Journal: The Journal of Comparative Neurology

Article Title: Nps ‐Expressing Neurons Receive Extensive Input From Auditory Brainstem Nuclei

doi: 10.1002/cne.70156

Figure Lengend Snippet: Cholera toxin b (CTb) injection sites in Nps ‐2A‐Cre;R26‐LSL‐L10GFP mice. Each CTb injection site is shown in a series of rostral (top row) to caudal (bottom row) levels in each of four cases: (a1–a3) 7209, (b1–b3) 7213, (c1–c3) 7216, and (d1–d3) 7220. The approximate level caudal to bregma is shown in the upper‐right corner of each panel. CTb is shown in red, L10GFP in green, and DAPI in blue. The scale bar in (a1) is 200 µm and applies to all panels.

Article Snippet: Cholera toxin b subunit (CTb) , Cholera toxin b subunit , List Biological Laboratories, goat polyclonal, Cat.# 703, Lot#: 7032A10; RRID:AB_10013220 , 1:10,000.

Techniques: Injection

Journal: Cell reports

Article Title: A spatial and projection-based transcriptomic atlas of paraventricular hypothalamic cell types

doi: 10.1016/j.celrep.2025.116904

Figure Lengend Snippet: (A) Experimental workflow for targeted single-nucleus RNA sequencing of parabrachial (PB)-projecting PVH neurons. (B) PB-projecting PVH neurons were classified with the Sim1 + sc/snRNA-seq reference atlas and projected into its UMAP space. The pie chart illustrates the proportion of PB-projecting PVH neurons that map to individual Sim1 + neuron clusters. (C) Three-level Sankey plot showing the mapping percentage of PB-projecting PVH neurons classified using the mouse Sim1 + sc/snRNA-seq reference atlas (left mapping to center; 5% cutoff) with corresponding Sim1 + MERFISH clusters identified via canonical correlation analysis (CCA; right mapping to center). Line thickness represents strength of mapping. (D) Schematic diagram of retrograde cholera toxin subunit B (CTB) injection into the PB and Cre-dependent AAV-EGFP-L10a injection into the PVH of a Mc4r -2A-Cre mouse (left). Representative coronal brain section showing the bilateral PBN CTB injection sites labeled by CTB IF. Scale bar, 250 μm (right). (E) Top panels show low-magnification (left) and high-magnification (right) views of EGFP IF, CTB IF, and Brs3 FISH labeling at ~−0.9 mm. Middle panels display Brs3 FISH, CTB IF, and EGFP IF, respectively, from left to right. Bottom panels display the overlay of Brs3 FISH with CTB IF (left), EGFP IF with CTB IF (center), and EGFP IF and Brs3 FISH (right). Yellow arrows indicate representative triple-labeled neurons. Low-magnification scale bar, 100 μm, high-magnification scale bar = 20 μm. (F) Schematic of Cre-dependent AAV-tetanus toxin light chain (TeTxLC) or AAV-EGFP injection into the PVH of Brs3 -IRES-Cre or wild-type mice (top left). Representative PVH injection site labeled with Cre-dependent AAV-EGFP-TeTxLC at ~−1.0 mm from bregma. Scale bar, 250 μm (top right). Experimental groups include wild-type (Cre-negative) mice injected with Cre-dependent AAV-TeTxLC ( n = 6), Brs3 -IRES-Cre mice injected with Cre-dependent AAV-GFP ( n = 10), and Brs3 -IRES-Cre mice injected with Cre-dependent TeTxLC ( n = 9). Body weights were monitored from the day of surgery (week 0) (lower left), and total body weight gained over 6 weeks post-surgery is depicted (lower right). Data shown as mean ± SEM. Statistical analysis was performed using one-way ANOVA followed by Tukey’s multiple comparisons test (** p < 0.01, **** p < 0.0001). (G) Schematic and representative trace from Arc Npy/Agrp → PVH Brs3 neuron channelrhodopsin-2 (ChR2)-assisted circuit mapping (CRACM) experiment. Light-evoked IPSCs were detected in 8/14 PVH Brs3 neurons. (H) Schematic showing injection of Cre-dependent ChR2 or mCherry into the PVH and optic fiber implants over the PB in Brs3 -IRES-Cre mice (left), with representative brain sections showing bilateral ChR2 expression in the PVH at ~−0.9 mm from bregma (left-center; scale bar, 100 μm), and bilateral fiber tracks in the PB (right-center; scale bar, 250 μm). Food intake (right) was measured over the first 3-h of the dark cycle, with or without photostimulation, in mCherry- ( n = 4) and ChR2-expressing mice ( n = 8). Data shown as mean ± SEM. Statistical analysis was performed using two-way repeated measures ANOVA followed by Sidak’s multiple comparisons test (** p < 0.01).

