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. 2013 Apr 11;27(6):990–1003. doi: 10.1210/me.2012-1338

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Copyright © 2013 by The Endocrine Society

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Figure 7. — Leptin modulation of neuronal transcript levels is abolished after treatment with LY294002 or wortmannin. rHypoE-19 neurons were pretreated with PBS vehicle, DMSO vehicle, 25 μM LY294002 (LY), or 1 μM wortmannin (WORT) for 1 hour before treatment with 10 nM leptin or PBS vehicle. First, it was ensured that induction of phospho-AKT after 15 minutes of insulin treatment is blocked by LY294002 and wortmannin (a). mRNA was isolated 1 hour (for AgRP only) or 8 hours after treatment as indicated and analyzed using qRT-PCR. Pretreatment with inhibitors attenuates leptin-induced increase of UCN2 (b), IR (c), IRS1 (d), IRS2 (e), and decrease in AgRP (f) transcript levels. mRNA levels were normalized to 18S rRNA and shown with SEMs (n = 4–9 independent experiments). *, P < .05; **, P < .01; **, P < .001, as per 2-way ANOVA with a Bonferroni post hoc test.