Journal: ACS Central Science

Article Title: Chemogenetic Approach Using Ni(II) Complex–Agonist Conjugates Allows Selective Activation of Class A G-Protein-Coupled Receptors

doi: 10.1021/acscentsci.8b00390

Figure Lengend Snippet: Selective activation of the engineered M1R in primary rat cortical astrocytes. (a) Schematic illustration of selective activation of the His 4 tag-fused M1R over endogenous mAChRs by coordination tethering. (b) Confocal microscopic images of astrocytes co-transfected with AcGFP (green) and M1R(1.18H4) immunostained by an anti-HA tag antibody (magenta). Nuclei were stained with Hoechst33258 (blue). Scale bar: 20 μm. Ca 2+ responses evoked by 30 nM (c) MAC(M1R) and (d) control-2 in astrocytes harboring the plasmids of (red) M1R(1.18H4), (blue) WT M1R, or (black) the control vector ( n = 18). Fura-2 ratiometric images of astrocytes expressing M1R(1.18H4) (e) before and (f) during perfusion of 30 nM MAC(M1R) . Red and blue represent high and low Ca 2+ concentration, respectively. Scale bar: 100 μm. (g) Average Δratio plot evoked by 5 min of treatment of 30 nM MAC(M1R) and control-2 ( n = 11–23). *** denote significant differences from the group of M1R(1.18H4)/ MAC(M1R) /inhibitor(−) (***: P < 0.001, one way analysis of variance with Dunnett’s post hoc test). F(6/117) = 23.05. Concentration–response curves of (h) M1R(1.18H4) and (i) WT M1R on (red) MAC(M1R) and (black) control-2 ( n = 20–25). The data represent the mean ± SEM. Reproducibility of all experiments was confirmed at least two times.

Article Snippet: Primary rat cortical astrocytes were obtained from P2 neonatal SD rat pups (both male and female were used) (Japan SLC) according to the protocol reported by Guaza et al .

Techniques: Activation Assay, Transfection, Staining, Control, Plasmid Preparation, Expressing, Concentration Assay

Journal: Scientific Reports

Article Title: Rhein and rhubarb similarly protect the blood-brain barrier after experimental traumatic brain injury via gp91 phox subunit of NADPH oxidase/ROS/ERK/MMP-9 signaling pathway

doi: 10.1038/srep37098

Figure Lengend Snippet: ( A ) Microscopic images of normal and scratch-wounded rat astrocytes. ( B ) The WB analysis showed that rhubarb and rhein reinforced neuroprotection by significantly downregulating the GFAP protein levels. ( C ) ROS production was increased within 24 h in the Vehicle group compared with the Control group. Rhubarb and rhein markedly reversed this trend in the scratch-wounded rat astrocytes. ( D ) The RT-PCR analysis showed that rhubarb and rhein significantly alleviated gp91 phox and ( E ) MMP-9 mRNA levels, and ( F ) aggravated the ZO-1 mRNA levels compared with the Vehicle group. ( G ) Representative WB analysis of gp91 phox , p-ERK1/2, ERK1/2, MMP-9 and ZO-1 expression in the scratch-wounded rat astrocytes. ( H ) Quantification of the WB indicated that rhubarb and rhein significantly decreased the gp91 phox , ( I ) phosphorylated ERK1/2 and ( J ) MMP-9 levels, and subsequently increased ( K ) ZO-1 expression in the scratch-wounded rat astrocytes. The values are expressed as the mean ± SD, n = 6/group, Δ p < 0.01 vs. the Control group. *p < 0.05 and # p < 0.01 vs. the Vehicle group.

Article Snippet: Primary rat cortical astrocytes were purchased and maintained according to the manufacturer’s protocol (OriCell TM , No. SCCAC-0001, Cyagen, USA).

Techniques: Control, Reverse Transcription Polymerase Chain Reaction, Expressing