Valspodar

Tacrine hydrochloride and MK571 were obtained from Cayman Chemical Company (Ann Arbor, MI). Collagenase (type I, class I), rat tail collagen (type I), Dulbecco’s modified Eagle’s medium (DMEM), and insulin were purchased from Invitrogen (Carlsbad, CA). Fluvoxamine maleate, verapamil hydrochloride, 5 - (and – 6) – carboxy - 2’, 7 ’-dichlorofluorescein diacetate (CDFDA), dexamethasone, tetraethylammonium chloride (TEA), bovine serum albumin (BSA), soybean trypsin inhibitor, and fetal bovine serum (FBS) were purchased from Sigma-Aldrich (St. Louis, MO). ITS culture supplement (10μg/ml insulin, 10μg/ml transferrin, 10ng/ml selenous acid) was obtained from BD Biosciences (San Jose, CA). Valspodar was obtained from XenoTech (Lenexa, KS). Total protein measurement’s reagents with the bicinchoninic acid (BCA) method were from Pierce (Rockford, IL). All other chemicals and reagents were of analytical grade and were readily available from commercial sources.

In vivo Aβ₄₀ clearance was investigated using the BEI method as described previously (Qosa et al., 2012 (link)). In brief, a stainless steel guide cannula was implanted stereotaxically into the right caudate nucleus of mice that had been anesthetized with intraperitoneal xylazine and ketamine (20 and 125 mg/kg, respectively) (Henry Schein Inc., NY). After 12 h recovery period, animals were re-anesthetized and tracer fluid (0.5 μl) containing ¹²⁵I-Aβ₄₀ (30 nM, PerkinElmer, MA) and ¹⁴C-inulin (0.02 μCi, American Radiolabeled Chemicals, MO) prepared in extracellular fluid buffer (ECF) was administered. Thirty minutes post ¹²⁵I-Aβ₄₀ injection (Cirrito et al., 2005 (link); Shibata et al., 2000 (link)); brain tissues were rapidly collected for ¹²⁵I-Aβ₄₀ analysis. To characterize role of P-gp and LRP1, 0.5 μl of ECF containing valspodar (40 μM; XenoTech, KS), a well-established P-gp inhibitor, or RAP (1 μM; Oxford Biomedical Research, MI), an LRP1 inhibitor, were intracerebrally administered 5 min prior to ¹²⁵I-Aβ₄₀ injection.