Effects of acutely inhibiting PI3K isoforms and mTOR on regulation of glucose metabolism in vivo

Abstract

In in vitro studies class-I PI3Ks (phosphoinositide 3-kinases), class-II PI3Ks and mTOR (mammalian target of rapamycin) have all been described as having roles in the regulation of glucose metabolism. The relative role each plays in the normal signalling processes regulating glucose metabolism in vivo is less clear. Knockout and knockin mouse models have provided some evidence that the class-I PI3K isoforms p110α, p110β, and to a lesser extent p110γ, are necessary for processes regulating glucose metabolism and appetite. However, in these models the PI3K activity is chronically reduced. Therefore we analysed the effects of acutely inhibiting PI3K isoforms alone, or PI3K and mTOR, on glucose metabolism and food intake. In the present study impairments in glucose tolerance, insulin tolerance and increased hepatic glucose output were observed in mice treated with the pan-PI3K/mTOR inhibitors PI-103 and NVP-BEZ235. The finding that ZSTK474 has similar effects indicates that these effects are due to inhibition of PI3K rather than mTOR. The p110α-selective inhibitors PIK75 and A66 also induced these phenotypes, but inhibitors of p110β, p110δ or p110γ induced only minor effects. These drugs caused no significant effects on BMR (basal metabolic rate), O2 consumption or water intake, but BEZ235, PI-103 and PIK75 did cause a small reduction in food consumption. Surprisingly, pan-PI3K inhibitors or p110α inhibitors caused reductions in animal movement, although the cause of this is not clear. Taken together these studies provide pharmacological evidence to support a pre-eminent role for the p110α isoform of PI3K in pathways acutely regulating glucose metabolism.

Figure 1. Acute effect of PI3K inhibitors on insulin tolerance

The indicated PI3K inhibitors were administered intraperitoneally (10 mg/kg of body mass) and 1 h later animals were injected with insulin (0.75 units/kg of body mass). Glucose levels in blood were measured in blood samples taken at the indicated time as described in the Experimental section. Results are means±S.E.M. (n≥6). Statistical significance was determined by repeated measures ANOVA (***P<0.001 compared with the vehicle control animals).

Figure 2. Acute effect of PI3K inhibitors on hepatic glucose output

The indicated PI3K inhibitors were administered intraperitoneally (10 mg/kg of body mass) and 1 h later animals were injected with pyruvate (2 g/kg of body mass). Glucose levels in blood were measured in blood samples taken at the indicated time as described in the Experimental section. Results are means±S.E.M. (n≥6). Statistical significance was determined by repeated measures ANOVA (***P<0.001 compared with the vehicle control animals).

Figure 3. Acute effect of PI3K inhibitors on glucose tolerance

The indicated PI3K inhibitors were administered intraperitoneally (10 mg/kg of body mass) and 1 h later animals were injected with glucose (2 g/kg of body mass). Glucose levels in blood were measured in blood samples taken at the indicated time as described in the Experimental section. Results are means±S.E.M. (n≥6). Statistical significance was determined by repeated measures ANOVA (*P < 0.05, **P<0.01 and ***P<0.001 compared with the vehicle control animals).

Figure 4. Acute effects of PI3K inhibitors on insulin levels during the GTT

The indicated PI3K inhibitors were administered intraperitoneally (10 mg/kg of body mass) and a GTT was performed as described in Figure 3. Insulin levels were measured in blood samples taken at the indicated time as described in the Experimental section. Results are means±S.E.M. (n≥6). Statistical significance was determined by repeated measures ANOVA (**P<0.01 and ***P<0.001 compared with the vehicle control animals).

Figure 5. Effect of PI3K inhibitors on oxygen consumption, BMR, food consumption and water intake

Animals were injected with the indicated PI3K inhibitors intraperitoneally (10 mg/kg of body mass) and were observed for 24 h following injection in a CLAMS metabolic cage as described in the Experimental section. The results are shown for oxygen consumption (A), BMR (B), food intake during the day (C), food intake during the night (D), water intake during the day (E) and water intake during the night (F). Results are means±S.E.M. (n≥6). Statistical significance was determined by one-way ANOVA and Dunnett's multiple comparison test (*P<0.05 and **P<0.01 compared with the vehicle control animals).

Figure 6. Effects of PI3K inhibitors on movement

The indicated PI3K inhibitors were administered intraperitoneally (10 mg/kg of body mass) and measurements of animal movement were made over a 24 h period in metabolic cages as described in the Experimental section. The results are shown for the total X-counts during the day (A), total X-counts during the night (B), total Z-counts during the day (C) and total Z-counts during the night (D). Results are means±S.E.M. (n≥6). Statistical significance was determined by one-way ANOVA and Dunnett's multiple comparison test (*P<0.05, **P<0.01 and ***P<0.001 compared with the vehicle control animals).