated release (Table 2). On the other hand, all adenosine receptor agonists tested, CHA, RPIA and CGS 21680, and riluzole, which inhibits glutamate release and GABA uptake, were without any effects (Table 3). Of compounds involved in the second messenger systems, genistein, with its main known activity as a tyrosine kinase inhibitor, was not effective during the period of 34– 50 (Table 4), whereas quinacrine, a nonselective inhibitor of both monoamine oxidases A and B, inhibited the unstimulated and K+-stimulated release in both age groups at all stages of superfusion (df =6,𝐹 = 23.648,𝑝 = 0.000). The protein kinase C activator PMA only inhibited the unstimulatedreleaseinadultmice(𝑝 = 0.036), whereas the protein kinase inhibitor chelerythrine was not effective. Journal of Amino Acids 3 Table 1: Effects of ion channel inhibitors on taurine release from hippocampal slices from 3-month-old and 7-day-old mice in hypoglycemia. Effectors Efflux rate constants (×10−3min−1)±SEM 3-month-old 7-day-old 𝑘1𝑘2𝑘1𝑘2 Basal (control) 2.36 ±0.11 (20) 1.93 ±0.18 (11) 1.34 ±0.08 (31) 1.20 ±0.11 (11) +50 mM K+(control) 2.92 ±0.21 (10) 1.90 ±0.10 (12) DIDS 0.5 mM 1.34 ±0.15∗∗ (6) 1.31 ±0.10∗(4) 1.12 ±0.14 (8) 1.16 ±0.13 (4) +50 mM K+1.81 ±0.12∗∗ (4) 1.45 ±0.06∗(4) SITS 2.0 mM 1.04 ±0.07∗∗ (8) 0.93 ±0.06∗∗ (4) 0.80 ±0.09∗∗ (6) 0.62 ±0.06∗∗ (4) +50 mM K+1.50 ±0.21∗∗ (4) 0.90 ±0.12∗∗ (4) 9-AC 0.2 mM 2.67 ±0.17 (5) 2.74 ±0.49 (4) 1.82 ±0.23 (8) 1.07 ±0.09 (4) +50 mM K+3.53 ±0.32 (4) 1.84 ±0.10 (4) The drugs were added at the beginning of superfusion and 50 mM K+at 30 min. The results show the efflux rate constants ±SEM (×10−3min−1)for the time intervals of 20–30 min (𝑘1) and 34–50 min without the excess of K+or in the presence of 50 mM K+(𝑘2) with the number of independent experiments in parenthesis. Abbreviations: DIDS, diisothiocyanostilbene-2󸀠2-disulphonate; SITS, 4-acetamido-4󸀠-isothiocyanostilbene-2󸀠2-disulphonate; 9AC, 9-anthracenecarboxylic acid. Significance of differences from the corresponding controls: ∗𝑝 < 0.05,∗∗𝑝 < 0.01. Table 2: Effects of nitric oxide generators on taurine release from hippocampal slices from 3-month-old and 7-day-old mice in hypoglycemia. Effectors Efflux rate constants (×10−3 min−1)±SEM 3-month-old 7-day-old 𝑘1𝑘2𝑘1𝑘2 Basal (control) 2.36 ±0.11 (20) 1.93 ±0.18 (11) 1.34 ±0.08 (31) 1.20 ±0.11 (11) +50 mM K+(control) 2.92 ±0.21 (10) 1.90 ±0.10 (12) SNAP 1.0 mM 4.31 ±0.22∗∗ (7) 5.44 ±0.14∗∗ (4) 2.84 ±0.11∗∗ (8) 2.17 ±0.07∗∗ (4) +50 mM K+5.35 ±0.15∗∗ (4) 2.34 ±0.08∗∗ (4) SNP 1.0 mM 3.17 ±0.13∗∗ (15) 4.23 ±0.38∗∗ (8) 1.80 ±0.21∗(7) 1.65 ±0.13∗(4) +50 mM K+4.41 ±0.77∗∗ (4) 1.98 ±0.10∗∗ (4) Hydroxylamine 5.0 mM 5.16 ±0.17∗∗ (7) 6.04 ±0.76∗∗ (7) 3.82 ±0.33∗∗ (8) 3.44 ±0.16∗∗ (4) +50 mM K+7. 