Journal: The Journal of Neuroscience

Article Title: Interdependent Conductances Drive Infraslow Intrinsic Rhythmogenesis in a Subset of Accessory Olfactory Bulb Projection Neurons

doi: 10.1523/JNEUROSCI.2520-15.2016

Figure Lengend Snippet: Expression of BK and CaV2.3 channels in AOB mitral cells. A–C, Confocal laser-scanning microscopy images of the mouse AOB (parasagittal cryosections). Immunofluorescence labeling (green) of CaV2.3 (A), the BK channel subunit α-1 (Sloα1; B), and the BK channel subunit β-4 (Sloβ4; C). Nuclei are counterstained with DRAQ5 (red). Dashed lines indicate both the lateral olfactory tract (LOT) and the glomerular layer (GL) in Aiii, Biii, and Ciii. Representative single optical section images show an overview of the AOB (Ai, Bi, Ci) and a high-magnification view (Aii, Bii, Cii) of the mitral cell layer (MCL). Note the robust labeling of cells in the MCL as well as the absence of immunofluorescence following peptide preadsorption (Aiii, Biii, Ciii; no DRAQ5 counterstaining). D, E, Post hoc immunolabeling of previously recorded intrinsically rhythmic neurons. Confocal images of AOB cryosections containing biocytin-filled iAMC somata and proximal dendrites (visualized by Alexa Fluor 488/633 streptavidin conjugate). Sections were counterstained against the BK channel α-1 (D) and β-4 (E) subunits, respectively. Merged images (Diii, Eiii) reveal BK expression in these representative iAMCs. Div, Eiv, Western blot analysis of total olfactory bulb (right lanes) and posterior brain (left lanes) protein extracts. Immunoblots were probed with anti-Sloα1 (D) and anti-Sloβ4 (E) antibodies, respectively. Expected band size is indicated above both blots. d, Dorsal; FC, frontal cortex; v, ventral.

Article Snippet: Primary antibodies were anti-Slo1 (K Ca 1.1, 1:500; Alomone Labs), anti-Sloβ4 (1:100; Alomone Labs), and anti-Ca V 2.3 (1:100; Alomone Labs).

Techniques: Expressing, Confocal Laser Scanning Microscopy, Immunofluorescence, Labeling, Immunolabeling, Western Blot

Journal: Nature Communications

Article Title: Priming mobilization of hair follicle stem cells triggers permanent loss of regeneration after alkylating chemotherapy

doi: 10.1038/s41467-019-11665-0

Figure Lengend Snippet: Bu-induced proliferation followed by Cy-induced apoptosis in organ-cultured HFs. a Schedule for Bu/– and Bu/Cy treatments and subsequent analysis at 2, 4, and 6 days after alkylating chemotherapy. b Hair shaft elongation and c , hair cycle score of organ-cultured human HFs after alkylating chemotherapy ( n = 38 biological replicates/timepoint, 3 independent experiments). Note that the score of the Bu/Cy group was lower than that of the control group on day 6. d Representative images of organ-cultured HFs of experimental groups. Control HFs maintained anagen morphology, and Bu-treated HFs prematurely entered catagen stage (black arrow); however, Bu/Cy-treated HFs showed completely shrunken bulb morphology (black arrowhead). Representative images and quantification of e , Ki67 + cells among K15 + HFSCs ( n = 5 biological replicates/group), f p53 + cells among integrin β4 + basal cells ( n = 6 biological replicates/group), and g , p53 + c-caspase-3 + cells ( n = 7 biological replicates/group) in the bulge after alkylating chemotherapy. HFSCs showed remarkable cellular proliferation after Bu treatment (white arrowhead) and were completely quenched with large-scale apoptosis after Bu/Cy treatment (white arrow; immunofluorescence; scale bar = 100 μm). Bu busulfan, Cy cyclophosphamide, Bu/Cy busulfan followed by cyclophosphamide, HF hair follicle, HPF high-power field, HFSC hair follicle stem cell, c-caspase-3 cleaved caspase-3. Data are mean ± SEM. Source data are provided as a Source Data file. * p < 0.05 (vs. Con); *** p < 0.001 (vs. Con, two-way analysis of variance with Dunnett’s multiple comparisons test) in b and c ; *** p < 0.001 (vs. Con, unpaired t test) in e , f , and g

Article Snippet: For immunofluorescence, frozen tissues were sectioned at a thickness of 4 μm and incubated overnight at 4 °C with the following primary antibodies diluted in antibody diluent reagent (Invitrogen): Fas (ab82419, 1:100; Abcam, MA, USA), Ki67 (SP6, 1:200; Spring Bioscience, Pleasanton, CA, USA or MIB-1, 1:200; Dako), K15 (LHK15, 1:200; Thermo Fisher), p53 (DO-7, 1:800; Novocastra, Newcastle, England or 1C12, 1:1000; Cell Signaling, Danvers, MA, USA), K14 (GTX104124, 1:1000; GeneTex, Irvine, CA, USA), cleaved caspase-3 (5A1E, 1:400; Cell Signaling), γ-H2AX (#2577, 1:800; Cell Signaling), Shh (EP1190Y, 1:500; Abcam), Lhx2 (C-20, 1:200; Santa Cruz Biotechnology), integrin β4 (H-101, 1:200; Santa Cruz Biotechnology), phospho Akt (#9275, 1:800; Cell Signaling), phospho p38 (#9211, 1:800; Cell Signaling), TYR (T311, 1:100; Thermo Fisher, Waltham, MA, USA), TRP1 (SC25543, 1:100; Santa Cruz Biotechnology), and MITF (C5, 1:100; Thermo Fisher).

Techniques: Cell Culture, Control, Immunofluorescence