Upper intestinal lipids trigger a gut-brain-liver axis to regulate glucose production.

Energy and glucose homeostasis are regulated by food intake and liver glucose production, respectively. The upper intestine has a critical role in nutrient digestion and absorption. However, studies indicate that upper intestinal lipids inhibit food intake as well in rodents and humans by the activation of an intestine-brain axis. In parallel, a brain-liver axis has recently been proposed to detect blood lipids to inhibit glucose production in rodents. Thus, we tested the hypothesis that upper intestinal lipids activate an intestine-brain-liver neural axis to regulate glucose homeostasis. Here we demonstrate that direct administration of lipids into the upper intestine increased upper intestinal long-chain fatty acyl-coenzyme A (LCFA-CoA) levels and suppressed glucose production. Co-infusion of the acyl-CoA synthase inhibitor triacsin C or the anaesthetic tetracaine with duodenal lipids abolished the inhibition of glucose production, indicating that upper intestinal LCFA-CoAs regulate glucose production in the preabsorptive state. Subdiaphragmatic vagotomy or gut vagal deafferentation interrupts the neural connection between the gut and the brain, and blocks the ability of upper intestinal lipids to inhibit glucose production. Direct administration of the N-methyl-d-aspartate ion channel blocker MK-801 into the fourth ventricle or the nucleus of the solitary tract where gut sensory fibres terminate abolished the upper-intestinal-lipid-induced inhibition of glucose production. Finally, hepatic vagotomy negated the inhibitory effects of upper intestinal lipids on glucose production. These findings indicate that upper intestinal lipids activate an intestine-brain-liver neural axis to inhibit glucose production, and thereby reveal a previously unappreciated pathway that regulates glucose homeostasis.

A balance of lipid-sensing mechanisms in the brain and liver.

Recent work has cast a spotlight on the brain as a nutrient-sensing organ that regulates the body's metabolic processes. Here we discuss the physiological and molecular mechanisms of brain lipid sensing and compare these mechanisms to liver lipid sensing. A direct comparison between the lipid-sensing mechanisms in the brain and liver reveals similar biochemical/molecular but opposing physiological mechanisms in operation. We propose that an imbalance between the lipid-sensing mechanisms in the brain and liver may contribute to obesity-associated type 2 diabetes.

Activation of N-methyl-D-aspartate (NMDA) receptors in the dorsal vagal complex lowers glucose production.

Diabetes is characterized by hyperglycemia due partly to increased hepatic glucose production. The hypothalamus regulates hepatic glucose production in rodents. However, it is currently unknown whether other regions of the brain are sufficient in glucose production regulation. The N-methyl-D-aspartate (NMDA) receptor is composed of NR1 and NR2 subunits, which are activated by co-agonist glycine and glutamate or aspartate, respectively. Here we report that direct administration of either co-agonist glycine or NMDA into the dorsal vagal complex (DVC), targeting the nucleus of the solitary tract, lowered glucose production in vivo. Direct infusion of the NMDA receptor blocker MK-801 into the DVC negated the metabolic effect of glycine. To evaluate whether NR1 subunit of the NMDA receptor mediates the effect of glycine, NR1 in the DVC was inhibited by DVC NR1 antagonist 7-chlorokynurenic acid or DVC shRNA-NR1. Pharmacological and molecular inhibition of DVC NR1 negated the metabolic effect of glycine. To evaluate whether the NMDA receptors mediate the effects of NR2 agonist NMDA, DVC NMDA receptors were inhibited by antagonist D-2-amino-5-phosphonovaleric acid (D-APV). DVC D-APV fully negated the ability of DVC NMDA to lower glucose production. Finally, hepatic vagotomy negated the DVC glycine ability to lower glucose production. These findings demonstrate that activation of NR1 and NR2 subunits of the NMDA receptors in the DVC is sufficient to trigger a brain-liver axis to lower glucose production, and suggest that DVC NMDA receptors serve as a therapeutic target for diabetes and obesity.

Sex-specific extraction of organic anions by the rat liver.

