Journal: Cells

Article Title: Small Extracellular Vesicles Promote Axon Outgrowth by Engaging the Wnt-Planar Cell Polarity Pathway

doi: 10.3390/cells14010056

Figure Lengend Snippet: sEVs promote neurite elongation and can colocalize with Wnt7b. ( A – C ) sEVs promote neurite elongation. ( A ) A schematic illustration of the two-compartment Xona microfluidic device. The somal compartment is connected to the axonal compartment through a 150 μm microgroove. ( B ) Cortical neurons (E15.5-16.5) were seeded in the somal compartment and cultured for 5 days prior to the addition of L cell-derived sEVs in either the somal or axonal compartment. Neurons were fixed 24 h later, and neuronal morphology was examined in Tuj1 stained neurons. Representative images are shown. Scale bar, 200 μm. ( C ) The length of the neurites growing in the microgroove and emerging in the axonal compartment was quantified for a minimum of 90 neurites. A dotted line marks both ends of the microgroove (150 μm). ( D – I ) sEVs can be internalized by neurons and colocalize with Wnts. Cortical neurons were treated with 10X concentrated conditioned media (CM) from L cells stably expressing CD81-EYFP, 4 h after plating for 29 h ( F ) or 24 h after plating for 30 min ( G – I ). In panel ( G ), after 30 min of treatment, neurons were washed and subsequently treated with regular complete media for 0, 2, 4, and 24 h. Representative images of neurons immunostained with GFP and Tuj1 ( F , G ) or GFP, Tuj1, and Wnt7b ( H ) are shown. Dashed boxes ( H ) indicate higher magnification of neurons. Arrowheads mark GFP puncta of internalized sEVs. Scale bar, 20 μm ( F ) or 40 μm ( G , H ). ( E ) Characterization of sEVs. The concentrated CM (10X) and sEV pellet from L cells were immunoblotted with anti-GFP antibody. ( I ) Pearson’s colocalization coefficient for ( H ). Neurons were identified using Tuj1 as a reference channel, and the colocalization coefficient was quantified using Nikon NIS-Elements software. Images ( F , G , H ) and the quantification ( I ) are representative of 30 neurons from 3 independent experiments. In all the violin plots, values are distinctly colored for each experiment, and the median is marked by a black line. Statistical significance: *** p < 0.001 using unpaired t -test ( I ) or one-way ANOVA with Dunnett’s post-test ( C ).

Article Snippet: Two-compartment microfluidic chambers containing a 150 μm microgroove were purchased from Xona Microfluidics (Durham, NC, USA, XC150).

Techniques: Cell Culture, Derivative Assay, Staining, Stable Transfection, Expressing, Software

Journal: bioRxiv

Article Title: Morphine-context associative memory and locomotor sensitization in mice are modulated by sex and context in a dose-dependent manner

doi: 10.1101/2023.11.03.565492

Figure Lengend Snippet: A) Experimental timeline of the conditioned place preference (CPP) paradigm. B) Illustration of the CPP apparatus in the low and high context version of the apparatus. In the low context version, the walls were distinctly patterned with horizontal stripes in the “left” compartment and zig zag stripes in the “right” compartment, but the flooring was smooth in both compartments. In the high context version, the flooring was also distinct between compartments, with the “left” compartment altered to a textured rubber flooring to distinguish it tactilely from the “right” compartment smooth flooring. C-D) % Time spent in the left compartment of the apparatus in the low ( C ) and high ( D ) context versions of the apparatus on the pretest, when mice could freely explore both compartments of the apparatus for the first time. Individual data points are color coded to reflect categorization of compartment bias on the pretest (pink: > 55%; teal: < 45%; black: between 45% and 55% in left compartment). *p < 0.05 for unpaired t-tests between males and females; # p < 0.05, #### p < 0.0001 for one-sample t-tests within group compared to the null hypothesis mean of 50%. E-F) Pretest locomotion in females and males, subcategorized by compartment bias of 55% as defined in C and D . Females had greater pretest locomotion than males in both the low ( E ) and high ( F ) context versions of the assay, but locomotion was unaffected by compartment bias. *p < 0.05, ****p < 0.0001 for main effects of sex in 2×2 ANOVAs.

Article Snippet: The CPP apparatus consisted of a standard mouse arena (27.3×27.3×20.3 cm) separated into two compartments by a two chamber place preference insert with removable guillotine door (Med Associates Inc., Fairfax, VT, USA).

Techniques: Conditioned Place Preference

Journal: bioRxiv

Article Title: Morphine-context associative memory and locomotor sensitization in mice are modulated by sex and context in a dose-dependent manner

doi: 10.1101/2023.11.03.565492

Figure Lengend Snippet: A-B) On the first day of conditioning (C1) in the low context ( A ) and high context ( B ) versions of the CPP assay, females displayed acute hyperlocomotion following the morphine injection on the afternoon conditioning session, regardless of whether the morphine was paired with the left or right compartment; this locomotor sensitivity was dose-dependent, as cohorts receiving higher doses of morphine displayed greater morphine-induced hyperlocomotion. Locomotion following saline injection on the morning session (6 hr earlier) was similar across morphine dose groups and paired compartment. C-D) Males displayed a similar dose-dependent hyperlocomotor response to morphine that was similar in both compartments of the apparatus in the low ( C ) and high ( D ) context assays, with no differences in saline locomotion. **p < 0.01, ****p < 0.0001 indicate main effect of morphine dose in 2×2 RM-ANOVAs; $$$ p ≤ 0.001, $$$$ p < 0.0001 indicate difference from 5 mg/kg morphine dose, ## p ≤ 0.01, ### p ≤ 0.001 indicates difference from 10 mg/kg morphine dose in post hoc t-tests between doses.

Article Snippet: The CPP apparatus consisted of a standard mouse arena (27.3×27.3×20.3 cm) separated into two compartments by a two chamber place preference insert with removable guillotine door (Med Associates Inc., Fairfax, VT, USA).

Techniques: Injection, Saline