Journal: Cell reports

Article Title: Inappropriate Intrusion of an Axonal Mitochondrial Anchor into Dendrites Causes Neurodegeneration

doi: 10.1016/j.celrep.2019.09.012

Figure Lengend Snippet: (A–F) Immunohistochemistry technique for capturing SNPH intrusion in PC dendrites. Shown is SNPH (green), Syt2 (red), and Calbindin (blue) labeling in 3.5-month-old WT (A, C, and E) and Shi (B, D, and F) mice. Scale bar, 10 μm. (G and H) High magnification of the maximum intensity projection image from the z stack through dendritic regions of WT (G) and Shi (H). (I and J) Orthogonal (slice) view of SNPH punctum (indicated by arrows in panels G and H) in the dendritic region from WT (I) and Shi (J) in x-z and y-z orientations, respectively. (K) Quantification of percentage area occupied by SNPH within the dendritic volume from 3 mice of each group. Data are shown as mean ± SEM. *p < 0.05. (L–N) Capturing SNPH intrusion by pre-tagging dendritic mitochondria in vivo using viral transduction. (L) Technique to selectively transduce PCs with AAV-Mito-mCherry. (M) Demonstration of successful pre-tagging of mitochondria in dendrites of a single PC by Calbindin staining. Scale bar, 10 μm. (N) Demonstration of how pre-tagged dendritic mitochondria in PCs are used to capture SNPH intrusion by 3D rotation. (O and P) Single dendritic tree in WT (O) or Shi (P) pre-tagged with Mito-mCherry (red) and SNPH intrusion (green) captured by co-rotation with Mito-mCherry. Merged images show the fraction of dendritic mitochondria anchored by intruded SNPH (yellow). (Q and R) Percentage of PCs with SNPH intrusions (Q) and percentage of SNPH bound to mitochondria per dendritic tree (R) from WT (n = 73) and Shi (n = 106) PCs. Data are shown as mean ± SEM. ***p < 0.005.

Article Snippet: Polyclonal Calbindin , Boster , M03047–2.

Techniques: Immunohistochemistry, Labeling, In Vivo, Transduction, Staining

Journal: Cell reports

Article Title: Inappropriate Intrusion of an Axonal Mitochondrial Anchor into Dendrites Causes Neurodegeneration

doi: 10.1016/j.celrep.2019.09.012

Figure Lengend Snippet: (A and B) Representative images of lentivirally transduced GFP-SNPH (1–469) (A) and GFP-SNPH (B) in PCs of SNPH-KO mice injected with saline (no harmaline, vehicle only). (C-H) Effect of harmaline on GFP-SNPH (1–469)-transduced (C) and GFP-SNPH-transduced (F) PC dendrites. Degenerating dendrites in GFP-SNPH-transduced PCs can be seen in (F). Also shown is Calbindin labeling of GFP SNPH (1–469) (D) and GFP-SNPH (G) from (C) and (F). Merged images of GFP SNPH (1–469) and GFP-SNPH with Calbindin are shown in (E) and (H), respectively. (I–K) Representative image of a harmaline-induced degenerating PC (white arrow in I) transduced with GFP-SNPH. Calbindin staining from the same section is shown in (J), whereas a merged image is shown in (K). Scale bars, 20 μm. (L) Quantification of dendritic shrinkage in GFP-SNPH (1–469)- and GFP-SNPH-transduced PCs in the absence (n = 3 mice, vehicle only) or presence of harmaline (n = 5 mice). Data are shown as mean ± SEM. ***p < 0.001.

Article Snippet: Polyclonal Calbindin , Boster , M03047–2.

Techniques: Injection, Saline, Labeling, Transduction, Staining

Journal: Cell reports

Article Title: Inappropriate Intrusion of an Axonal Mitochondrial Anchor into Dendrites Causes Neurodegeneration

doi: 10.1016/j.celrep.2019.09.012

Figure Lengend Snippet:

Article Snippet: Polyclonal Calbindin , Boster , M03047–2.

Techniques: Virus, Plasmid Preparation, Recombinant, Software, Imaging

Journal: iScience

Article Title: Dual mechanisms of supporting cell regeneration in the neonatal mouse cochlea

doi: 10.1016/j.isci.2026.115113

Figure Lengend Snippet: ScRNA-seq reveals damage-induced transcriptomes of the greater epithelial ridge and inner phalangeal cells (A) UMAP showing integration of Epcam + cochlear epithelial cells from P4-P7 control ( Ki67 CreERT2/+ ; R26R tdTomato/+ ) and damage ( Ki67 CreERT2/+ ; Lgr5 DTR/+ ; R26R tdTomato/+ ) cochlea, resulting in 18 individual clusters. (B) UMAPs showing gene expression profiles of medial and lateral GER markers. (C) UMAPs showing cell clusters integrated from control and damage cochlea at individual time points. Arrows indicate cell clusters predominantly or exclusively found in the damaged cochlea. (D) Percent distribution of cells from each time point and condition by cluster. To account for differences in the number of profiled cells in each dataset, percent contribution to each cluster was normalized to the dataset size. Blue asterisk denotes proliferating GER cells, red asterisks denote native and responding DC/IPC populations. Note that cluster 10 (native DC/IPCs) are mainly composed of cells from control cochlea, whereas cluster 8 (responding DC/IPC) is entirely composed of cells from damaged cochlea. (E) UMAPs showing a reduction of Lgr5 + cells in the damaged cochlea, as well as an increase in Ki67-tdTomato + cells in the damaged lateral GER and Fabp3 + responding DC/IPCs. Cluster 17 (proliferating cells) express Mki67 . (F) Overlaid expression of “cell cycle” and “pathways of neurodegeneration—multiple diseases” gene signatures, extracted from KEGG Pathways mmu04110 and mmu05022 using clusterProfiler. Cell cycle gene signature is highly enriched in cluster 17 and moderately enriched in cluster 8. See also . (G) Volcano plots showing differentially expressed genes between damage and control GER and IPhC clusters at all four time points (excluding proliferating cell cluster). (H) Immunostaining showing that Galectin1 and Calb1 mark medial GER (white brackets) in all three turns of the control cochlea (P6-7). After damage, domains expressing Galectin1 and Calb1 expand laterally into the central GER. Note that Calb1 also labels hair cells. (I) Crabp1 is expressed in medial GER in all three turns of P4 and P7 control cochlea. At P4 after damage, its expression robustly increased and expanded to the central and lateral GER, before returning to similar levels and spatial pattern as controls at P7. (J) Immunostaining shows that Jag1 expression initially decreases in the lateral GER relative to controls at P4, before upregulation at P7 to more intense levels than controls. (K) Quantification of Crabp1-expressing regions shows a significant increase after damage relative to controls at P7 ( n = 3). Unpaired t tests were used for (K). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. Brackets represent GER. IHC = inner hair cell (arrows), OHC = outer hair cell, GER = greater epithelial ridge. Scale bars represent 50 μm.

Article Snippet: Rabbit anti-calbindin (Calb1) , Cell Signaling Technologies , Cat#13176: RRID: AB_2687400.

Techniques: Control, Gene Expression, Expressing, Immunostaining