Journal: Scientific Reports

Article Title: A bitopic mTORC inhibitor reverses phenotypes in a tuberous sclerosis complex model

doi: 10.1038/s41598-025-08345-z

Figure Lengend Snippet: Rapalink-1 Treatment in a TSC Mouse Model. ( A ) Schematic diagram of conditional Tsc2 and inducible RFP genes. CRE recombinase causes Tsc2 exons 2–4 to recombine, and RFP to be expressed. ( B ) Schematic diagram of CAG-CRE and CAG-GFP (Green) plasmid electroporation. Plasmid is injected intraventricularly into P0-1 mice. Electrodes placed on the head of the pup subsequently cause plasmids to be taken up by NSCs lining the lateral ventricles when an electrical field is applied. C , D ) Macroscopic images of coronal sections of P60 mice following electroporation. Black indicates RFP expression whereas blue indicates RFP saturation. E - G ) Images of 20× Tsc2 wt/wt coronal section demonstrating induction of RFP (red, E), pS6 staining (green, F), and composite with GFP expression (blue, G). H - J ) Images of 20× Tsc2 mut/mut coronal section demonstrating induction of RFP (red, H), pS6 staining (green, I), and composite with GFP expression (blue, J). K-L ) 4× digital zoom of H-I showing cells with neuronal morphology and high pS6. Chevrons indicate macroscopic neurons, and arrowhead indicates a giant cell. E-J) Scale bar = 75 μm. K-M) Scale bar = 75 μm.

Article Snippet: Rabbit Monoclonal pS6 Ser 240/244 , Cell Signaling Technology , Cat#4838; RRID: AB_659977; Clone: 61H9.

Techniques: Plasmid Preparation, Electroporation, Injection, Expressing, Staining

Journal: Scientific Reports

Article Title: A bitopic mTORC inhibitor reverses phenotypes in a tuberous sclerosis complex model

doi: 10.1038/s41598-025-08345-z

Figure Lengend Snippet: Rapalink-1 Inhibits mTORC1 Activity. A-C) 20× images of RFP (red, A), pS6 (green, B), and composite with GFP (blue, C) in a coronal section of a Tsc2 mut/mut P90 vehicle treated mouse brain. Rectangle denotes magnified region in D-F. D-F) 4× digital zoomed in images of A-C demonstrating neuronal morphology. G-I) 20× images of RFP (red, G), pS6 (green, H), and composite with GFP (blue, I) in a coronal section of a brain from a Tsc2 mut/mut P90 Rapalink-1 (3 mg/kg) treated mouse. J-L) 4× digital zoomed in images of G-I demonstrating neuronal morphology. M) Quantification of pS6 in RFP positive cells. *= p < 0.05, ****= p < 0.0001. Data is represented as mean ± SEM. A-C and G-I scale bar = 75 μm. D-F and J-L scale bar = 18.75 μm.

Article Snippet: Rabbit Monoclonal pS6 Ser 240/244 , Cell Signaling Technology , Cat#4838; RRID: AB_659977; Clone: 61H9.

Techniques: Activity Assay

Journal: Scientific Reports

Article Title: A bitopic mTORC inhibitor reverses phenotypes in a tuberous sclerosis complex model

doi: 10.1038/s41598-025-08345-z

Figure Lengend Snippet: Rapalink-1 Reduces Cell Size. A-D) 20× images of GFP (blue, A), RFP (red, B), pS6 (green, C), and composite (D) in Tsc2 mut/mut P90 control brains. E-H) 20× images of GFP (blue, E), RFP (red, F), pS6 (green, G), and composite (H) in Tsc2 mut/mut P90 Rapalink-1 (3 mg/kg) brains. I, J) Images from D and H magnified 400% showing examples of giant cells. K) Quantification of neuron soma size relative to controls. Data is represented as mean ± SEM. Scale bar = 75 μm. ****= p < 0.0001.

Article Snippet: Rabbit Monoclonal pS6 Ser 240/244 , Cell Signaling Technology , Cat#4838; RRID: AB_659977; Clone: 61H9.

Techniques: Control

Journal: Scientific Reports

Article Title: A bitopic mTORC inhibitor reverses phenotypes in a tuberous sclerosis complex model

doi: 10.1038/s41598-025-08345-z

Figure Lengend Snippet: Rapalink-1 Reduces Hamartoma Size. A-D) 20× images of GFP (blue), pS6 (green), and RFP (red) in Tsc2 mut/mut P90 brains treated with vehicle or E-H) 20× images of GFP (blue), pS6 (green), and RFP (red) in Tsc2 mut/mut P90 3.0 mg/kg Rapalink-1 treated brains. I) Quantification of lesion area. J) Schematic diagram of molecular pathways regulated by Tsc1/Tsc2 in TSC model in control and Rapalink-1 treated brains. Tsc1/Tsc2 turn off mTORC1 pathway under normal conditions. However, in the TSC SEGA model (left), Tsc1/Tsc2 encoded protein products cannot turn mTORC1 off leading to ectopic/cytomegalic neurons with excessive mTORC1 activity, hypertrophic dendrites, giant cells, and SEGA-like lesions (i.e. striatal hamartomas). Rapalink-1 appears to partially rescue these changes with mTORC1 inhibited, neuron size decreased, and reduced SEGA-like lesions. Data are represented as mean ± SEM. Scale bar = 150 μm (A, B) or 37.5 μm (C, D). *= p < 0.05.

Article Snippet: Rabbit Monoclonal pS6 Ser 240/244 , Cell Signaling Technology , Cat#4838; RRID: AB_659977; Clone: 61H9.

Techniques: Control, Activity Assay

Journal: bioRxiv

Article Title: Development of the visual system in social poison frog tadpoles

doi: 10.1101/2022.10.18.512729

Figure Lengend Snippet: ( A ) Begging tadpoles had higher pS6 staining in the ganglion cell layer (GCL) and inner nuclear layer (INL) compared to non-begging tadpoles. ( B ) Photomicrographs showing pS6 staining in the GCL and INL, but not in the outer nuclear layer (ONL). Dotted lines approximately separate layers. Scale bar = 10 um. ( C ) The number of pS6-stained cells positively correlated to tadpole stage. ( D ) While metamorphic begging tadpoles had higher activation in the ganglion cell layer, there was no difference in pS6 staining in the retina of control, begging, and aggressive pre-metamorphic tadpoles. Different lowercase letters represent significant differences (P<0.05).

Article Snippet: Slides were allowed to come to room temperature, washed with 1x PBS 3×10min, blocked in 1x PBS with 5% normal goat serum (NGS) and 0.3% triton-X, incubated in 1:5000 rabbit anti-pS6 (Invitrogen; cat# 44-923G) prepared in blocking solution overnight at 4°C.

Techniques: Staining, Activation Assay