03 ±0.28∗∗ (4) 3.84 ±0.10∗∗ (4) The effectors were added at the beginning of superfusion and 50 mM K+at 30 min. The results show the efflux rate constants ±SEM (×10−3 min−1)forthe time intervals of 20–30 min (𝑘1) and 34–50 min without the excess of K+or in the presence of 50 mM K+(𝑘2) with the number of independent experiments in parenthesis. SNAP: S-nitroso-N-acetylpenicillamine; SNP: sodium nitroprusside. Significance of differences from the corresponding controls: ∗𝑝 < 0.05, ∗∗𝑝 < 0.01. Table 3: Effects of adenosine agonists and riluzole on taurine release from hippocampal slices from 3-month-old and 7-day-old mice in hypoglycemia. Effectors Efflux rate constants (×10−3 min−1)±SEM 3-month-old 7-day-old 𝑘1𝑘2𝑘1𝑘2 Basal (control) 2.36 ±0.11 (20) 1.93 ±0.18 (11) 1.34 ±0.08 (31) 1.20 ±0.11 (11) +50 mM K+(control) 2.92 ±0.21 (10) 1.90 ±0.10 (12) CHA 0.5 mM 2.98 ±0.12 (6) 1.85 ±0.30 (6) 1.48 ±0.07 (10) 1.11 ±0.06 (7) +50 mM K+2.42 ±0.27 (4) 2.00 ±0.06 (4) R-PIA 0.1 mM 2.46 ±0.17 (6) 1.92 ±0.21 (6) 1.97 ±0.19 (6) 1.41 ±0.12 (4) +50 mM K+3.63 ±0.51 (4) 2.12 ±0.11 (4) CGS 21680 10.0 mM 2.59 ±0.22 (4) 2.20 ±0.20 (4) 1.75 ±0.16 (6) 1.38 ±0.20 (4) +50 mM K+3.44 ±0.32 (4) 2.28 ±0.11 (4) Riluzole 0.1 mM 2.47 ±0.17 (7) 1.86 ±0.27 (4) 1.86 ±0.15∗(6) 1.38 ±0.10 (4) +50 mM K+2.50 ±0.35 (4) 2.23 ±0.15 (4) The agonists were added at the beginning of superfusion and 50 mM K+at 30 min. The results show the efflux rate constants ±SEM (×10−3 min−1)for the time intervals of 20–30 min (𝑘1) and 34–50 min without the excess of K+or in the presence of 50 mM K+(𝑘2) with the number of independent experiments in parenthesis. CHA: N6-cyclohexyladenosine; R-PIA: (R(−)N6-(2-phenylisopropyl)adenosine; CGS 21680: 4-[2-[[6-amino-9-(N-ethyl-𝛽-Dribofuranuronamidosyl)-9H-purin-2-yl]amino]ethyl]benzenepropanoic acid hydrochloride. Significant difference from the control: ∗𝑝 < 0.05. 4Journal of Amino Acids Table 4: Effects of compounds involved in the second messenger systems on taurine release from mouse hippocampal slices in hypoxia. Concentration (mM) Efflux rate constants (×10−3 min−1)±SEM 3-month-old 7-day-old 𝑘1𝑘2𝑘1𝑘2 Basal (control) 2.36 ±0.11 (20) 1.93 ±0.18 (11) 1.34 ±0.08 (31) 1.20 ±0.11 (11) +50 mM K+(control) 2.92 ±0.21 (10) 1.90 ±0.10 (12) Genistein 0.001 2.59 ±0.13 (7) 2.06 ±0.18 (7) 1.92 ±0.16∗(7) 1.44 ±0.16 (4) +50 mM K+3.07 ±0.25 (4) 2.27 ±0.06 (4) Quinacrine 0.01 1.20 ±0.08∗∗ (7) 0.83 ±0.05∗∗ (4) 0.75 ±0.07∗∗ (8) 0.61 ±0.06∗∗ (4) +50 mM K+1.12 ±0.13∗∗ (4) 0.79 ±0.05∗∗ (4) PMA 0.00001 2.31 ±0.20 (8) 1.77 ±0.11 (4) 1.80 ±0.15∗(8) 1.33 ±0.17 (4) +50 mM K+1.95 ±0.17∗(4) 1.85 ±0.03 (4) Chelerythrine 0.001 2.49 ±0.17 (7) 2.07 ±0.14 (4) 1.73 ±0.18 (8) 1.53 ±0.04 (4) +50 mM K+3.40 ±0.51 (4) 2.01 ±0.09 (4) IBMX 1.0 1.85 ±0.16 (8) 2.51 ±0.29 (4) 1.99 ±0.24 (6) 1.33 ±0.11 (4) +50 mM K+2.70 ±0.13 (4) 1.85 ±0.03 (4) RO 20-1724 0.2 3.46 ±0.12∗∗ (8) 3.43 ±0.15∗∗ (4) 1.96 ±0.26 (7) 1.27 ±0.17 (4) +50 mM K+4.56 ±0.30∗∗ (4) 2.65 ±0.33∗(4) Zaprinast 0.1 2.31 ±0.15 (8) 2.00 ±0.20 (8) 1.00 ±0.13 (6) 0.66 ±0.12∗(4) +50 mM K+2.53 ±0.06 (4) 2.04 ±0.23 (4) ODQ 0.01 2.51 ±0.18 (7) 2.01 ±0.13 (4) 1.50 ±0.17 (8) 1.19 ±0.08 (8) +50 mM K+2.50 ±0.25 (4) 2.04 ±0.06 (4) Alloxan 5.0 2.81 ±0.31 (7) 2.19 ±0.06 (4) 1.70 ±0.17 (6) 1.58 ±0.25 (4) +50 mM K+3.13 ±0.10 (4) 1.86 ±0.07 (4) The agonists were added at the beginning of the superfusion and 50 mM K+at 30 min. The results show the efflux rate constants ±SEM (×10−3 min−1)forthe time intervals of 20–30 min (𝑘1)andfor34–50minwithouttheexcessofK +or in the presence of 50 mM K+(𝑘2) with the number of independent experiments in parenthesis. PMA: 4𝛽-phorbol 12-myristate 13-acetate; IBMX: 3-isobutyl-1-methylxanthine; ODQ: 1H-[1,2,4]oxadiazolo[4,3]quinoxalin-1-one; RO 20-1724: 4-(3-butoxy-4-methoxyphenyl)-2-imidazolidone. Significance of differences from the corresponding controls: ∗𝑝 < 0.05,∗∗𝑝 < 0.01. The nonspecific inhibitor of cAMP and cGMP phosphodiesterases and the nonselective adenosine receptor antagonist IBMX was not effective during the period of 34–50 min. The selective inhibitor of cGMP-insensitive phosphodiesterase, type IV, RO 20-1724 enhanced both the unstimulated (𝑝= 0.008)andstimulated(𝑝 = 0.005) release in adult mice and alsoslightlythestimulatedrelease(𝑝 = 0.038)indeveloping mice (Table 4). The selective inhibitor of cGMP-specific phosphodiesterases V and VI (PDE5/6) and the agonist at the G protein-coupled receptor 35 zaprinast significantly (𝑝= 0.035) reduced the unstimulated release in 7-day-old mice, whereas ODQ, the potent and selective inhibitor of NOsensitive guanylyl cyclase, was without effect. Alloxan, the unspecific inhibitor of adenylyl cyclases, was likewise not effective during the period of 34–50 min. 4. Discussion The present results show taurine release to be enhanced in hypoxia at both ages studied, the effect being more pronounced in the developing hippocampus. The release is largelymediatedbyanionchannels,sincetheanionchannel inhibitors markedly reduced it. The nitric acid generators strongly stimulated the release. The effects of the second messenger system were more variable. These actions were also in many cases different in adult and developing mice. The enhancement could be due to several mechanisms, including Ca2+-dependent exocytosis, Ca2+-independent release via reversal of carrier-mediated uptake, indiscriminate opening of ion channels which allow the passage of taurine molecules, or leakage through damaged plasma membranes. Depolarization by K+-stimulation was now able to further potentiate taurine release at both ages under hypoglycemia. This may signify release via exocytosis, which is also preserved in hippocampal slices when they are exposed to even more drastic cell-damaging conditions such as ischemia (the absence of glucose and oxygen atmosphere) [17, 18]. In developing mice taurine release in the absence of glucose is approximately the same as in the presence of nitrogen atmosphere without glucose, whereas in adults these ischemic conditions have had even more impact [18]. Both neurons and glial cells have been shown to contain taurine [22], and thus only a part of the released taurine may originate from neurons. K+stimulation of taurine release has also been shown to be associated with cell swelling [23]. Indeed, the