Journal: Nature Communications

Article Title: Optimized design and in vivo application of optogenetically functionalized Drosophila dopamine receptors

doi: 10.1038/s41467-023-43970-0

Figure Lengend Snippet: a OptoPAD setup allowing light stimulation upon feeding action. Flies expressing optoXRs in a subset of MBONs (MBONγ5β’2a,β2mp and β2mp-bilateral) related to behavioral valence receive a light stimulus (1 s 525 nm 400 μW/cm 2 ) every time they feed on the sucrose drop. b Cumulative sips over time for flies expressing optoDop1R2 V2 using MB011B-Gal4 without or with light stimulation (mean ± SEM, n = 58, 63 animals). c Total sips at 60 min for flies expressing optoDop1R2 V2 using MB011B-Gal4 without or with light stimulation ( n = 58, 63 animals, two-tailed Mann-Whitney test). d Total sips at 60 min for flies expressing optoDop1R1 V2 using MB011B-Gal4 without or with light stimulation ( n = 65, 65 animals, two-tailed Mann-Whitney test). e Total sips at 60 min for flies expressing Dop1R2 RNAi control or with MB011B-Gal4 ( n = 54, 50 animals, two-tailed Mann-Whitney test). f Total sips at 60 min for flies expressing Dop1R1 RNAi control or with MB011B-Gal4 ( n = 41, 47 animals, two-tailed Mann-Whitney test). All boxplots depict 75th (top), median (central line) and 25th (bottom) percentile, whiskers depict 99th (top) and 1st (bottom) percentile. All violin plots with single data points depict data distribution, dotted lines depict 75th (top) and 25th (bottom) percentile, solid central line the median. Source data and statistical details are provided as a Source Data file.

Article Snippet: For activation of the optoDop1R2 V2 , larvae were subjected to 10s blue light stimulation (470 nm, 720 μW/cm2, CoolLED) twice with an interval of 30s between each pulse.

Techniques: Expressing, Two Tailed Test, MANN-WHITNEY, Control

Journal: The Journal of Neuroscience

Article Title: Extrasynaptic NMDA Receptors Bidirectionally Modulate Intrinsic Excitability of Inhibitory Neurons

doi: 10.1523/JNEUROSCI.2065-21.2022

Figure Lengend Snippet: EU1794-4 mostly acts on extrasynaptic NMDARs in GABAergic neurons. A , Ambient NMDAR responses in the presence of M-8324 or vehicle. HC, Holding current with neurons clamped at 40 mV. Veh, 0.20 ± 0.046 pA/pF; N (cells) = 6. M-8324, 0.65 ± 0.13 pA/pF; N (cells) = 7. * p <0.05 (unpaired t test). B , M-8324 increased extrasynaptic NMDAR responses induced by puffed NMDA. Peak and area of puffed NMDA-induced response in the presence of M-8324 or Veh. Veh, Peak, 87 ± 6.77%; Area, 112.62 ± 8.79%; N (cells) = 5. M-8324, Peak, 77.81 ± 8.92%; Area, 112.03 ± 10.28%; N (cells) = 6. * p <0.05 (unpaired t test). C , Ambient NMDAR responses in the presence of EU1794-4 (30 μ m ) or vehicle, with sample traces on the right. Veh, 0.24 ± 0.03 pA/pF; N (cells) = 5. EU1794-4, 0.42 ± 0.05 pA/pF; N (cells) = 8. Calibration: 10 s, 50 pA. * p <0.05 (unpaired t test). D , EU1794-4's impact on evoked NMDAR-EPSCs, compared with vehicle, with sample traces on the right. Calibration: 100 ms, 20 pA. Peak amplitude, Veh, 95.22 ± 5.01%; EU1794-4, 75.71 ± 9.43%; p = 0.10. Area, Veh, 95.24 ± 4.95%; EU1794-4, 79.46 ± 8.92%; p = 0.16. N (cells) = 7 (Veh), 8 (EU1794-4). E , Activity dependence of EU1794-4 on NMDAR-EPSCs. Neither peak nor area was altered after 20 Hz synaptic stimulation in the presence of EU1794-4 (30 μ m ). Veh, Peak, 91.17 ± 5.87%; Area, 90.98 ± 13.4%; N (cells) = 5; EU1794-4, Peak, 97.84 ± 6.5%; Area, 113.7 ± 23.58%; N (cells) = 5. F , EU1794-4-Fluo increased extrasynaptic NMDAR responses. Ambient NMDAR responses were significantly larger in the presence of EU1794-4-Fluo (300 μ m ), compared with the Veh. Veh, 0.15 ± 0.05; N (cells) = 5; EU1794-4-Fluo, 0.57 ± 0.11; N (cells) = 5. * p <0.05 (unpaired t test). G , Left, Staining of EU1794-4-Fluo and synaptophysin on cultured neurons. Representative images of synaptophysin (red), EU1794-4-Fluo (green), and merge between them. Right, Percentage of (synaptophysin + EU1794-4-Fluo)/total EU1794-4-Fluo; 2.09 ± 0.47%, N (cells) = 9. ## p <0.01, compared with fluorescent density of the total EU1794-4-Fluo (paired t test). Data are mean ± SEM.

Article Snippet: PV neurons were identified by their responses to 10 pulses (50 ms duration, 50 ms interstimulus interval) of blue LED stimulation (470 nm, Thorlabs).

Techniques: Activity Assay, Staining, Cell Culture

Journal: The Journal of Neuroscience

Article Title: Extrasynaptic NMDA Receptors Bidirectionally Modulate Intrinsic Excitability of Inhibitory Neurons

doi: 10.1523/JNEUROSCI.2065-21.2022

Figure Lengend Snippet: Impact of EU1794-4 on neuronal spiking in vivo . A , Experimental setup. A mouse was head-fixed via a headpost (P) but could run freely on a rotatable plate. Sound (S) was applied to one ear, and patch recording (R) was performed in the contralateral A1. Blue light (L) and drug infusion tube (I) were positioned next to recording site in A1. B , Left, Confocal images represent tdTomato-labeled PV neurons (red) and expression of ChR2-YFP (green) in a representative brain section. Scale bar, 500 μm. Right top, Raster plot of spikes in a representative PV neuron to pulses of blue LED light stimulation (blue bars, 50 ms each pulse). Right bottom, Corresponding poststimulus spike time histogram. C , Peak/trough amplitude ratio plotted against trough-to-peak (T–P) interval of spike waveform. Each data point represented an individual neuron. Solid symbols represent mean ± SD. Inset, Spike waveforms of a representative PV neuron. Black traces were 20 superimposed spikes. Red dotted vertical lines indicate the timing of trough and peak. Blue arrows point to peak and trough. Peak/trough ratio: Pyr, 0.19 ± 0.08; PV, 0.69 ± 0.16. Trough-peak interval: Pyr, 0.81 ± 0.16; PV, 0.24 ± 0.05. Scale bar, 0.5 ms. D , Noise-evoked responses (raster plots) of PV neurons in A1 before and after EU1794-4 injection (red arrow). Dashed lines indicate the onset and offset of acoustic stimulation. Inset, Twenty randomly selected superimposed spike waveforms. Noise-evoked ( E ) and spontaneous ( F ) spike frequency of recorded PV neurons before, 15 min, and recovery after EU1794-4 infusion. N (cells) = 16. ** p < 0.01 (one-way repeated-measures ANOVA with Bonferroni test). G-I , Similar to D-F , but for pyramidal neurons. N (cells) = 18. ** p < 0.01, * p < 0.05, (one-way repeated-measures ANOVA with Bonferroni test). Data are mean ± SEM.

Article Snippet: PV neurons were identified by their responses to 10 pulses (50 ms duration, 50 ms interstimulus interval) of blue LED stimulation (470 nm, Thorlabs).

Techniques: In Vivo, Labeling, Expressing, Injection

Journal: Cell reports

Article Title: In Vitro Recapitulation of Murine Thymopoiesis from Single Hematopoietic Stem Cells

doi: 10.1016/j.celrep.2020.108320

Figure Lengend Snippet: (A–C) Different subsets in the hematopoietic hierarchy from HSCs to T cell progenitors were isolated from the bone marrow and the thymus of C57BL/6 WT mice and seeded into M-ATOs. From the bone marrow: HSC (hematopoietic stem cell) (Lin − Sca1 + cKit + CD48 CD150 + IL7R − ); MPP (multi-potent progenitor) (Lin − Sca1 + c-Kit + CD48 − CD150 − IL7R − ); LMPP (lymphoid-primed multi-potent progenitor) (Lin − Sca1 + c-Kit + IL7R + Flk2 + ); and CLP (common lymphoid progenitor) (Lin − Sca1 Lo c-Kit Lo IL7R + Flk2 + ). From the thymus: ETP (Lin − CD4 − CD8 − c-Kit hi CD44 hi CD25 − ); DN2 (Lin − CD4 − CD8 − c-Kit hi CD44 hi CD25 + ); and DN3 (Lin − CD4 − CD8 − CD44 − CD25 + ). Representative phenotypes of M-ATO-derived cells are shown at weeks 1 (A), 2 (B), and 6 (C). Data are representative of three biological replicates. (D) Frequencies of T cell populations shown as percentage of total CD45 + TCRγδ − Lin − cells initiated from the different hematopoietic subsets in week 1, week 2, and week 6 M-ATOs. Error bars denote ± SD (n = 3 independent experiments).

Article Snippet: Direct mouse Lin depletion Kit , Miltenyi Biotech , Cat# 130–110-470.

Techniques: Isolation, Derivative Assay

Journal: Cell reports

Article Title: In Vitro Recapitulation of Murine Thymopoiesis from Single Hematopoietic Stem Cells

doi: 10.1016/j.celrep.2020.108320

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Direct mouse Lin depletion Kit , Miltenyi Biotech , Cat# 130–110-470.

Techniques: Virus, Recombinant, Plasmid Preparation, Selection, Isolation, Cell Stimulation, Proliferation Assay, Activation Assay, Software

Journal: Scientific Reports

Article Title: Mesoscopic cortical network reorganization during recovery of optic nerve injury in GCaMP6s mice

doi: 10.1038/s41598-020-78491-z

Figure Lengend Snippet: Cortical responses following optic nerve injury. Upper panel for amplitude and lower panel for peak response. ( A) Cortical peak response (ΔF/F 0 × 100) in the contralateral hemisphere to the flash stimulation in the injured eye. ( B) Cortical peak response (ΔF/F 0 × 100) in the ipsilateral hemisphere to the flash stimulation in the healthy eye. ( C ) Peak latency in the contralateral hemisphere to the flash stimulation in the injured eye. ( D) Peak latency in the ipsilateral hemisphere to the flash stimulation in the healthy eye. (*) Indicates Kruskal Wallis test, which provided p values under 0.05. V1 primary visual cortex; A, AM, PM anterior, anteromedial, and posteromedial regions of the secondary visual cortex; AC anterior cingulate cortex; RS retrosplenial cortex.

Article Snippet: A 470 nm visual stimulation LED fixed on a 20 mm Star Base (Rebel LED, 65 lm, 700 mA, LUXEONStarLEDS, Quadica Developments Inc.) was placed at 5 cm from one eye.

Techniques: