Journal: The Journal of Neuroscience

Article Title: Sparser and Less Efficient Hippocampal-Prefrontal Projections account for Developmental Network Dysfunction in a Model of Psychiatric Risk Mediated by Gene-Environment Interaction

doi: 10.1523/JNEUROSCI.1203-21.2021

Figure Lengend Snippet: Organization and developmental dynamics of hippocampal innervation of PL in neonatal and prejuvenile dual-hit GE mice. A , Schematic representation of tracing protocols. Mice were injected unilaterally with the retrograde tracer FG into PL or with either the anterograde tracer BDA or the transsynaptic tracer WGA into HP. B , left, Fluorescent images of FG (white) injection site in the PL of 50-µm-thick coronal slice from a P10 mouse when costained with NeuN (green). Right, Corresponding NeuN-stained coronal slices (50-µm-thick) including the dHP, iHP, and vHP with retrogradely stained cells (white). Scale bar: 500 μm. Insets, FG-stained neurons (white) shown at higher magnification. Scale bar: 100 μm. C , Photograph of a representative BDA injection into the i/vHP of a P10 mouse visualized by streptavidin staining (green) and costained with DAPI (blue) in a 50-µm-thick coronal slice. D , 3D reconstruction of hippocampal axons (tDimer, red) into the PL (cyan surface) in a cleared P10 mouse brain. PL volume was delimited according to nuclei staining (DRAQ5; data not shown). Inset, logarithmic violin plots depicting the relative space occupancy of hippocampal fibers within PL, normalized to the number of transfected neurons in i/vHP. E , left, Orthogonal views of the Z-stack (YZ, XZ) images illustrating BDA-positive boutons (green) that colocalized with vGLUT1 (red). Right, Confocal images displaying BDA-positive boutons (green) on GABA-positive (red) somata (arrows) and GABA-negative neurons (asterisk). F , left, Representative example of WGA staining (magenta) in a 50-µm-thick coronal slice including the PFC of a P10 mouse that transsynaptically labeled neurons targeted by hippocampal axons. Dotted lines mark the borders of the two subdivisions as well the PL layers. Right, Photographs displaying the colocalization of WGA (magenta) and GABA (cyan) staining for GABA-positive neurons (circle). GABA-negative but WGA-positive neurons are marked by squares. Gi , Photograph of BDA-labeled hippocampal terminals (black) targeting deep layers of PL from a P10 CON (left) and a P10 GE (right) mouse, respectively. ii , Schematic illustrating the extent of the BDA injections into the i/vHP of neonatal CON (gray) and dual-hit GE (green) mice. iii , Violin plots of the normalized density of hippocampal terminals (in µm/mm 2 /cell) in layer 2/3 and layer 5/6 of PL averaged for all investigated neonatal CON and GE mice. H , Same as G , for prejuvenile mice. I , Photograph of FG-labeled neurons (white) in the CA1 area of i/vHP in a 50-µm-thick NeuN-stained (green) coronal slice from a P10 CON and a P10 GE mouse, respectively. J , Violin plots of the normalized density of PL-projecting neurons in the hippocampal CA1 area averaged for all investigated neonatal (left) and prejuvenile (right) CON and GE mice. PL: prelimbic cortex, IL: infralimbic cortex, Cg: cingulate cortex, SO: stratum oriens, SP: stratum pyramidale, SR: stratum radiatum. Single data points are represented as dots and the red horizontal bars in violin plots correspond to the median and the 25th and 75th percentiles; * p < 0.05.

Article Snippet: Subsequently, slices were incubated overnight with mouse monoclonal Alexa Fluor 488-conjugated antibody against NeuN (1:100, MAB377X; Merck Millipore), Alexa Fluor 488-conjugated streptavidin (1:1000, Merck Millipore), or rabbit polyclonal primary antibody against GABA (1:1000, A2052; Sigma-Aldrich), polyclonal guinea-pig antibody against VGLUT1 (1:1000, Synaptic Systems), rabbit polyclonal primary antibody against lectin (1:1000, A2052; Sigma-Aldrich) followed by 2-h incubation with Alexa Fluor 568 goat anti-rabbit IgG secondary antibody (1:500, A11008; Merck Millipore), Alexa Fluor 568 goat anti-guinea pig (1:500, Invitrogen), Alexa Fluor 568 donkey anti-rabbit (1:500, Life Technologies) and Alexa Fluor 568-conjugated streptavidin (1:1000, Merck Millipore).

Techniques: Injection, Staining, Transfection, Labeling

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: A pathogenic dyskerin mutation impairs proliferation and activates a DNA damage response independent of telomere length in mice

doi: 10.1073/pnas.0803559105

Figure Lengend Snippet: Localization of γ-H2AX foci to telomeres in Dkc1Δ15 (Δ15) MEFs. (A) Colocalization of γ-H2AX foci (Alexa Fluor 568, red) and telomeres by telomere PNA-FISH (FITC, green) in WT and Dkc1Δ15 MEFs after treatment with 5 μM etoposide for 30 min. DNA was counterstained with DAPI (blue). Magnified views of merged images showed details of the colocalization. (B) Quantitation of colocalization of signals from (A) of γ-H2AX foci and telomeres. More than 100 cells were analyzed.

Article Snippet: FITC, Texas red, Alexa Fluor 568, and DAPI images were overlapped by using ISIS FISH imaging software (Metasystems).

Techniques: Quantitation Assay