volume-regulated anion channels may contribute, together with the reversed function of astrocytic glutamate transporters,upto80percentofthetotalreleaseoftaurine and excitatory amino acids in global cerebral ischemia in rats [24]. Intracellular swelling activates stretch-sensitive (volumeregulated) ion channels and is accompanied by the release Journal of Amino Acids 5 of both inorganic and organic osmolytes, including taurine [25]. For instance, the volume-regulated anion channels have been shown to be the predominant contributors to the release of excitatory amino acids in the ischemic cortical penumbra [26]. The swelling-induced increase in taurine release is a diffusional process without any carrier involvement [27]. In both the developing and adult hippocampus the presynaptically acting ionotropic glutamate agonists significantly increase the release of taurine in a receptor-mediated manner [28, 29]. For instance, of the ionotropic receptors, NMDA receptor activation in particular has been assumed to play a central role in hypoglycemia-induced glutamate release [30]. Activation of NMDA receptors allows Ca2+ to enter the cells. NO synthase is a Ca2+-dependent enzyme [31], being activated in the presence of Ca2+ andthenproducing NO.NOstimulatessolubleguanylatecyclaseandinthis manner foments the production of 3󸀠,5󸀠-cyclic guanosine monophosphate [32], which enhances taurine release [33]. The present activatory effect of the NO generators studied is in concert with this sequence of events. NO also regulates neuronal activity by activating calcium-dependent potassium channels [34, 35]. NO can be the endogenous opener of leak K+channels in the hippocampus as shown in basal forebrain cholinergic neurons [36]. The increase in extracellular K+ enhances taurine release. The NO donors did not markedly foment the K+-evoked release now. The effect was apparently already maximal at the K+concentration used in this study. NO produced by an exogenous NO donor has been shown to block both tetrodotoxin-sensitive and tetrodotoxinresistant Na+currents in baroreceptor neurons [37] and NO to block different Na+channels in baroreceptor neurons [38]. In contrast, NO donors SNP and SNAP have significantly increased the mean persistent current in excised inside-out patches from cultured hippocampal neurons [39]. Such results suggest that, depending on cell type, NO may modulate Na+currents differently. Taurine efflux has been shown to be dependent on the presence of both Na+and Cl−ions [40]. The reduction of release by the Cl−channel antagonists SITS and DIDS indicates that the release may also occur through anion channels, similar to the volumesensitive taurine release in astrocytes and neurons [41, 42]. The present attenuating effect of quinacrine on the release is furthermore in line with this assumption since quinacrine has been shown to inhibit the hypotonically induced whole cell Cl−currents in human submandibular gland (HSG) cells [43]. The various second messengers exerted