Wistar rat hippocampal slices were prepared with a chilled cutting solution containing choline chloride, sodium carbonate, glucose, sodium ascrobate, magnesium sulfate, sodium pyruvate, posstassium chloride, calcium chloride, sodium phosphate. The slices were incubated with physiological saline and gassed with 95% oxygen, 5% carbon dioxide.
For experiments NBQX, bicuculline and serine (Sigma) were added to the physiological saline.
Whole-cell patch electrodes containing Fluo5F and Alexa Fluor 594 (Molecular Probes), Cesium metholsulfate, HEPES, sodium phosphocreatine, glutathione, magnesium chloride, sodium ATP and sodium GTP.
Paired pulses and single pulses were alternated every 5 seconds.
Custom Ti:sapphire laser (Mira, Coherent)scanning microscope and Zeiss lens was used. Fluorescent image acquisition was controlled by software from Matlab (Mathworks). Stimulated synapses were identified by an on-line analysis program written in IGOR (Wavemetrics).
Statistics used the Wilcoxon-two sample test.