Hydrocortisone

Cell culture reagents were purchased from Life Technologies [Trypsin-EDTA (0.05%), TrypLE, N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid (HEPES), Anti-Anti, B-27 supplement along with Neurobasal, Dulbecco’s modified Eagle’s media (DMEM) and Minimum Essential Media (MEM) medias], Sigma-Aldrich [Cytosine β-D-arabinofuranoside (Ara-C), poly-D-lysine (70–150 kDa), Hank’s Balanced Salt Solution (HBSS), 1,4-Piperazinediethanesulfonic acid (PIPES), 3-(N-morpholino)propanesulfonic acid (MOPS), Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA), insulin, N-Acetyl Cysteine (NAC), hydrocortisone, sodium pyruvate and GlutaMAX], Worthington (Trypsin, Papain, Papain Dissociation Kit, and DNase), Atlanta Biologicals [Horse Serum (HS) and Fetal Bovine Serum (FBS)]. Zinc indicators (FluoZin-3 AM and Newport Green DCF) were obtained from Life Technologies. Vitamin MEM solution, amino acid MEM solution, ZnCl₂ (0.1 M stock solution), N,N,N′,N′-Tetrakis (2-pyridylmethyl)ethylenediamine (TPEN), 2-Mercaptopyridine N-oxide sodium salt (pyrithione) were all purchased from Sigma-Aldrich. For GluA2 surface labeling, Alexa 488-CAM2 and Alexa 647-CAM2 were designed and synthesized by the Hamachi research group (Kyoto University; Wakayama et al., 2017 (link)).

MCF10A human breast epithelial cells were maintained in Dulbecco’s modified Eagle medium (DMEM)/Ham’s F-12 (50/50) medium supplemented with epidermal growth factor 20 ng/mL, insulin $10\mu \mathrm{g}/\mathrm{mL}$ , hydrocortisone $0.5\mu \mathrm{g}/\mathrm{mL}$ , cholera toxin 100 ng/mL, 5% horse serum (Atlanta Biologicals), and 1% antibiotics and antimycotics (Mediatech, Manassas, Virginia). Triple-negative breast tumor cell lines (HCC1395, BT-20, MDA-MB-231, and Hs578T) were cultured in RPMI 1640 (HCC1395), EMEM (BT-20), or DMEM (MDA-MB-231, Hs578T) medium supplemented with 9% fetal bovine serum (Atlanta Biologicals) and 1% antibiotics and antimycotics. All cell lines were purchased from American Type Culture Collection (ATCC, Manassas, Virginia). Cells were maintained at 37°C in a cell culture incubator with 5% ${\mathrm{CO}}_2$ .