Page 70 - 76_03

ALBERTO BARTOLOMÉ Y COLS. AN. R. ACAD. NAC. FARM.

Figure 2. Insulin and glucose action on the PI3K/TSC/mTORC1 pathway. Cells
were stimulated with insulin 10 nM for 5 min (IR +/+, Rec A and Rec B), or glucose
5 mM (IR -/-) for 15 min. Specific inhibitors were added 30 min prior to stimulation
(wortmannin 40 nM, U0126 5 µM). Protein extracts were collected and submitted to
Western-blot with the use of specific antibodies. Blots are representative of three in-
dependent experiments. Quantification of the most significative points is shown. Data
are presented as means ± S.E.M. *P < 0.05 compared to control points.

3.4. Energetic status-dependent modulation of mTORC1

AMPK is the energetic sensor of the cell and it is activated by a
rise in AMP/ATP ratio (29). Thus, AMPK-Thr172 phosphorylation is de-
creased by glucose and increased by 2 deoxylucose (2-DG) in a dose-
dependent manner as well as by 4 mM AICAR in IR +/+ ß cells.
Activation of AMPK by 2-DG or AICAR addition to the culture medi-
um inhibited mTOR and p70S6K phosphorylation. Conversely, inac-

364