Cyclothiazide ctz

Salts were of analytical grade and obtained from Sigma-Aldrich. TTX was obtained from Carl Roth, Tetraethylammonium chloride (TEA) and 4-aminopyridine (4AP) from Sigma-Aldrich. AMPA, Cyclothiazide (CTZ), 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline (NBQX), N-Methyl-D-aspartate (NMDA) and Glycine were obtained from Tocris Bioscience. For voltage-gated conductances, the basic extracellular solution contained (in mM): NaCl 137, KCl 2.9, CaCl₂ 2.1, MgCl₂ 1.2, HEPES 10, Glucose 10 (pH 7.8). The pipette solution was composed of (in mM): KCl 130, MgCl₂ 4, EGTA 10, HEPES 10 and Na₂-ATP 2 (pH 7.4). For glutamate receptors the basic extracellular solution contained (mM): NaCl 134, KCl 2.9, CaCl₂ 2.1, MgCl₂ 1.2, HEPES 10, Glucose 10 (pH 7.8). The pipette solution was composed of (in mM): CsCl 134, MgCl₂ 2, EGTA 10, HEPES 10, Na₂-ATP 4 and Li-GTP 0.4 (pH 7.4). During the measurements, solutions were applied by the DAD VM SuperFast perfusion system (Scientific Instruments) and driven away with the use of a CVC2000 vacuum pump.

Neurobasal^®® medium, B-27 supplement, 100× antibiotic–antimycotic (10.000 units/mL penicillin, 10.000 µg/mL streptomicyn and 25 µg/mL Fungizone^®®) and calcein-AM (calcein acetoxymethyl ester) were purchased from ThermoFisher Scientific (Waltham, MA, USA). NMDA (N-methyl-D-aspartic acid), Sephin1, guanabenz, HBSS, glycine, poly-L-ornithine, glutamine, thapsigargin and tunycamicin were obtained from MilliporeSigma/Merck (Burlington, MA, USA). AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) and cyclothiazide (CTZ) were obtained from Tocris Biosciences (Bristol, UK).

Reverse microdialysis was used to administer compounds directly into the hippocampus. Drugs were diluted into the perfusion buffer of artificial CSF and 0.15% BSA, allowing the drugs to diffuse into the brain continuously for the duration of the experiment at the same time that Aβ is collected. Due to the complexity of determining the final concentration of compound delivered to the brain, only the starting concentrations of drugs in the perfusion buffer are given. We estimate approximately 10% of the drug is delivered across the probe membrane where it is further diluted in the brain CSF. AMPA (0.5, 2, 5, 7.5, and 10 μM), MK801 (100 μM), NMDA (40 μM), and thiorphan (10 μM) were purchased from Sigma. Cyclothiazide (CTZ; 300 μM), tetrodotoxin (TTX; 5 μM), NBQX (100 μM), and GM6001 (25 μM) were purchased from Tocris Bioscience (Ellisville, MO). LY411575 (Sigma-Aldrich) was diluted in corn oil and administered subcutaneously at 5 mg/kg.