Short term voluntary overfeeding disrupts brain insulin control of adipose tissue lipolysis

Abstract

Insulin controls fatty acid (FA) release from white adipose tissue (WAT) through direct effects on adipocytes and indirectly through hypothalamic signaling by reducing sympathetic nervous system outflow to WAT. Uncontrolled FA release from WAT promotes lipotoxicity, which is characterized by inflammation and insulin resistance that leads to and worsens type 2 diabetes. Here we tested whether early diet-induced insulin resistance impairs the ability of hypothalamic insulin to regulate WAT lipolysis and thus contributes to adipose tissue dysfunction. To this end we fed male Sprague-Dawley rats a 10% lard diet (high fat diet (HFD)) for 3 consecutive days, which is known to induce systemic insulin resistance. Rats were studied by euglycemic pancreatic clamps and concomitant infusion of either insulin or vehicle into the mediobasal hypothalamus. Short term HFD feeding led to a 37% increase in caloric intake and elevated base-line free FAs and insulin levels compared with rats fed regular chow. Overfeeding did not impair insulin signaling in WAT, but it abolished the ability of mediobasal hypothalamus insulin to suppress WAT lipolysis and hepatic glucose production as assessed by glycerol and glucose flux. HFD feeding also increased hypothalamic levels of the endocannabinoid 2-arachidonoylglycerol after only 3 days. In summary, overfeeding impairs hypothalamic insulin action, which may contribute to unrestrained lipolysis seen in human obesity and type 2 diabetes.

FIGURE 1.

Brain insulin fails to suppress lipolysis after 3-day HFD feeding. A, experimental protocol. B and C, Ra glycerol at base line (B) and the 1 milliunit·kg⁻¹·min⁻¹ clamp (C) (n ≥ 4/group). D, fold change of plasma free FA levels from base line. Pancreatic clamp was started at the 120-min time point (indicated by the arrow; n ≥ 5/group). All error bars represent S.E. *, p < 0.05; **, p < 0.01; and ***, p < 0.001 versus RC MBH vehicle + 1 milliunit·kg⁻¹·min⁻¹ clamp group.

FIGURE 2.

Brain insulin fails to suppress Hsl activation after 3-day HFD feeding. A and B, Western blot analyses and quantification of epidydimal fat pads harvested after euglycemic pancreatic clamps (n ≥ 5/group). A, quantification of WAT Hsl phosphorylation. B, Western blot and quantification of WAT Akt signaling. C and D, WAT (C) and liver (D) insulin signaling as assessed by Western blot analysis. Epidydimal fat and liver samples were harvested 15 min after an intraperitoneal (IP) bolus of insulin or saline. Before the intraperitoneal injection of insulin, RC- and HFD-fed rats were infused with MBH insulin or vehicle for 6 h (n = 3/group; n = 2/group for saline injected controls). N.S., not statistically significant, as indicated by brackets. E, insulin levels at base line and during the clamp (n ≥ 4/group). F and G, correlation of caloric intake on HFD and Ra glycerol at base line (F) and during the clamp period (G) (both n = 11). All error bars represent S.E.

FIGURE 3.

Short term overfeeding impairs the ability of MBH insulin to suppress hepatic GP. A, glucose infusion rate (GIR) required to maintain euglycemia (n ≥ 5/group). B, area under the curve (AUC) of graph in A (n ≥ 5/group). C, GP at base line and during the 1 milliunit·kg⁻¹·min⁻¹ clamp period (n ≥ 4/group). D, GP suppression (n ≥ 4/group). E, glucose disposal during the clamp period (n ≥ 4/group). All error bars represent S.E. *, p < 0.05; **, p < 0.01; and ***, p < 0.001 versus RC MBH vehicle + 1 milliunit·kg⁻¹·min⁻¹ clamp group unless otherwise indicated. #, p < 0.05; ##, p < 0.01; and ###, p < 0.001 versus HFD groups.

FIGURE 4.

Lipolytic flux correlates with the degree of hepatic GP. A, correlation of Ra glycerol and hepatic GP during clamps of HFD- and RC-fed rats infused with either MBH insulin or vehicle (n = 34). B–D, plasma glucagon (B), leptin (C), and adiponectin (D) levels during the base line and clamp periods. Adiponectin levels were also measured prior to MBH insulin or vehicle infusion (pre-MBH) (n ≥ 3/group). All error bars represent S.E. *, p < 0.05; and **, p < 0.01 as indicated.

FIGURE 5.

HFD feeding increases 2-AG levels in rat hypothalami after only 3 days. A–C, measurements of key endocannabinoids in MBH tissue samples (n ≥ 7/group). D, Ra glycerol flux of 3-day HFD-fed male rats, which were intracerebroventricularly (ICV) infused with either rimonabant or vehicle for 6 h. Ra glycerol was measured at base line and after induction of a 3 milliunits·kg⁻¹·min⁻¹ hyperinsulinemic clamp. All error bars represent S.E. *, p < 0.05 versus RC-fed group. N.S., not statistically significant.