a-b. Time-course of 20 or 100 μM NMDA induced CREB phosphorylation in ACSF (a) or conditioned medium (b) with 1, 3 or 9 mM Mg2+; Mg2+ elevation enhanced NMDA-induced CREB phosphorylation at Ser133 (pCREB). Magnesium ion concentration in medium is around 0.8 mM. Adding 2.2 and 8.2 mM extra magnesium in conditioned medium led to approximately 3 mM and 9 mM external magnesium concentrations. Neurons was incubated with extra Mg2+ containing ACSF or condition medium for one hour before the NMDA application. c. Mg2+ enhances NMDA-induced CREB phosphorylation in mouse brain slices. Acute mouse brain slices was preconditioned with ACSF containing 3 mM Mg2+ for 1 hour. Brain slices were then incubated with 20 μM NMDA for 10 minutes. (*, P<0.05; **, P<0.01, N=3). Error bar in this figure and all other figures indicate standard error of the mean (SEM). Con: indicating control. In supplemental figure legends, C indicates control for space reasons.