Coverslips were mounted on a closed perfusion chamber used to apply different media to the cells. Standard media (ACSF) consisted of HEPES-buffered standard physiological perfusion medium14 (link) and containing NaCl 140 mM, KCl 3 mM, CaCl2 3 mM, MgCl2 2 mM, Glucose 5 mM, HEPES 10 mM, adjusted to a pH of 7.3 with NaOH.
All experiments were performed at room temperature. Glutamate, Glycine, D-Serine, L-Lactate, D-Lactate, Pyruvate, MK801, Ifenprodil, dithiothreitol (DTT), 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB), Stiripentol were provided by Sigma-Aldrich and added to ACSF. For most experiments, 2 successive pulses of glutamate (1–100 μM) and glycine (100 μM) are applied to the cells spaced out by a minimum delay of 20 min. The amplitude of the 1st and the 2nd Ca2+ response induced by the glutamate/glycine cocktail are compared. For the experiments performed at 1 mM of glutamate (+100 μM of glycine), only a single pulse is applied due to a higher number of excito-toxic responses. Finally, for some experiments, 5 mM of Glucose are added to the ACSF (final concentration of Glucose: 10 mM; annotated 5 mM extra Glucose ACSF).
All experiments were performed at room temperature. Glutamate, Glycine, D-Serine, L-Lactate, D-Lactate, Pyruvate, MK801, Ifenprodil, dithiothreitol (DTT), 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB), Stiripentol were provided by Sigma-Aldrich and added to ACSF. For most experiments, 2 successive pulses of glutamate (1–100 μM) and glycine (100 μM) are applied to the cells spaced out by a minimum delay of 20 min. The amplitude of the 1st and the 2nd Ca2+ response induced by the glutamate/glycine cocktail are compared. For the experiments performed at 1 mM of glutamate (+100 μM of glycine), only a single pulse is applied due to a higher number of excito-toxic responses. Finally, for some experiments, 5 mM of Glucose are added to the ACSF (final concentration of Glucose: 10 mM; annotated 5 mM extra Glucose ACSF).
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