In vivo microdialysis was performed as previously described (Cirrito et al., 2003 (link); Castellano et al., 2011 (link)). In brief, microdialysis guide cannula and probes with a 38-kD molecular weight cutoff membrane (BR-2 probes; Bioanalytical Systems) were inserted into the left hippocampus at the following coordinates: bregma, −3.1 mm; midline, −2.5 mm; and tip, 3.2 mm below dura at a 12° angle. Perfusion buffer (0.15% BSA in artificial cerebrospinal fluid [in mM: 1.3 CaCl, 1.2 MgSO4, 3 KCl, 0.4 KH2PO4, 25 NaHCO3, and 122 NaCl, pH 7.35]) was perfused at a 1-µl/min flow rate with a syringe pump (Stoelting Co). Microdialysis samples were collected every 60–90 min using a refrigerated fraction collector (Univentor). Mice were housed in a RaTurn Caging system (Bioanalytical Systems) with ad libitum food and water for the duration of the experiment. LA-1 and Compound E (AsisChem) were perfused directly into the hippocampus through the microdialysis probe (reverse microdialysis) at a concentration of 100 µM and 200 nM, respectively.
Quantitative measurements of Aβ collected from in vivo microdialysis fractions were performed by sandwich ELISA, as previously described (Cirrito et al., 2011 (link)). ELISA plates were coated with a mouse anti-Aβ40 selective antibody, mHJ2, and detected with a biotinylated central domain mouse anti-Aβ (amino acids 13–28) antibody, mHJ5.1.
Quantitative measurements of Aβ collected from in vivo microdialysis fractions were performed by sandwich ELISA, as previously described (Cirrito et al., 2011 (link)). ELISA plates were coated with a mouse anti-Aβ40 selective antibody, mHJ2, and detected with a biotinylated central domain mouse anti-Aβ (amino acids 13–28) antibody, mHJ5.1.