Journal: bioRxiv

Article Title: C-type lectin-like receptor 2 (CLEC-2)-dependent DC migration is controlled by tetraspanin CD37

doi: 10.1101/227918

Figure Lengend Snippet: (A) WT (left), Cd37-/- (middle) or CD11c ΔCLEC”2 (right) DCs were stimulated in a 3D collagen gel with (bottom row) or without (upper row) recombinant podoplanin-Fc (rPDPN-Fc). Cells were stained for F-actin (red) and nucleus (blue) and imaged with a Leica SP5 confocal fluorescence microscope. One representative cell is shown for each condition (overview with more cells is provided in Supplementary Figure 2). Scale bar = 10μm. (B-D) Number (B) and length (C) of actin protrusions, and morphology index (D) of WT (left), Cd37-/- (middle) or CD11c ΔCLEC ” 2 (right) BMDCs upon rPDPN-Fc stimulation (grey boxes) compared to no ligand (white boxes). Data are shown as Tukey Box & whiskers from 3 independent experiments, total n=3 mice per genotype. In Tukey Box & whiskers, black dots are determined as outliers; i.e. data points outside the 25 th and 75 th percentile, minus or plus the 1.5 interquartile range, respectively. Two-way ANOVA with Tukey’s multiple comparisons, ***p<0.001, ****p<0.0001.

Article Snippet: BMDCs (0.3x10 6 ) were seeded into a 3D collagen (type I, rat tail)/matrigel matrix (both from Corning, via Thermo Fisher Scientific, Loughborough, UK) supplemented with 10% minimum essential medium alpha medium (MEMalpha, Invitrogen, via Thermo Fisher Scientific, Loughborough, UK) and 10% FCS (Greiner Bio-One, Stonehouse, UK) on glass-bottomed cell culture plates (MatTek Corporation, Bratislava, Slovakia).

Techniques: Recombinant, Staining, Fluorescence, Microscopy

Journal: bioRxiv

Article Title: A system of feed-forward cerebellar circuits that extend and diversify sensory signaling

doi: 10.1101/2023.04.11.536335

Figure Lengend Snippet: A) Representative image of UBC in whole cell patch clamp recording filled with Alexa Fluor 488. B) In GRP UBCs, mossy fiber stimulation evoked fast excitatory post synaptic currents and a slow inward current that became more prominent with faster stimulation rates. C) Summary of the decay constants of the slow EPSCs generated by a train of 10 stimuli at a 50 Hz rate in GRP UBCs. D) In current clamp, 50 Hz stimulation produced a burst of spikes that outlasted the stimulus. E) The fast EPSCs and part of the slow EPSC was blocked by an AMPA receptor antagonist (50 μM GYKI53655). The remaining slow EPSC was blocked by an mGluR1 antagonist (1 μM JNJ16259685). F) Inward currents in GRP UBCs were consistently blocked by AMPA receptor and mGluR1 receptor antagonists. G) In P079 UBCs, mossy fiber stimulation evoked slow outward currents. H) Summary of the decay constants of the slow outward currents generated by a train of 10 stimuli at a 50Hz rate in P079 UBCs. I) In current clamp, 50 Hz stimulation generated a pause in spontaneous action potential firing in P079 UBCs. J) The outward current was blocked by an mGluR2 antagonist (1 μM LY341495). K) Outward currents in P079 UBCs were entirely blocked by 1 μM LY341495. L) Peak amplitudes of the slow currents were inward in all recorded GRP UBCs and outward in all recorded P079 UBCs. M) The capacitance was significantly higher in P079 UBCs compared to GRP UBCs. N) The input resistance was not different between GRP UBCs and P079 UBCs. O) Frequency-intensity curves show that GRP UBCs are more excitable and are able to fire at faster rates than P079 UBCs. P) Example traces showing the response of GRP and P079 UBCs to 40-pA depolarizing and −100-pA hyperpolarizing 500 ms current steps. The GRP UBCs fire at a higher rate than P079 UBCs during a 40-pA depolarizing current step. The P079 UBCs have a more prominent voltage sag in response to the −100-pA hyperpolarizing current step than the GRP UBCs. Q) The sag ratio was larger in P079 UBCs than GRP UBCs. Stimulation artifacts have been removed for clarity. Error bars are SEM.

