Palmitic Acid Hydroxystearic Acids Activate GPR40, Which Is Involved in Their Beneficial Effects on Glucose Homeostasis.

Palmitic acid hydroxystearic acids (PAHSAs) are endogenous lipids with anti-diabetic and anti-inflammatory effects. PAHSA levels are reduced in serum and adipose tissue of insulin-resistant people and high-fat diet (HFD)-fed mice. Here, we investigated whether chronic PAHSA treatment enhances insulin sensitivity and which receptors mediate PAHSA effects. Chronic PAHSA administration in chow- and HFD-fed mice raises serum and tissue PAHSA levels ∼1.4- to 3-fold. This improves insulin sensitivity and glucose tolerance without altering body weight. PAHSA administration in chow-fed, but not HFD-fed, mice augments insulin and glucagon-like peptide (GLP-1) secretion. PAHSAs are selective agonists for GPR40, increasing Ca+2 flux, but not intracellular cyclic AMP. Blocking GPR40 reverses improvements in glucose tolerance and insulin sensitivity in PAHSA-treated chow- and HFD-fed mice and directly inhibits PAHSA augmentation of glucose-stimulated insulin secretion in human islets. In contrast, GLP-1 receptor blockade in PAHSA-treated chow-fed mice reduces PAHSA effects on glucose tolerance, but not on insulin sensitivity. Thus, PAHSAs activate GPR40, which is involved in their beneficial metabolic effects.Supported by NIH grants R01 DK43051, P30 DK57521 (B.B.K.), and R01 DK106210 (B.B.K. and A. Saghatelian); a grant from the JPB Foundation (B.B.K.); and T32DK07516 (B.B.K. and J.L.)

Ca2+/Calmodulin-Dependent Protein Kinase Kinase Is Not Involved in Hypothalamic AMP-Activated Protein Kinase Activation by Neuroglucopenia

Hypoglycemia and neuroglucopenia stimulate AMP-activated protein kinase (AMPK) activity in the hypothalamus and this plays an important role in the counterregulatory responses, i.e. increased food intake and secretion of glucagon, corticosterone and catecholamines. Several upstream kinases that activate AMPK have been identified including Ca2+/Calmodulin-dependent protein kinase kinase (CaMKK), which is highly expressed in neurons. However, the involvement of CaMKK in neuroglucopenia-induced activation of AMPK in the hypothalamus has not been tested. To determine whether neuroglucopenia-induced AMPK activation is mediated by CaMKK, we tested whether STO-609 (STO), a CaMKK inhibitor, would block the effects of 2-deoxy-D-glucose (2DG)-induced neuroglucopenia both ex vivo on brain sections and in vivo. Preincubation of rat brain sections with STO blocked KCl-induced α1 and α2-AMPK activation but did not affect AMPK activation by 2DG in the medio-basal hypothalamus. To confirm these findings in vivo, STO was pre-administrated intracerebroventricularly (ICV) in rats 30 min before 2DG ICV injection (40 µmol) to induce neuroglucopenia. 2DG-induced neuroglucopenia lead to a significant increase in glycemia and food intake compared to saline-injected control rats. ICV pre-administration of STO (5, 20 or 50 nmol) did not affect 2DG-induced hyperglycemia and food intake. Importantly, activation of hypothalamic α1 and α2-AMPK by 2DG was not affected by ICV pre-administration of STO. In conclusion, activation of hypothalamic AMPK by 2DG-induced neuroglucopenia is not mediated by CaMKK

Increased Insulin Sensitivity and Reduced Adiposity in Phosphatidylinositol 5-Phosphate 4-Kinase β(−/−) Mice

Phosphorylated derivatives of the lipid phosphatidylinositol are known to play critical roles in insulin response. Phosphatidylinositol 5-phosphate 4-kinases convert phosphatidylinositol 5-phosphate to phosphatidylinositol 4,5-bis-phosphate. To understand the physiological role of these kinases, we generated mice that do not express phosphatidylinositol 5-phosphate 4-kinase β. These mice are hypersensitive to insulin and have reduced body weights compared to wild-type littermates. While adult male mice lacking phosphatidylinositol 5-phosphate 4-kinase β have significantly less body fat than wild-type littermates, female mice lacking phosphatidylinositol 5-phosphate 4-kinase β have increased insulin sensitivity in the presence of normal adiposity. Furthermore, in vivo insulin-induced activation of the protein kinase Akt is enhanced in skeletal muscle and liver from mice lacking phosphatidylinositol 5-phosphate 4-kinase β. These results indicate that phosphatidylinositol 5-phosphate 4-kinase β plays a role in determining insulin sensitivity and adiposity in vivo and suggest that inhibitors of this enzyme may be useful in the treatment of type 2 diabetes

STO-609 does not affect counter-regulatory responses to neuroglucopenia <i>in vivo</i>.

<p>Saline or STO-609 (5, 20 or 50 nmol) was injected ICV in fed rats 30 min before ICV injection of saline or 2DG (40 µmol) as described in “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036335#s4" target="_blank">Materials and Methods</a>”. Glycemia (A) and 1-h food intake (B) were measured. n = 5−9 rats per group. Data are means ± SEM. *<i>p</i><0.05 vs. saline-saline group.</p

STO-609 does not block 2DG-induced AMPK activation in ARC and VMH/DMH.

<p>Saline or STO-609 (50 nmol) was injected ICV in fed rats 30 min prior to saline or 2DG (40µmol) ICV injection. Hypothalamic nuclei were dissected 10 min after saline or 2DG injection. α1 (A) and α2 (B) AMPK activities were measured in microdissected Arcuate and VMH/DMH nuclei. n = 9 rats per group. Data are means ± SEM. *<i>p</i><0.05 vs. saline-saline group.</p

CaMKKα and CaMKKβ protein expression in hypothalamic nuclei and non-hypothalamic brain areas.

<p>Tissue lysates (40 µg protein) were subjected to Western blotting with anti-CaMKKα or anti-CaMKKβ antibodies (Santa Cruz) as described in “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036335#s4" target="_blank">Materials and Methods</a>”. Hip; hippocampus, Arc; arcuate nucleus, VMH/DMH; ventromedial hypothalamus/dorsomedial hypothalamus, PVN; paraventricular nucleus, LH; lateral hypothalamus, HB; hindbrain.</p