Article Snippet: Goat polyclonal anti-cholera toxin subunit B , List Biological Laboratories , Cat#: 703; RRID:AB_10013220.

Techniques: RNA Sequencing, Injection, Labeling, Expressing

Journal: ASN NEURO

Article Title: GABA Receptor Activation in Müller Glia as a Molecular Switch for Controlling VEGF-A in the Retina

doi: 10.1080/17590914.2026.2618997

Figure Lengend Snippet: GABA A R blockade inhibits GABA-induced VEGF-A modulation in MGC. (A) Immunofluorescence images of MGC treated for 48 hours with 100 µM GABA alone or co-treated with 10 µM gabazine (GABA A R antagonist) or 5 µM CGP55845 (GABA B R antagonist), stained for VEGF-A (red). Nuclei were counterstained with DAPI (blue). Insets (white boxes) show 2x digital zoom. Scale bars: 200 µm (main image) and 100 µm (insets). (B) Mean VEGF-A fluorescence intensity for each condition. (C) Histogram showing the percentage distribution of VEGF-A fluorescence intensity in MGC cultures exposed to the treatments in A. (D) ELISA measurements of VEGF-A release in media from MGC treated for 48 hours with 100 µM GABA alone or co-treated with 10 µM gabazine (GABA A R antagonist), 5 µM CGP55845 (GABA B R antagonist), or 10 µM TPMPA (GABA rho R antagonist). One-way ANOVA with Tukey’s post hoc: * p < 0.05, ** p < 0.01, **** p < 0.0001 versus control.

Article Snippet: Cells were then incubated overnight at 4 °C with primary antibodies against CRALBP (1:100, sc-59487, Santa Cruz Biotechnology, USA), Sox-2 (1:100, sc-365964, Santa Cruz Biotechnology, USA), Nestin (1:250, ab221660, Abcam, USA), GABA A R .1-6 subunits (1:100, sc-376282, Santa Cruz Biotechnology, USA), GABA B R B1 subunit (1:100, sc-398901, Santa Cruz Biotechnology, USA), VEGF-A (1:250, sc-7269, Santa Cruz Biotech, USA) and HIF-1α (1:500, 36169, Cell Signaling).

Techniques: Immunofluorescence, Staining, Fluorescence, Enzyme-linked Immunosorbent Assay, Control

Journal: ASN NEURO

Article Title: GABA Receptor Activation in Müller Glia as a Molecular Switch for Controlling VEGF-A in the Retina

doi: 10.1080/17590914.2026.2618997

Figure Lengend Snippet: Selective GABA A R activation regulates VEGF-A in MGC. (A) Representative immunofluorescence images of MGC treated for 48 hours with 100 µM muscimol (GABA A R agonist) or 100 µM baclofen (GABA B R agonist), with or without gabazine at 10 µM or 5 µM CGP55845 (GABA A R and GABA B R antagonists, respectively), immunostained against VEGF-A (red). Nuclei were counterstained with DAPI (blue). Insets (white boxes) show 2× digital zoom. Scale bar: 200 µm (main panel) and 100 µm (insets). (B) Mean VEGF-A fluorescent intensity. (C) Percent distribution histogram of single-cell VEGF-A fluorescence intensities. (D) Western blot and quantification of cellular VEGF-A in MGC treated with 100 µM muscimol for 48 hours. E. Quantification of Vegfa mRNA in MGC treated with 100 µM muscimol for 48 hours, normalized to Gapdh ( n = 4 independent cultures). F. ELISA quantification of secreted VEGF-A in conditioned media from MGC treated with 100 µM muscimol or 100 µM baclofen, with or without gabazine at 10 or 5 µM CGP55845 for 48 hours. Data were analyzed using Student’s t-test and one-way ANOVA with Tukey’s post hoc test: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, n.s.: not significant versus control.

Article Snippet: Cells were then incubated overnight at 4 °C with primary antibodies against CRALBP (1:100, sc-59487, Santa Cruz Biotechnology, USA), Sox-2 (1:100, sc-365964, Santa Cruz Biotechnology, USA), Nestin (1:250, ab221660, Abcam, USA), GABA A R .1-6 subunits (1:100, sc-376282, Santa Cruz Biotechnology, USA), GABA B R B1 subunit (1:100, sc-398901, Santa Cruz Biotechnology, USA), VEGF-A (1:250, sc-7269, Santa Cruz Biotech, USA) and HIF-1α (1:500, 36169, Cell Signaling).

Techniques: Activation Assay, Immunofluorescence, Fluorescence, Western Blot, Enzyme-linked Immunosorbent Assay, Control