The capacity for hepatic elimination of some compounds is different in males and females and differential expression of a number of sinusoidal and canalicular transporters exists. However, the specific events underlying the functional differences are not understood. To determine how sex influences sinusoidal and canalicular organic anion transport, bile duct-cannulated livers from mature Sprague-Dawley rats of both sexes were single-pass perfused with saline containing the model organic anions bromosulphophthalein (BSP), carboxyfluorescein (CF), carboxyfluorescein diacetate (CFDA) or 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS). Assay of effluent perfusate anion concentration showed that BSP, but not DIDS, extraction was significantly higher in male versus female rats. At 20 min perfusion with 50 microM BSP the mean effluent concentration was 5.6 and 20.1 microM in, respectively, male and female rats. HPLC confirmed that the effluent perfusate concentration of BSP was higher in female as compared with male rats and was not contributed to by its glutathione conjugate. With 25 microM DIDS, the effluent concentration reached 7.3 (male) and 8.2 microM (female), indicating high extraction efficiency. In contrast to BSP and DIDS, CF extraction was very low (<20%) so that differences between male and females could not be assessed. Biliary BSP and CF excretion were, respectively, 3.5- and 4-fold higher in male rats. Neither sinusoidal efflux nor biliary excretion of CF was sex-dependent with a higher cytoplasmic load of CF (during CFDA perfusion). Our results suggest that differences in sinusoidal uptake are responsible for the sex-specific hepatic excretion of some organic anions.

In vivo effects of polyunsaturated, monounsaturated, and saturated fatty acids on hepatic and peripheral insulin sensitivity.

OBJECTIVE: Free fatty acids (FFAs) cause insulin resistance and are often elevated in obesity. Chronic ingestion of diets rich in saturated fat induces more insulin resistance than diets rich in unsaturated fat, however, it remains unclear whether different FFAs cause distinct levels of insulin resistance in the short-term, which is relevant to the feeding and fasting cycle. Protein kinase C (PKC)-δ is implicated in hepatic insulin resistance. Therefore, we investigated the effects of short-term elevation of fatty acids with different degrees of unsaturation on hepatic insulin action and liver PKC-δ membrane translocation, a marker of activation. MATERIALS/METHODS: Triglyceride emulsions of Soybean Oil+Heparin (polyunsaturated (POLY)), Olive Oil+Heparin (monounsaturated (MONO)), Lard Oil+Heparin (saturated (SATU)), or saline (SAL) were infused intravenously for 7h to elevate plasma FFA concentrations ~3-4 fold in rats. During the last 2h of infusion, a hyperinsulinemic-euglycemic clamp with tritiated glucose methodology was performed to examine hepatic and peripheral insulin sensitivity. RESULTS: Surprisingly, SATU, MONO, and POLY impaired peripheral insulin sensitivity (glucose utilization divided by insulin) to a similar extent. Furthermore, all lipids induced a similar degree of hepatic insulin resistance compared to SAL. Although there were changes in hepatic content of lipid metabolites, there were no significant differences in liver PKC-δ membrane translocation across fat groups. CONCLUSIONS: In summary, in the short-term, FFAs with different degrees of unsaturation impair peripheral insulin sensitivity and induce hepatic insulin resistance as well as hepatic PKC-δ translocation to the same extent.

Glucose transporter-1 in the hypothalamic glial cells mediates glucose sensing to regulate glucose production in vivo.

