The Journal of Neuroscience, January 24, 2007, ():
Endoplasmic Reticulum Stress Inhibition Protects against Excitotoxic Neuronal Injury in the Rat Brain
J. Neurosci. Sokka et al.
27: 901
Supplemental Data
Files in this Data Supplement:
- supplemental material
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Supporting Fig. 1.
(A) 25然 KA was added to the hippocampal neurons and the integrity of ER was analyzed by the ER-tracker. Arrows show disintegration of ER membranes 1h after KA. Scale bar: 40μm.
(B) Neurons were stimulated for 1h using different concentrations of KA or glutamate (Glu). Viability of cells were analysed by 0.5痢/ml propidium iodine (PI) to show integrity of plasma membrane combined with 4痢/ml Hoescht blue to stain nuclei. Quantification of number of PI positive cells counting more than 100 cells in triplicate wells. No significant change in KA treated cells compared with controls (n=3). 300然 Glu induced a significant increase in PI stained cells compared with controls (p<0.05, n=6). Insert, arrows show PI stained cells in Glu treated cultures. Scale bar: 50μm.
- supplemental material
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Supporting Fig. 2.
In vivo experiments. (A) KA induced transient PERK and eIF2α phosphorylation in the rat hippocampus. CA3, hippocampal CA3 area. Scale bar 125痠.
(B) c-Jun phosphorylation was observed in CA3 neurons at 12h but was abolished by 24h. Scale bar: 25痠.
- supplemental material
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Supporting Fig. 3.
Dose response curve.
Hippocampal neurons were treated for 90 min with different concentrations of KA and glutamate (Glu). Immunblottings for p-eIF2α (A) and Bip (B) were done as described in Methods. Note an increase in both protein species with 25然 KA and 75然 Glu.
- supplemental material
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Supporting Fig. 4.
Calcium responses in hippocampal neurons pretreated with vehicle (a, c, e) or 50然 Salubrinal (b, d, f) and measured by using Fura-2 as described in Methods.
Cells were pretreated with the compounds for 24h (a, b, e, f) or for 30min (c, d) before stimulations with 50然 KA (a-d) or 50然 glutamate (e, f). There were no significant differences in peak amplitude of Ca2+ responses induced by KA or glutamate in Sal treated cells compared with controls. Number of cells was: (a, b) n=17; (c, d) n=11; (e) n=5; (f) n=4.