less pronounced effects on taurine release. In keeping with this the effects of different metabotropic glutamate receptor agonists and antagonists have also not been particularly effective in taurine release in the hippocampus [44]. The presynaptic adenosine receptors, particularly of the A1class, are known to regulate neurotransmitter release [45]. The A1receptors have also enhanced taurine release in the adult hippocampus, but only when it was subjected to K+stimulation in ischemia [46]. In the present study the nonspecific antagonist of adenosine receptors likewise exhibited only minor effects. 5. Conclusions The release of taurine in the hypoglycemic hippocampus of both young and adult mice appears to be a complex process mediated by several different mechanisms, such as exocytosis, carrier reversal, diffusion via ion channels, and probably leakage through partially damaged cell membranes. Hypoglycemia also evokes the release of excitatory amino acids [6, 18]. Glutamate release overactivates its receptors and functions as an excitotoxic agent. There is a rationale here in that the activation of excitatory amino acid receptors causes a concomitant increase in taurine release. Taurine is inhibitory in nature and counteracts this excessive excitation. In the developing brain taurine release is much larger in magnitude than in the adult brain, which could be a reason for the wellknowngreatertoleranceofdevelopingnervoustissuetolack of glucose. Abbreviations 9-AC: 9-Antracenecarboxylate AIDA: (RS)-1-Aminoindan-1,5dicarboxylate AMPA: 2-Amino-3-hydroxy-5-methyl-4isoxazolepropionate 2R,4R-APDC: (2R,4R)-4-Aminopyrrolidine-2,4dicarboxylate cAMP: Cyclic adenosine monophosphate ATPase: Adenosine triphosphatase cGMP: Cyclic guanosine monophosphate CGS 21680: 4-[2-[[6-Amino-9-(N-ethyl-𝛽-Dribofuranuronamidosyl)-9Hpurin-2yl]amino]ethyl]benzenepropanoic acid hydrochloride CHA: N6-Cyclohexyladenosine DHPG: (S)-3,5-Dihydroxyphenylglycine DIDS: Diisothiocyanostilbene-2,2󸀠- disulphonate DMPX: 3,7-Dimethyl-1-propargylxanthine EDTA: Ethylenediaminetetraacetate EGLU: (2S)-2-Cyclopropyl-4phosphonophenylglycine GABA: 𝛾-Aminobutyrate GAT: GABA transporter HA: Hydroxylamine Hepes: N-2-Hydroxyethylpiperazine-N󸀠- 2-ethanesulphonic acid IBMX: 3-Isobutyl-1-methylxanthine L-AP4: L(+)-2-Amino-4phosphonobutyrate L-NAME: N6-Nitro-L-argininemethylester L-SOP: O-Phospho-L-serine MK-801 (dizocilpine): (5S,10R)-(+)-Methyl-10,11dihydro-5Hdibenzo(a,d)cyclohepten-5,10amine 6Journal of Amino Acids NBQX: 2,3-Dioxo-6-nitro-1,2,3,4tetrahydrobenzo[f]quinoxaline-7sulfonamide NMDA: N-Methyl-D-aspartate ODQ: 1H-(1,2,4)Oxadiazolo(4,3-a)quinoxalin-1one PDE: Phosphodiesterase PMA: 4𝛽-Phorbol 12-myristate 13-acetate PKC: ProteinkinaseC RO 20-1724: 4-(3-Butoxy-4-methoxyphenyl)-2imidazolidone R-PIA: R(−)N6-(2-Phenylisopropyl)adenosine SITS: 4-Acetamido-4󸀠-isothiocyanostilbene2,2󸀠-disulphonate SNAP: S-Nitroso-N-acetylpenicillamine SNP: Sodium nitroprusside. 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