Journal: Cells

Article Title: Disruption of Circadian Rhythms by Ambient Light during Neurodevelopment Leads to Autistic-like Molecular and Behavioral Alterations in Adult Mice

doi: 10.3390/cells10123314

Figure Lengend Snippet: Daily rhythms of animal locomotor activities and clock gene expression are disrupted in SD mice. ( A ) A diagram indicating the strategy to generate the SD mice. ( B ) Top: Representative actograms of mouse wheel-running activities in control (CTR) and Short Day (SD) mice. Middle: Morlet wavelet transform analysis below indicates that CTR mice entrained to the 12 h/12 h light/dark cycle and exhibited a circadian period of 24 h. In contrast, SD mice entrained to the 8 h/8 h light/dark cycle and exhibited a circadian period of 8 h. Bottom: Average activities of CTR (n = 4) and SD (n = 3) mice across 24 h. Note that the SD mice exhibited a higher level of total activities in 24 h as compared with the CTR mice. ( C ) Representative microscopic images of immunostaining for PER1 and PER2 in the SCN. ZT1: one hour after light-on for CTR and SD mice; ZT13 and ZT9: 1 h after light-off for CTR and SD mice. Quantitative analysis of staining intensity is shown to the right. The levels at ZT13 and ZT9 were normalized according to the levels at ZT1 in the CTR and the SD mice, respectively. Note that PER1 and Per2 levels were not different between ZT1 and ZT9 in SD mice. Data are displayed as individual values and mean ± SEM. n = 3 in CTR and n = 4–5 in SD. **** p < 0.0001; n.s., not significant vs. ZT1. ( D ) Western blotting images indicate that PER1, PER2, and BMAL1 exhibited circadian oscillations in the hippocampi of the CTR, but not the SD, mice. Quantitation of protein levels is shown below. For this experiment mice were kept in constant darkness for 48 h and hippocampal samples were harvested every 4 h in the next 24 h. Data are displayed as mean ± SEM. n = 3 in CTR and SD. ** p < 0.01; * p < 0.05 vs. CTR.

Article Snippet: Immunohistochemistry was performed as published in [ ], using one of the following primary antibodies: rabbit anti-phospho-eIF4E (Ser209) antibody (1:1000 final dilution; Novus Biologicals, NBP1-19923; RRID: AB_1641951); rabbit anti-phospho-S6 (Ser240/244) antibody (1:1000 final dilution; Cell Signaling Tech, 2215); rabbit anti-PER1 antibody (1:3000, Millipore, AB2201); or goat anti-PER2 antibody (1:300, Santa Cruz, sc-7728).

Techniques: Gene Expression, Control, Immunostaining, Staining, Western Blot, Quantitation Assay

Journal: Scientific Reports

Article Title: Persistent neuronal Ube3a expression in the suprachiasmatic nucleus of Angelman syndrome model mice

doi: 10.1038/srep28238

Figure Lengend Snippet: ( a ) Representative confocal images of immunofluorescent signals of UBE3A and AVP in coronal sections containing the SCN from Ube3a m +/ p + and Ube3a m –/ p + mice sacrificed at ZT8 on a 12:12 LD cycle. ( b ) High-magnification images of regions in white boxes in a . Arrows identify cells co-expressing UBE3A and AVP. ( c ) Quantification of average intensity of AVP immunofluorescence in SCN regions in Ube3a m + /p + and Ube3a m –/ p + mice (p = 0.484, Student’s t test, n = 3 mice). ( d ) Quantification of AVP-UBE3A colocalization in the SCN of Ube3a m –/ p + mice (AVP-positive neurons positive for UBE3A, n = 28–149 cells from 3 sections each from 3 mice; UBE3A-positive neurons positive for AVP: n = 44–274 cells from 3 sections each from 3 mice). ( e ) Representative images of UBE3A and VIP immunofluorescent signals in coronal sections containing the SCN from Ube3a m + /p + and Ube3a m –/ p + mice sacrificed at ZT4 on a 12:12 LD cycle. ( f ) High-magnification images of region in white boxes in e . Arrows identify cells co-expressing UBE3A and VIP; double-headed arrows indicate UBE3A-positive cells that lack VIP expression. ( g ) Quantification of average intensity of VIP immunofluorescence in SCN regions in Ube3a m + /p + and Ube3a m –/ p + mice (p = 0.568, Student’s t test, n = 3 mice). ( h ) Quantification of VIP-UBE3A colocalization in the SCN of Ube3a m –/ p + mice (VIP-positive neurons positive for UBE3A: n = 4–56 cells from 1–3 sections each from 5 mice; UBE3A-positive neurons positive for VIP: n = 50–243 cells from 1–3 sections each from 5 mice). Scale bars, 100 μm ( a , e ), 25 μm ( b , f ).

Article Snippet: The following primary antibodies were used: mouse anti-UBE3A (1:1,000, clone 3E5, Sigma Cat. #SAB1404508); mouse anti-NEUN (1:750, clone A60, Millipore, Billerica, MA, USA, Cat. #MAB377); rabbit anti-SOX2 (1:2,000, Active Motif, Carlsbad, CA, USA, Cat. #39823); rabbit anti-VIP (1:2,000, Immunostar, Hudson, WI, USA, Cat. #20077); rabbit anti-AVP (1:2,000, Millipore Cat. #AB1565); mouse anti-BMAL1 (1:2,000, Bethyl Laboratories, Montgomery, TX, USA, Cat. #A302–616 A); rabbit anti-PER2 (1:5,000, clone R38; Millipore, Cat. #AB2202); rabbit Anti-GFAP (1:750, Dako North America, Inc., Carpinteria, CA, USA, Cat. #Z0334); and chicken anti-GFP (1:2,000, Aves Labs, Tigard, OR, USA, Cat. #GFP-1020).

Techniques: Expressing, Immunofluorescence