OBJECTIVE: Circulating glucose inhibits glucose production in normal rodents and humans, but this glucose effectiveness is disrupted in diabetes due partly to sustained hyperglycemia. We hypothesize that hyperglycemia in diabetes impairs hypothalamic glucose sensing to lower glucose production, and changes of glucose transporter-1 (GLUT1) in the hypothalamic glial cells are responsible for the deleterious effects of hyperglycemia in vivo. RESEARCH DESIGN AND METHODS: We tested hypothalamic glucose effectiveness to increase hypothalamic glucose concentration and lower glucose production in rats induced with streptozotocin (STZ) uncontrolled diabetes, STZ and phlorizin, and whole-body and hypothalamic sustained hyperglycemia. We next assessed the content of glial GLUT1 in the hypothalamus, generated an adenovirus expressing GLUT1 driven by a glial fibrillary acidic protein (GFAP) promoter (Ad-GFAP-GLUT1), and injected Ad-GFAP-GLUT1 into the hypothalamus of rats induced with hyperglycemia. Pancreatic euglycemic clamp and tracer-dilution methodologies were used to assess changes in glucose kinetics in vivo. RESULTS: Sustained hyperglycemia, as seen in the early onset of STZ-induced diabetes, disrupted hypothalamic glucose sensing to increase hypothalamic glucose concentration and lower glucose production in association with reduced GLUT1 levels in the hypothalamic glial cells of rats in vivo. Overexpression of hypothalamic glial GLUT1 in STZ-induced rats with reduced GLUT1 acutely normalized plasma glucose levels and in rats with selectively induced hypothalamic hyperglycemia restored hypothalamic glucose effectiveness. CONCLUSIONS: Sustained hyperglycemia impairs hypothalamic glucose sensing to lower glucose production through changes in hypothalamic glial GLUT1, and these data highlight the critical role of hypothalamic glial GLUT1 in mediating glucose sensing to regulate glucose production.

Central lactate metabolism regulates food intake.

The central nervous system regulates food intake (FI) and body weight (BW), but the associated mechanisms remain to be elucidated. Here we report that central injections of lactate reduced FI and BW in rodents. Inhibition of central lactate metabolism to pyruvate with the lactate dehydrogenase inhibitor oxamate abolished the central effects of lactate on FI and BW. Conversely, central injections of pyruvate recapitulated the effects of lactate. Finally, inhibition of central lactate metabolism prevented the ability of circulating lactate to lower FI and BW. Together, the data indicate that activation of central lactate metabolism lowers FI and BW.

Activation of central lactate metabolism lowers glucose production in uncontrolled diabetes and diet-induced insulin resistance.

OBJECTIVE: Hypothalamic lactate metabolism lowers hepatic glucose production and plasma glucose levels in normal rodents. However, it remains unknown whether activation of hypothalamic lactate metabolism lowers glucose production and plasma glucose levels in rodents with diabetes and obesity. RESEARCH DESIGN AND METHODS: We performed intracerebroventricular (ICV) administration of lactate to enhance central lactate metabolism in 1) early-onset streptozotocin-induced uncontrolled diabetic rodents, 2) experimentally induced hypoinsulinemic normal rodents, and 3) early-onset diet-induced insulin-resistant rodents. Tracer-dilution methodology was used to assess the impact of ICV lactate on the rate of glucose production in all three models. RESULTS: We first report that in the absence of insulin treatment, ICV lactate administration lowered glucose production and glucose levels in rodents with uncontrolled diabetes. Second, ICV lactate administration lowered glucose production and glucose levels in normal rodents with experimentally induced hypoinsulinemia. Third, and finally, ICV lactate administration lowered glucose production in normal rodents with diet-induced insulin resistance. CONCLUSIONS: Central lactate metabolism lowered glucose production in uncontrolled diabetic and normal rodents with hypoinsulinemia and in rodents with diet-induced insulin resistance. These data suggest that insulin signaling is not required for central lactate to lower glucose production and that the activation of hypothalamic lactate metabolism could consequently bypass insulin resistance and lower glucose levels in early-onset diabetes and obesity.

Hypothalamic protein kinase C regulates glucose production.

OBJECTIVE: A selective rise in hypothalamic lipid metabolism and the subsequent activation of SUR1/Kir6.2 ATP-sensitive K(+) (K(ATP)) channels inhibit hepatic glucose production. The mechanisms that link the ability of hypothalamic lipid metabolism to the activation of K(ATP) channels remain unknown. RESEARCH DESIGN AND METHODS: To examine whether hypothalamic protein kinase C (PKC) mediates the ability of central nervous system lipids to activate K(ATP) channels and regulate glucose production in normal rodents, we first activated hypothalamic PKC in the absence or presence of K(ATP) channel inhibition. We then inhibited hypothalamic PKC in the presence of lipids. Tracer-dilution methodology in combination with the pancreatic clamp technique was used to assess the effect of hypothalamic administrations on glucose metabolism in vivo. RESULTS: We first reported that direct activation of hypothalamic PKC via direct hypothalamic delivery of PKC activator 1-oleoyl-2-acetyl-sn-glycerol (OAG) suppressed glucose production. Coadministration of hypothalamic PKC-delta inhibitor rottlerin with OAG prevented the ability of OAG to activate PKC-delta and lower glucose production. Furthermore, hypothalamic dominant-negative Kir6.2 expression or the delivery of the K(ATP) channel blocker glibenclamide abolished the glucose production-lowering effects of OAG. Finally, inhibition of hypothalamic PKC eliminated the ability of lipids to lower glucose production. CONCLUSIONS: These studies indicate that hypothalamic PKC activation is sufficient and necessary for lowering glucose production.

