Journal: Alcoholism, clinical and experimental research

Article Title: Regulation of Kisspeptin Synthesis and Release in the Preoptic/Anterior Hypothalamic Region of Prepubertal Female Rats: Actions of IGF-1 and Alcohol

doi: 10.1111/acer.13539

Figure Lengend Snippet: Effects of IGF-1 and acute ALC exposure on IRS-1 protein in the POA/RHA of prepubertal female rats. A) Representative Western blot of phosphorylated (p)- and total-IRS-1 proteins from control (lanes 1-3), IGF-1 (lanes 4-6), ALC (7-9) and IGF-1 + ALC (lanes 10-12) animals. B) Each bar represents the mean ± SEM of the densitometric quantification of all bands assessing p-IRS-1 protein normalized to total IRS-1 protein. IGF-1 (checked bar) induced an increase in p-IRS-1 over control (solid bar) animals. Note that 3g/kg exposure to ALC (open bar) did not alter basal expression of p-IRS-1, but that the IGF-1 induced increase of p-IRS-1was blocked by acute ALC exposure (hatched bar). The number of animals represented by each bar is 6. **p<0.01.

Article Snippet: Following transfer, membranes were blocked with 5% nonfat dried milk/0.1% Tween 20 in PBS (pH 7.4) for 3 hr and subsequently incubated at 4 C overnight with rabbit anti-total-IRS-1 (1:500; Cohesion Biosciences Ltd, London, UK) or anti-p-IRS-1 (Ser307; 1:500; Assay Biotechnology Company, Inc, Sunnyvale, CA), rabbit anti-total (1:1000) or anti-p-Akt (Ser473, 1:3000; Cell Signaling Tech., Danvers, MA), rabbit anti-total (1:2000) or anti-p-TSC2 (Thr1462, 1:250; Cell Signaling Tech., Danvers, MA), rabbit anti-Rheb (1μg/ml; Abcam Inc., Cambridge, MA), rabbit anti-total or anti-p-mTOR (Ser2448, 1:1000; Cell Signaling Tech., Danvers, MA) and rabbit anti-Kisspeptin (3μg/ml; Novus Biologicals, Littleton, CO).

Techniques: Western Blot, Expressing

Journal: Alcoholism, clinical and experimental research

Article Title: Regulation of Kisspeptin Synthesis and Release in the Preoptic/Anterior Hypothalamic Region of Prepubertal Female Rats: Actions of IGF-1 and Alcohol

doi: 10.1111/acer.13539

Figure Lengend Snippet: Schematic drawing showing the effects of IGF-1 and ALC on prepubertal Kp synthesis and release. A) IGF-1 activates the IGF-1R/IRS-1 complex, which subsequently initiates critical phosphorylations within the Akt mediated pathway to mTOR; hence resulting in increased Kp synthesis and release. B) ALC blocks the activation of IGF-1R/IRS-1 complex by inhibiting the phosphorylation of IRS-1resulting in the inhibition of key downstream phosphorylations within the Akt pathway to mTOR, thus suppressing Kp synthesis and release. IGF-1, Insulin-like growth factor 1; IGF-1R; insulin-like growth factor receptor 1; IRS-1, insulin receptor substrate 1; Kp, kisspeptin; mTOR, mammalian target of rapamycin; phosphorylated site, p; ras homologue enriched in brain, Rheb; TSC1, tuberous sclerosis complex 2.

Article Snippet: Following transfer, membranes were blocked with 5% nonfat dried milk/0.1% Tween 20 in PBS (pH 7.4) for 3 hr and subsequently incubated at 4 C overnight with rabbit anti-total-IRS-1 (1:500; Cohesion Biosciences Ltd, London, UK) or anti-p-IRS-1 (Ser307; 1:500; Assay Biotechnology Company, Inc, Sunnyvale, CA), rabbit anti-total (1:1000) or anti-p-Akt (Ser473, 1:3000; Cell Signaling Tech., Danvers, MA), rabbit anti-total (1:2000) or anti-p-TSC2 (Thr1462, 1:250; Cell Signaling Tech., Danvers, MA), rabbit anti-Rheb (1μg/ml; Abcam Inc., Cambridge, MA), rabbit anti-total or anti-p-mTOR (Ser2448, 1:1000; Cell Signaling Tech., Danvers, MA) and rabbit anti-Kisspeptin (3μg/ml; Novus Biologicals, Littleton, CO).

Techniques: Activation Assay, Inhibition