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. Author manuscript; available in PMC: 2018 Dec 1.
Published in final edited form as: Glia. 2017 Sep 14;65(12):2003–2023. doi: 10.1002/glia.23210

Figure 10. Involvement of PI-3K/AKT in the nociceptin-dependent upregulation of GLAST expression.

Figure 10

(A) Astrocyte cultures were first pre-incubated for 2 hrs in DMEM-F12 alone; and then for increasing times (0-60 min) in CDM alone (white bars), CDM with 1μM nociceptin (Noci), CDM with 30μM LY294002 (LY) (PI-3K inhibitor) or CDM with 1μM Noci and 30μM LY. Cell lysates were then subjected to western blot analysis with anti-phosphorylated AKT (pAKT) antibody. The results are the mean ± SEM from 3 different cultures; control vs. Noci at 60 min, ***p<0.001. (B) Immunocytochemistry was used to evaluate GLAST expression in rat astrocytes after a 24 hr treatment with CDM alone, or CDM supplemented with one of the following: 1μM Noci, 30μM LY, or 1μM Noci + 30μM LY. The bar graph shows the number of GLAST positive cells as a % of the GFAP(+) cells/field under each condition. Results are the mean ± SEM from twelve fields/well, 3 wells/condition. Control vs. LY, n.s.; control vs. Noci,***p<0.001; Noci vs. Noci + LY, ***p<0.001. (C) Astrocytes treated for 24 hrs as in (B), were subjected to western blot analysis for GLAST. GLAST levels in the bar graph are expressed as change relative to control values. The results are the mean ± SEM, n=3, Control vs. LY, n.s.; control vs. Noci,***p<0.001; Noci vs. Noci + LY, ***p<0.05.