S ampa

(S)-AMPA (Sigma-Aldrich, MO) and ZK 200775, a competitive AMPA receptor antagonist (Tocris Bioscience, MO) were dissolved in artificial cerebrospinal fluid (aCSF) consisting of (in mM): 148 NaCl, 2.7 KCl, 1.2 CaCl₂, and 0.85 MgCl₂, pH adjusted to 7.0–7.5.

Mouse hippocampal neurons from wild type E16–18 animals were isolated and cultured for 13 days in vitro in Neurobasal media with B27 Supplement, Glutamax, and penicillin/streptomycin (Invitrogen). Neuron cultures were then treated with Aβ in the absence or presence of PICK1 inhibitorsfor 24 hours. Calcium influx was measured using the FLIPR Calcium 4 Assay (Molecular Devices). The cells were loaded with dye for 1 hour at 37 °C and then stimulated with 40 μM s-AMPA (Sigma). Fluorescent recordings were taken using the FLIPR Tetra (Molecular Devices). Baseline measurements before AMPA stimulation were recorded and fluorescent intensity measurements were taken for roughly 3 minutes during and after stimulation. Calcium influx was calculated as the area under the curve and plotted using PRISM GraphPad software.

Veratridine, dihydrokainic acid (DHK), 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide disodium salt (NBQX), (S)-α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (s-AMPA) and citalopram hydrobromide were purchased from Tocris (Bristol, UK). 4-chloro-dl-phenylalanine methyl ester hydrochloride (pCPA) was purchased from Sigma-Aldrich (Tres Cantos, Spain).
For intracerebral microinfusion, Veratridine was dissolved in 12.5% dimethyl sulfoxide in artificial cerebrospinal fluid and pH adjusted to 6.5; DHK and s-AMPA were dissolved in PBS 10 × as previously reported;²⁹ NBQX and citalopram were dissolved in artificial cerebrospinal fluid. The microinfusion volume was 0.5 μl per side in all cases. pCPA was administered intraperitoneally, was daily prepared by dissolving in saline and pH was adjusted to 6.0 with 0.1 m NaOH.^{30 (link)}