Journal: Journal of Inflammation Research

Article Title: Early-Life Stress Induced by Neonatal Maternal Separation Leads to Intestinal 5-HT Accumulation and Causes Intestinal Dysfunction

doi: 10.2147/JIR.S488290

Figure Lengend Snippet: 5-HT promoted neurogenesis in the enteric nervous system. ( A ) The mRNA expression of 5-HT 3 , 5-HT 4 , and 5-HT 7 in the colon (n=6). ( B ) The protein expression of 5-HT 4 and GAPDH in the colon (n=3). ( C ) The mRNA expression of Tubb3, Sox2, Nestin, Olig2, and Gfap in the muscularis (n=6). ( D ) The protein expression of Tuj1, HUC/D, and GAPDH in the muscularis (n=3). ( E ) Immunofluorescence staining of HUC/D (green) and Tuj1 (red) in the LMMP (scale bar=100μm), and the number of HUC/D ( F ) and Tuj1 ( G ) positive cells was quantified. ( H ) Immunofluorescence staining of Tuj1 in the colon (Tuj1, red) and nucleic acid (Dapi, blue) (scale bar=100 μm), and ( I ) the number of Tuj1-positive cells was quantified. ( J ) Immunofluorescence staining of S100β in LMMP (S100β, red) and nucleic acid (Dapi, blue) (scale bar=100 μm), and ( K ) the number of S100β-positive cells was quantified. Data is presented as the mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Subsequently, the cells were incubated with 5-HT (1 μM, H9523, Sigma‒Aldrich, Shanghai, China) along with the 5-HT 4 receptor antagonist GR113808 (10 nM, HY-103152, MCE, USA) or the Wnt pathway inhibitor IWP-2 (20 μM, HY-13912, MCE, USA) in serum-free media for 2 days.

Techniques: Expressing, Immunofluorescence, Staining

Journal: Journal of Inflammation Research

Article Title: Early-Life Stress Induced by Neonatal Maternal Separation Leads to Intestinal 5-HT Accumulation and Causes Intestinal Dysfunction

doi: 10.2147/JIR.S488290

Figure Lengend Snippet: 5-HT 4 mediates 5-HT activation of the Wnt signaling pathway to promote neurogenesis. ( A and B ) The protein expression levels of β-catenin, APC, Axin1, GSK-3β, and GAPDH in the muscularis muscle were determined by Western blotting (n=3). ( C ) Western blotting was used to determine the protein expression of β-catenin, and GAPDH in N2a cells (n=3). ( D ) The expression of Tubb3, Sox2, and Nestin in N2a cells was measured by real-time qPCR (n=6). ( E – H ) The protein expression of β-catenin, HUC/D, Tuj1, and GAPDH in N2a cells was examined by Western blotting, and the relative protein levels were normalized to those of GAPDH (n=3). Data is presented as the mean ± SEM. * P < 0.05; ** P < 0.01.

Article Snippet: Subsequently, the cells were incubated with 5-HT (1 μM, H9523, Sigma‒Aldrich, Shanghai, China) along with the 5-HT 4 receptor antagonist GR113808 (10 nM, HY-103152, MCE, USA) or the Wnt pathway inhibitor IWP-2 (20 μM, HY-13912, MCE, USA) in serum-free media for 2 days.

Techniques: Activation Assay, Expressing, Western Blot

Journal: Journal of Inflammation Research

Article Title: Early-Life Stress Induced by Neonatal Maternal Separation Leads to Intestinal 5-HT Accumulation and Causes Intestinal Dysfunction

doi: 10.2147/JIR.S488290

Figure Lengend Snippet: Overview of the role of 5-HT signaling in ELS-induced intestinal dysfunction. Elevated serotonin levels in the gut due to early stress contribute to intestinal dysfunction in mice. These effects may occur through activation of the 5-HT 4 receptor, stimulation of the Wnt pathway, the promotion of neurogenesis, an increase in excitatory neurons, and subsequent impacts on intestinal motility.

Article Snippet: Subsequently, the cells were incubated with 5-HT (1 μM, H9523, Sigma‒Aldrich, Shanghai, China) along with the 5-HT 4 receptor antagonist GR113808 (10 nM, HY-103152, MCE, USA) or the Wnt pathway inhibitor IWP-2 (20 μM, HY-13912, MCE, USA) in serum-free media for 2 days.

Techniques: Activation Assay

Journal: PLoS ONE

Article Title: Regulation of Amyloid Precursor Protein Processing by Serotonin Signaling

doi: 10.1371/journal.pone.0087014

Figure Lengend Snippet: Overview of agonists, antagonists and inhibitors used to investigate the proteins contributing to the induction of sAPPα after 5-HT 4d receptor stimulation.

Article Snippet: GF109203X, SQ22536, D609, ionomycin, 4,5,6,7-tetrabromo-1H-benzotriazole (TBB), GR113808, NF449, gallein and batimastat were obtained from Tocris.

Techniques: In Vitro

Journal: PLoS ONE

Article Title: Regulation of Amyloid Precursor Protein Processing by Serotonin Signaling

doi: 10.1371/journal.pone.0087014

Figure Lengend Snippet: (A) Prucalopride induced sAPPα secretion in SH-SY5Y human neuroblastoma cells is specific for the 5-HT 4 receptor. SH-SY5Y cells, transfected with pEAK12-AP-APP and pcDNA3.1-5-HT 4d , were treated with 1 µM prucalopride and 5-HT (5-HT 4 receptor agonists) in the absence or presence of 1 µM GR113808 (5-HT 4 receptor antagonist) or PMA and secretion of sAPPα was analyzed via measuring SEAP. (B) SEAP levels were measured in supernatants of SH-SY5Y cells, transfected with pEAK12-AP-APP and pcDNA3.1-5-HT 4d (WT), pcDNA3.1-5-HT 4d DRY117/118AAY (DRY) or pcDNA3.1-5-HT 4d Δ346 (Δ346) mutants and stimulated with 1 µM prucalopride or 5-HT. (C), (D) and (F) SEAP levels were measured in supernatants of SH-SY5Y cells, transfected with pEAK12-AP-APP and pcDNA3.1-5-HT 4d and treated with 1 µM prucalopride or 5-HT in the absence or presence of 100 µM CTB (Gα s inhibitor) (C), 100 µM NF449 (Gα s inhibitor) (D) or 100 µM gallein (Gβγ inhibitor) (F). (E) SEAP levels were measured in SH-SY5Y cells, transfected with pEAK12-AP-APP, pcDNA3.1-5-HT 4d and pcDNAI-Amp-Gα s DN or pcDNA3.1 at a ratio of 2∶1∶4, respectively, and treated with 1 µM prucalopride or 5-HT. Values shown are mean ± SEM of 6 individual wells and were normalized to vehicle control. * P <0.05, ** P <0.01, *** P <0.001, one-way ANOVA with Tukey-Kramer or Dunnet's post-hoc test.

Article Snippet: GF109203X, SQ22536, D609, ionomycin, 4,5,6,7-tetrabromo-1H-benzotriazole (TBB), GR113808, NF449, gallein and batimastat were obtained from Tocris.

Techniques: Transfection, Control