Journal: Neuropsychopharmacology

Article Title: Prenatal exposure to the mineralocorticoid receptor antagonist spironolactone disrupts hippocampal area CA2 connectivity and alters behavior in mice

doi: 10.1038/s41386-024-01971-7

Figure Lengend Snippet: a On the first day of pregnancy (gestational day 0; GD0), each Amigo2-GFP dam was paired with a male C57BL/6J mouse. Pregnancy was confirmed by the presence of a copulation plug after pairing. Spiro pellets (15 mg/pellet) were implanted in pregnant dams at gestational day 12 (+/−1; GD12). High-performance liquid chromatography-mass spectrometry (HPLCMS) was used to confirm the presence of spiro and metabolites in a subset of dams and pups collected on the day of birth (first postnatal day; P0). Other litters were allowed to age until P60 for behavioral assays prior to immunofluorescent studies. b RNA in situ hybridization of the gene encoding MR, Nr3c2 (magenta), in a murine hippocampus at P0. Scale bar is 200 µm. c Log 2 (x + 1) transformed peak integrated values (arbitrary units; a.u.) of spiro and metabolites in serum and brain homogenates of treated (red circles) and control (black circles) subjects. Displayed analytes include spiro and metabolites canrenone (can), 7α-Thiomethylspironolactone (7-alpha), and 6-beta-7-alpha-thiomethylspironolactone (6-beta). Two-way ANOVA analysis revealed that there was a main effect of treatment on transformed integrated peak values (two-way, ANOVA, F (1,6) = 16363, p < 0.0001). Post hoc analysis revealed that integrated areas differed between control and treatment groups for can, 7-alpha, and 6-beta (Šídák multiple comparisons test, p < 0.0001 for all except for spiro, which was not detected). One metabolite of spiro, 6-beta, was detected in pup brains of a spiro-treated litter on the day of birth (one-tailed, unpaired t-test, t (4) = 22.94, p < 0.0001). Data are presented in box and whisker (min to max, line at median) and floating bar (min to max, line at mean) plots in Prism for serum and brain homogenate data, respectively. d Table showing mass spectral measurements of spiro and metabolites measured by LC-MS.

Article Snippet: The following stock reagents were created into standard solutions to confirm accurate mass and retention time of analytes: spiro (Cayman Chemical Company; 9000324), canrenone (“can”; Cayman Chemical Company; 21307), 7α-thiomethylspironolactone (“7-alpha”; Santa Cruz; sc-207187), and 6β-Hydroxy-7α-thiomethylspironolactone (“6-beta”; Santa Cruz; sc-210569).

Techniques: High Performance Liquid Chromatography, Mass Spectrometry, RNA In Situ Hybridization, Transformation Assay, Control, One-tailed Test, Whisker Assay, Liquid Chromatography with Mass Spectroscopy

Journal: Research

Article Title: Cortical Organoid-on-a-Chip with Physiological Hypoxia for Investigating Tanshinone IIA-Induced Neural Differentiation

doi: 10.34133/research.0273

Figure Lengend Snippet: Formation and validation of the human cortical organoid under hypoxic conditions. (A) Representative bright-field pictures of the cortical organoids generated on the chip within the hypoxic environment. Scale bars, 500 μm. (B) The immunofluorescence images of NPCs (SOX2 and NESTIN), proliferative marker Ki67, cortical layer (TBR1, SATB2, and CTIP2), differentiated neurons (TUJ1), postsynapse (αPSD95 and drebrin), Wnt signaling pathway (TCF7L2), and inhibitory neurons (GABA) within 40-d cortical organoids. Scale bars, 100 μm (in “Merge”) and 50 μm (in “Enlarged”).

Article Snippet: The primary antibodies included NESTIN (mouse, Santa Cruz Biotechnology, sc-20978, 1:400), SOX2 (rabbit, Cell Signaling Technology, 3579, 1:400), TUJ1 (mouse, BioLegend, 801201, 1:500), CTIP2 (rat, Abcam ab18465, 1:500), TBR1 (rabbit, Abcam, ab31940, 1:200), GFAP (mouse, Beyotime, AG259-1, 1:200), SATB2 (rabbit, Proteintech, 21307-1-AP, 1:500), HIF-1α (mouse, Santa Cruz Biotechnology, sc-13515, 1:200), and GABA (rabbit, Sigma, A2052, 1:400).

Techniques: Biomarker Discovery, Generated, Immunofluorescence, Marker