Article Snippet: Primary antibodies included chicken polyclonal anti-GFP (1:2000, Aves Labs, GFP-1020, AB_10000240), goat polyclonal anti-mCherry (1:2000, Sicgen, AB0040, AB_2333093), mouse monoclonal anti-rat mGluR1a (1:800, BD Pharmingen, 556389, AB_396404), rabbit polyclonal anti-calretinin (1:2000, Swant, 7697, AB_10000342).

Techniques: Patch Clamp, Generated, Produced

Journal: bioRxiv

Article Title: A system of feed-forward cerebellar circuits that extend and diversify sensory signaling

doi: 10.1101/2023.04.11.536335

Figure Lengend Snippet: A) Sagittal section of lobe X showing genetically expressed mCitrine (green) from the P079 mouse, tdTomato (red) from the GRP-Cre/Ai9 mouse, and immunohistochemical labeling of mGluR1 (magenta). Maximum intensity projections. B) Example of a P079 UBC (soma indicated with O, brush indicated with arrowhead), GRP UBC (soma indicated with *, brush indicated with arrowhead). The GRP UBCs expresses mGluR1 in the somatic membrane and dendritic brush. Most P079 UBCs do not express mGluR1. Single image planes. C) Sagittal section of lobe X showing genetically expressed mCitrine (green) from the P079 mouse, tdTomato (red) from the GRP-Cre/Ai9 mouse, and immunohistochemical labeling of calretinin (magenta). Maximum intensity projections. D) Example of a P079 UBC (soma indicated with O), GRP UBC (soma indicated with *). The P079 UBCs express calretinin in their cytoplasm. GRP UBCs do not express calretinin.

Article Snippet: Primary antibodies included chicken polyclonal anti-GFP (1:2000, Aves Labs, GFP-1020, AB_10000240), goat polyclonal anti-mCherry (1:2000, Sicgen, AB0040, AB_2333093), mouse monoclonal anti-rat mGluR1a (1:800, BD Pharmingen, 556389, AB_396404), rabbit polyclonal anti-calretinin (1:2000, Swant, 7697, AB_10000342).

Techniques: Immunohistochemical staining, Labeling, Membrane

Journal: bioRxiv

Article Title: A system of feed-forward cerebellar circuits that extend and diversify sensory signaling

doi: 10.1101/2023.04.11.536335

Figure Lengend Snippet: A) Coronal section of the dorsal cochlear nucleus showing genetically expressed mCitrine (green) from the P079 allele, tdTomato (red) from the GRP-Cre/Ai9 alleles, and immunohistochemical labeling of mGluR1 (magenta). Maximum intensity projections. B) Example of a P079 UBC (soma indicated with O, brush indicated with arrowhead), GRP UBC (soma indicated with *, brush indicated with arrowhead). The GRP UBCs expresses mGluR1 in the somatic membrane and dendritic brush. Most P079 UBCs do not express mGluR1, although some that are weakly labeled do appear to express mGluR1. C) Coronal section of the dorsal cochlear nucleus showing genetically expressed mCitrine (green) from the P079 allele, tdTomato (red) from the GRP-Cre/Ai9 alleles, and immunohistochemical labeling of calretinin (magenta). Maximum intensity projections. D) Example of a P079 UBC (soma indicated with O), GRP UBC (soma indicated with *). The P079 UBCs express calretinin in their cytoplasm. GRP UBCs do not express calretinin.

Article Snippet: Primary antibodies included chicken polyclonal anti-GFP (1:2000, Aves Labs, GFP-1020, AB_10000240), goat polyclonal anti-mCherry (1:2000, Sicgen, AB0040, AB_2333093), mouse monoclonal anti-rat mGluR1a (1:800, BD Pharmingen, 556389, AB_396404), rabbit polyclonal anti-calretinin (1:2000, Swant, 7697, AB_10000342).

Techniques: Immunohistochemical staining, Labeling, Membrane

Journal: bioRxiv

Article Title: A system of feed-forward cerebellar circuits that extend and diversify sensory signaling

doi: 10.1101/2023.04.11.536335

Figure Lengend Snippet: A-B) GRP ON UBC terminals (red) contacting mGluR1(+) ON UBCs (magenta). Somas indicated with *. Single image sections.

Article Snippet: Primary antibodies included chicken polyclonal anti-GFP (1:2000, Aves Labs, GFP-1020, AB_10000240), goat polyclonal anti-mCherry (1:2000, Sicgen, AB0040, AB_2333093), mouse monoclonal anti-rat mGluR1a (1:800, BD Pharmingen, 556389, AB_396404), rabbit polyclonal anti-calretinin (1:2000, Swant, 7697, AB_10000342).

Techniques: