Journal: Alcohol (Fayetteville, N.Y.)

Article Title: Alcohol consumption does not impact delta and kappa opioid receptor-mediated synaptic depression in dorsolateral striatum of adult male mice

doi: 10.1016/j.alcohol.2024.06.002

Figure Lengend Snippet: A) Adult male A2aCre-TdTomato and D1Cre-TdTomato mice underwent 3 weeks of water or alcohol (20% ethanol) drinking-in-the-dark (DID). Mice drank for 2 h/day on Monday through Thursday and for 4 h on Friday. Mice had access to normal water bottles in cages at all other times. B) Daily water consumption across the 15 sessions of water DID. C) Total water consumption each of the 3 weeks of water DID. There were no significant differences in water consumption between the two genotypes. D) Daily alcohol consumption across the 15 sessions of alcohol DID. There was a significant effect of genotype (F (1, 9) = 13.66; P=0.0049) and time (F (14, 126) = 13.62; P<0.0001) and a significant time x genotype interaction (F (14, 126) = 3.062, P=0.0004). Post-hoc analyses revealed that D1Cre-TdTomato mice drank more alcohol than A2aCre-TdTomato mice only on the fifth session of DID (P<0.0001). E) Total alcohol consumption each of the 3 weeks of alcohol DID. There was a significant effect of genotype (F (1, 9) = 13.66; P=0.0049) and a significant time x genotype interaction (F (2, 18) = 3.848, P=0.0406). Post-hoc analyses revealed that D1Cre-TdTomato mice drank more alcohol the first week of DID than A2aCre-TdTomato mice (P=0.0312). For D1Cre-TdTomato mice: water n=6 mice, ethanol n=6 mice. For A2aCre-TdTomato mice: water n=4 mice, ethanol n=5 mice.

Article Snippet: Mice were generated via breeding transgenic male TdTomato reporter mice (Ai14 mice, Jackson Laboratory strain # 007914) with transgenic female A2aCre or D1Cre mice.

Techniques:

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: Alcohol consumption does not impact delta and kappa opioid receptor-mediated synaptic depression in dorsolateral striatum of adult male mice

doi: 10.1016/j.alcohol.2024.06.002

Figure Lengend Snippet: A) Representative brain slice image and time course of kappa opioid receptor-mediated LTD as induced by the kappa agonist U69,593 (300 nM, 5 min bath application). Representative traces of electrically-evoked excitatory postsynaptic current amplitudes (eEPSCs) illustrate baseline currents and currents following agonist application (at right in ( A ). B) U69593 produced a significant reduction in eEPSC amplitudes in water drinking mice (t=5.493, df=6, P=0.0015). C) U69593 produced a significant reduction in eEPSC amplitudes in alcohol drinking mice (t=4.875, df=7, P=0.0018). D) There was not a significant difference in the degree of LTD produced by U69,593 between water and alcohol drinkers (t=1.361, df=13, P=0.1967). Water=7 recordings from 4 A2aCre-TdTomato mice; Alcohol=8 recordings from 5 A2aCre-TdTomato mice. E) Time course of delta opioid receptor-mediated LTD as induced by the delta agonist DPDPE (300 nM, 5 min bath application). Representative traces of electrically-evoked excitatory postsynaptic current amplitudes (eEPSCs) illustrate baseline currents and currents following agonist application (at right in ( E ). F) DPDE produced a significant reduction in eEPSC amplitudes in water drinking mice (t=6.006, df=6, P=0.0010). G) DPDPE produced a significant reduction in eEPSC amplitudes in alcohol drinking mice (t=3.581, df=8, P=0.0072). ( H) There was not a significant difference in the degree of LTD produced by DPDPE between water and alcohol drinkers (t=2.075, df=14, P=0.0568). Water=7 recordings from 4 A2aCre-TdTomato mice; Alcohol=9 recordings from 5 A2aCre-TdTomato mice. Summary data in A and E represent mean ± SEM. Summary data in D and H illustrates the minimum to the maximum with the box representing the median and the interquartile ranges. Scale bars in A and E are 200 pA and 50 ms.

Article Snippet: Mice were generated via breeding transgenic male TdTomato reporter mice (Ai14 mice, Jackson Laboratory strain # 007914) with transgenic female A2aCre or D1Cre mice.

Techniques: Slice Preparation, Produced

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: Alcohol consumption does not impact delta and kappa opioid receptor-mediated synaptic depression in dorsolateral striatum of adult male mice

doi: 10.1016/j.alcohol.2024.06.002

Figure Lengend Snippet: A) Representative brain slice image and time course of kappa opioid receptor-mediated LTD as induced by the kappa agonist U69,593 (300 nM, 5 min bath application). Representative traces of electrically-evoked excitatory postsynaptic current amplitudes (eEPSCs) illustrate baseline currents and currents following agonist application (at right in ( A ). B) U69593 produced a significant reduction in eEPSC amplitudes in water drinking mice (t=6.931, df=7, P=0.0002). C) U69593 produced a significant reduction in eEPSC amplitudes in alcohol drinking mice (t=4.727, df=8, P=0.0015). D) There was not a significant difference in the degree of LTD produced by U69,593 between water and alcohol drinkers (t=1.054, df=15, P=0.3086. Water=9 recordings from 4 D1-TdTomato mice; Alcohol=8 recordings from 6 D1Cre-TdTomato mice. E) Time course of delta opioid receptor-mediated LTD as induced by the delta agonist DPDPE (300 nM, 5 min bath application). Representative traces of electrically-evoked excitatory postsynaptic current amplitudes (eEPSCs) illustrate baseline currents and currents following agonist application (at right in ( E ). F) DPDE produced a significant reduction in eEPSC amplitudes in water drinking mice (t=5.344, df=7, P=0.0011). G) DPDPE produced a significant reduction in eEPSC amplitudes in alcohol drinking mice (t t=3.182, df=8, P=0.0130). ( H) There was not a significant difference in the degree of LTD produced by DPDPE between water and alcohol drinkers (t=1.014, df=15, P=0.3268). Water=8 recordings from 4 D1Cre-TdTomato mice; Alcohol=9 recordings from 6 D1Cre-TdTomato mice. Summary data in A and E represent mean ± SEM. Summary data in D and H illustrates the minimum to the maximum with the box representing the median and the interquartile ranges. Scale bars in A and E are 200 pA and 50 ms.

Article Snippet: Mice were generated via breeding transgenic male TdTomato reporter mice (Ai14 mice, Jackson Laboratory strain # 007914) with transgenic female A2aCre or D1Cre mice.

Techniques: Slice Preparation, Produced

Journal: Journal of Extracellular Vesicles

Article Title: Targeted Intracellular Delivery via Precision Programming of ARRDC1‐Mediated Microvesicles

doi: 10.1002/jev2.70199

Figure Lengend Snippet: Targeted delivery of ARMMs to CD8 + T cells in vivo. (A) Schematic of mCD8DARPin‐decorated ARMMs carrying an ARRDC1‐GFP fusion protein as cargo. (B) Western blot analysis showing incorporation of NiV‐G‐mCD8DARPin into ARMMs. Lysates from producer cells and EVs were subjected to Western blot analysis using indicated antibodies. (C) Targeted delivery of GFP protein to mouse CD8⁺ splenocytes. Mouse splenocytes were incubated for 24 h with NT‐ARMM(GFP) or mCD8‐ARMM(GFP) at a vesicle‐to‐cell ratio of 2 × 10⁵. Cells were stained with anti‐mCD8 antibody and analysed by flow cytometry. Representative plots from three independent experiments are shown. (D) Quantification of GFP⁺ CD8⁺ cells from flow cytometry data shown in (C). ** p < 0.01 . n.s.: not significant. (E) Schematic of mCD8DARPin‐ARMMs carrying ARRDC1‐Cre fusion protein. Lower panel depicts the Ai14 reporter system, where Cre‐mediated excision of a floxed stop cassette induces tdTomato expression. (F) Flow cytometry analysis of tdTomato + cells from mouse spleen. Ai14 mice were injected retro‐orbitally with NT‐ARMM(Cre) or mCD8‐ARMM(Cre) at 1 × 10 11 vesicles per mouse. After 7 days, splenic T cells were isolated, stained with APC‐conjugated anti‐mCD8 antibody, and analysed by flow cytometry. Representative plots from three independent experiments are shown. (G) Quantification of tdTomato⁺ CD8⁺ splenocytes from (F). Data are shown as mean ± SEM ( n = 3 mice per group). * p < 0.05; n.s.: not significant .

Article Snippet: To evaluate in vivo targeting of CD8+ cells, 8‐week‐old female Ai14 mice (Jackson Labs) received retro‐orbital injections of mCD8DARPin‐ARMMs (Cre) (6 × 10 10 EVs in 150 μL PBS).

Techniques: In Vivo, Western Blot, Incubation, Staining, Flow Cytometry, Expressing, Injection, Isolation

Journal: Journal of Extracellular Vesicles

Article Title: Targeted Intracellular Delivery via Precision Programming of ARRDC1‐Mediated Microvesicles

doi: 10.1002/jev2.70199

Figure Lengend Snippet: Targeted in vivo delivery of ARMMs to GluA4 + interneurons. (A) Schematic of GluA4DARPin‐decorated ARMMs carrying Cre recombinase as cargo. (B) Western blot analysis showing incorporation of NiV‐G‐GluA4DARPin and Cre protein into ARMMs. HEK293T cells were co‐transfected with NiV‐G‐GluA4DARPin, NiV‐F, and either ARRDC1‐GFP or ARRDC1‐Cre. EVs were isolated 72 h post‐transfection by ultracentrifugation. Cell and EV lysates were analysed by Western blotting. (C) Schematic showing stereotaxic injection of ARMMs into mouse brain. (D) Representative motor cortex sections stained with anti‐parvalbumin (PV) antibody and 4′,6‐diamidino‐2‐phenylindole (DAPI). Ai14 reporter mice were stereotaxically injected with NT‐ARMMs (Cre), GluA4‐ARMMs (Cre), or VSVG‐ARMMs (Cre) at 2 × 10 11 vesicles per mouse. Three weeks post‐injection, brains were harvested for immunostaining and imaging to assess tdTomato expression. Images show separate and merged channels from mice injected with each ARMM variant. Arrows indicate PV⁺/tdTomato⁺ double‐positive cells. Scale bars: 25 µm. (E) Quantification of tdTomato⁺ cells within PV⁺ or PV − neuronal populations based on multiple images, including the one shown in (D). Data are shown as mean ± SEM ( n = 3 mice per group).* p < 0.05, ** p < 0.01.

Article Snippet: To evaluate in vivo targeting of CD8+ cells, 8‐week‐old female Ai14 mice (Jackson Labs) received retro‐orbital injections of mCD8DARPin‐ARMMs (Cre) (6 × 10 10 EVs in 150 μL PBS).

Techniques: In Vivo, Western Blot, Transfection, Isolation, Injection, Staining, Immunostaining, Imaging, Expressing, Variant Assay

Journal: bioRxiv

Article Title: Direct piriform-to-auditory cortical projections shape auditory-olfactory integration

doi: 10.1101/2024.07.11.602976

Figure Lengend Snippet: Injection of AAVrg-hSyn-eGFP into the left auditory cortex (ACx) of Cdh23 mice. B) Representative fluorescent micrographs of ACx injection site (left), posterior piriform (pPir) ROI (middle), anterior piriform (aPir) ROI (right). Scale bars = 500 µm. C) Number of cells labeled with GFP in aPir and pPir (n=7 mice; n.s. p=0.375, Wilcoxon signed-rank test). D) Injection of AAVrg-hSyn-Cre into the left ACx of Ai14 mice. E) Representative fluorescent micrographs of ACx injection site (left), pPir ROI (middle), aPir ROI (right). Scale bars = 500 µm. F) Number of cells labeled with tdTomato in aPir and pPir (n=7 mice; *p=0.020, paired t-test).

Article Snippet: To label ACx-projecting neurons in Pir in Ai14 mice, we injected AAVrg-hSyn-Cre-WPRE-hGH (Addgene #105553-AAVrg; RRID:Addgene_105553; 150-250 nL) into left ACx.

Techniques: Injection, Labeling

Journal: bioRxiv

Article Title: Direct piriform-to-auditory cortical projections shape auditory-olfactory integration

doi: 10.1101/2024.07.11.602976

Figure Lengend Snippet: Injection of AAV1-hSyn-Cre into the left piriform cortex (Pir) of Ai14 mice. B) Representative fluorescent micrographs of the Pir injection site (left) and auditory cortical ROIs (right). Scale bars = 500 µm. C) Number of cells labeled with tdTomato across auditory cortical regions (n=6 mice; *p=0.0141, Friedman test). D) Histograms of the depth distribution of labeled cells in each auditory cortical region (all cells from 6 mice).

Article Snippet: To label ACx-projecting neurons in Pir in Ai14 mice, we injected AAVrg-hSyn-Cre-WPRE-hGH (Addgene #105553-AAVrg; RRID:Addgene_105553; 150-250 nL) into left ACx.

Techniques: Injection, Labeling

Journal: Neuroscience Bulletin

Article Title: A Novel Long-range Excitatory Neural Circuit from the Magnocellular Red Nucleus to Spinal Dorsal Horn Facilitates Neuropathic Pain-like Behaviors in Male Mice

doi: 10.1007/s12264-025-01553-7

Figure Lengend Snippet: RMC CaMKIIα neurons were activated in male mice with neuropathic pain after common peroneal nerve (CPN) ligation. A Schematic of the CPN model. B Image showing c-Fos (green) co-labeled with CaMKIIα (red) in RMC of male mice after CPN ligation and quantification of the percentage of c-Fos + neurons ( n = 4 mice). Scale bar, 20 μm. C Representative images of c-Fos + (red) neurons in the magnocellular red nucleus (RMC) after sham and CPN in Fos-Cre ERT2 ;Ai14 transgenic male mice and quantification of the percentage of c-Fos + neurons ( n = 3 mice). Scale bar, 100 μm. D Representative photomicrographs of the viral injection site in RMC. Scale bar, 100 μm. E Heatmap of calcium signals recorded in RMC neurons transfected with AAV-hSyn1-GCaMP6s-P2A-nls-dTomato, aligned to pinch stimulation in CPN mice, alongside representative calcium signal traces from a single animal (3 trials). F Representative fluorescence signals (Δ F / F ) (the average Δ F / F response across 3 trials from a single mouse) of GCaMP6s (red) and control (blue) recorded from RMC CaMKIIα neurons. G Quantification of Ca 2+ signal in RMC neurons with fiber photometry (the statistical summary (mean ± SEM) of peak of Δ F / F across 3 trials for individual mice in each group) ( n = 9 trials for Sham+GcaMP6s, n = 9 trials for CPN+GcaMP6s from 3 mice each group). H Schematic of electrophysiological recording from RMC in brain slices of mice after CPN. I The representative field potentials of RMC neurons in sham and CPN-ligated mice and the stimulation time points. J Representative action potentials of RMC neurons in sham and CPN-ligated mice. K Quantification of action potentials of RMC neurons as shown in ( I ) ( n = 21 neurons for the sham group from 4 mice, n = 22 neurons for the CPN group from 4 mice). L , M Quantification of peak and decay time of field potentials of RMC neurons in sham and CPN-treated mice ( n = 12 groups of neurons for the sham group from 5 mice, n = 13 groups of neurons for the CPN group from 3 mice). N Representative traces of mEPSC recordings in RMC neurons from sham and CPN-ligated mice. O Quantification graphs of frequency and amplitude of mEPSC in RMC neurons from sham and CPN-ligated mice ( n = 20 neurons for the sham group from 3 mice, n = 41 neurons for the CPN group from 4 mice). P Cumulative probability plots of mEPSC frequency of RMC neurons from sham and CPN-ligated mice. Data are presented as the mean ± SEM, t -test or two-way ANOVA, * P < 0.05, ** P < 0.01.

Article Snippet: Fos-Cre ERT2 ; Ai14 mice were obtained by crossing transgenic Fos-Cre ERT2 mice (Shanghai Model Organisms Center, Shanghai, China) with Ai14 mice (Cre-dependent tdTomato reporter, Shanghai Model Organisms Center, Shanghai, China). tdTomato expression was induced in Fos-Cre ERT2 ; Ai14 mice by 4-hydroxytamoxifen (4-OHT) treatment [50 mg/kg, MedChemExpress (MCE), New Jersey, USA] intraperitoneally (i.p.).

Techniques: Ligation, Labeling, Transgenic Assay, Injection, Transfection, Fluorescence, Control

Journal: Neuroscience Bulletin

Article Title: A Novel Long-range Excitatory Neural Circuit from the Magnocellular Red Nucleus to Spinal Dorsal Horn Facilitates Neuropathic Pain-like Behaviors in Male Mice

doi: 10.1007/s12264-025-01553-7

Figure Lengend Snippet: RMC CaMKIIα -DH CaMKIIα neural circuit facilitated the neuropathic pain in male mice after CPN ligation. A Representative images showing neurons in the dorsal horn (DH) that were microinjected with AAV2-retro-hSyn-Cre-P2A-GFP, exhibiting co-localization with AAV9-EF1α-DIO- hM4D(Gi)-mCherry (Retro-hSyn-Cre+DIO-hM4D(Gi)) viruses in RMC. Scale bars, 100 μm. B , C Quantification of thermal paw-withdraw latency ( B ) and PWT ( C ) in CPN-ligated mice upon RMC-DH circuit chemo-inhibition ( n = 6 mice for the Control group, n = 7 mice for the Retro-hSyn-Cre+DIO-hM4D(Gi) group). D Representative image showing neurons in the DH that were transduced by scAAV2/1-hSyn-Cre-pA injected into the RMC of an Ai14 mouse and were co-localized with the excitatory neuronal marker CaMKIIα. Scale bars, 50 μm. Data were mean ± SEM, two-way RM ANOVA, * P < 0.05.

Article Snippet: Fos-Cre ERT2 ; Ai14 mice were obtained by crossing transgenic Fos-Cre ERT2 mice (Shanghai Model Organisms Center, Shanghai, China) with Ai14 mice (Cre-dependent tdTomato reporter, Shanghai Model Organisms Center, Shanghai, China). tdTomato expression was induced in Fos-Cre ERT2 ; Ai14 mice by 4-hydroxytamoxifen (4-OHT) treatment [50 mg/kg, MedChemExpress (MCE), New Jersey, USA] intraperitoneally (i.p.).

Techniques: Ligation, Inhibition, Control, Injection, Marker

Journal: Hearing research

Article Title: Generation of a ChAT Cre mouse line without the early onset hearing loss typical of the C57BL/6J strain

doi: 10.1016/j.heares.2020.107896

Figure Lengend Snippet: Breeding ChATCre,Cdh23WT mice with Ai14 reporter mice produced tdTomato-labeled cells (magenta) in known cholinergic nuclei. A. A low magnification image shows the distribution of labeled cholinergic cells in the SOC (magenta). Labeled cells were most numerous in the LSO and VNTB, but were also visible around the margins of the LSO (arrows). In addition to the labeled cells, small bundles of labeled cholinergic axons from the abducens nucleus are visible (arrowheads). Scale bar (A) = 250 μm. B-G. High magnification images of tdTomato-labeled cells from the B. ventral nucleus of the trapezoid body, C. lateral superior olive, D. facial nucleus, E. motor trigeminal nucleus, F. laterodorsal tegmental nucleus, and G. pedunculopontine tegmental nucleus. Dorsal is up; lateral is to the left. Scale bar (B-G) = 20 μm.

Article Snippet: Finally, the distribution of fluorescently labeled cells in the brains of ChAT Cre,Cdh23WT : Ai14 mice matches the known distribution of cholinergic cells ( VanderHorst and Ulfake, 2006 ).

Techniques: Produced, Labeling

Journal: Hearing research

Article Title: Generation of a ChAT Cre mouse line without the early onset hearing loss typical of the C57BL/6J strain

doi: 10.1016/j.heares.2020.107896

Figure Lengend Snippet: A majority of cholinergic cells are labeled by the reporter, tdTomato, in ChATCre,Cdh23WT:Ai14 mice. Images show cholinergic cells labeled with anti-ChAT immunochemistry (ChAT+, cyan) and tdTomato-labeled cells (magenta). The proportion of ChAT+ cells that were double labeled with tdTomato varied from nucleus to nucleus. Arrows show examples of ChAT+ neurons that were not labeled by the tdTomato. A. ventral nucleus of the trapezoid body. B. lateral superior olive. C. facial nucleus. D. motor trigeminal nucleus. E. laterodorsal tegmental nucleus. F. pedunculopontine tegmental nucleus. Scale bar (A-F) = 20 μm.

Article Snippet: Finally, the distribution of fluorescently labeled cells in the brains of ChAT Cre,Cdh23WT : Ai14 mice matches the known distribution of cholinergic cells ( VanderHorst and Ulfake, 2006 ).

Techniques: Labeling

Journal: Hearing research

Article Title: Generation of a ChAT Cre mouse line without the early onset hearing loss typical of the C57BL/6J strain

doi: 10.1016/j.heares.2020.107896

Figure Lengend Snippet: Most cholinergic cells express Cre, as indicated by tdTomato fluorescence. The percentage of cholinergic cells (ChAT+) that were successfully labeled by the ChAT-driven Cre expression of tdTomato (tdT+) in three mice is shown for samples from two sections through each of three brainstem regions: facial nucleus, motor trigeminal nucleus (Motor V) and the pontomesencephalic cholinergic nuclei. Columns 3 and 4 show the number of ChAT+ cells that co-contained the tdTomato fluorescent protein (# ChAT+/tdT+) or that did not contain the tdTomato signal (# ChAT+/tdT−).

Article Snippet: Finally, the distribution of fluorescently labeled cells in the brains of ChAT Cre,Cdh23WT : Ai14 mice matches the known distribution of cholinergic cells ( VanderHorst and Ulfake, 2006 ).

Techniques: Fluorescence, Labeling, Expressing