Modulation of hepatocellular swelling-activated K+ currents by phosphoinositide pathway-dependent protein kinase C.

K(+) channels participate in the regulatory volume decrease (RVD) accompanying hepatocellular nutrient uptake and bile formation. We recently identified KCNQ1 as a molecular candidate for a significant fraction of the hepatocellular swelling-activated K(+) current (I(KVol)). We have shown that the KCNQ1 inhibitor chromanol 293B significantly inhibited RVD-associated K(+) flux in isolated perfused rat liver and used patch-clamp techniques to define the signaling pathway linking swelling to I(KVol) activation. Patch-electrode dialysis of hepatocytes with solutions that maintain or increase phosphatidylinositol 4,5-bisphosphate (PIP(2)) increased I(KVol), whereas conditions that decrease cellular PIP(2) decreased I(KVol). GTP and AlF(4)(-) stimulated I(KVol) development, suggesting a role for G proteins and phospholipase C (PLC). Supporting this, the PLC blocker U-73122 decreased I(KVol) and inhibited the stimulatory response to PIP(2) or GTP. Protein kinase C (PKC) is involved, because K(+) current was enhanced by 1-oleoyl-2-acetyl-sn-glycerol and inhibited after chronic PKC stimulation with phorbol 12-myristate 13-acetate (PMA) or the PKC inhibitor GF 109203X. Both I(KVol) and the accompanying membrane capacitance increase were blocked by cytochalasin D or GF 109203X. Acute PMA did not eliminate the cytochalasin D inhibition, suggesting that PKC-mediated I(KVol) activation involves the cytoskeleton. Under isotonic conditions, a slowly developing K(+) current similar to I(KVol) was activated by PIP(2), lipid phosphatase inhibitors to counter PIP(2) depletion, a PLC-coupled alpha(1)-adrenoceptor agonist, or PKC activators and was depressed by PKC inhibition, suggesting that hypotonicity is one of a set of stimuli that can activate I(KVol) through a PIP(2)/PKC-dependent pathway. The results indicate that PIP(2) indirectly activates hepatocellular KCNQ1-like channels via cytoskeletal rearrangement involving PKC activation.

Oleate-induced decrease in hepatocyte insulin binding is mediated by PKC-delta.

We have previously shown that free fatty acids (FFA) impair hepatic insulin extraction in vivo and thus generate hyperinsulinemia, a suspected risk factor for atherosclerosis and cancer. Hepatic insulin extraction is a receptor-mediated event, which is initiated by hepatocyte insulin binding. In the present study, we investigated the effect of FFA on insulin binding in freshly isolated rat hepatocytes maintained at 10 mM glucose. Hepatocyte insulin binding decreased after 1 h exposure to oleate in a concentration-dependent manner reaching a maximum (35-40%) at 125 microM. Inhibition of FFA oxidation by >90% with the carnitine palmitoyltransferase I (CPT-I) inhibitor methylpalmoxirate (MP, 30 microM) did not prevent the effect of oleate. However, when hepatocytes were treated with the PKC inhibitor bisindolylmaleimide (BIM, 1 microM) the effect of oleate was abolished. Subcellular fractionation and immunoblotting of specific PKC isoforms revealed that oleate-induced hepatic PKC-delta membrane translocation, but did not translocate-epsilon, -theta, -alpha, -betaI and -betaII. These results indicate that PKC-delta activation mediated the FFA-induced decrease in hepatocyte insulin binding under our conditions, and thus provides a mechanistic basis for FFA-induced hyperinsulinemia.