The hepatic energy state, defined by adenine nucleotide levels, couples metabolic pathways with energy requirements. This coupling is fundamental in the adaptive response to many conditions and is impaired in metabolic disease. We have found that the hepatic energy state is substantially reduced following exercise, fasting, and exposure to other metabolic stressors in C57BL/6 mice. Glucagon receptor signaling was hypothesized to mediate this reduction because increased plasma levels of glucagon are characteristic of metabolic stress and because this hormone stimulates energy consumption linked to increased gluconeogenic flux through cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) and associated pathways. We developed what we believe to be a novel hyperglucagonemic-euglycemic clamp to isolate an increment in glucagon levels while maintaining fasting glucose and insulin. Metabolic stress and a physiological rise in glucagon lowered the hepatic energy state and amplified AMP-activated protein kinase signaling in control mice, but these changes were abolished in glucagon receptor-null mice and mice with liver-specific PEPCK-C deletion. 129X1/Sv mice, which do not mount a glucagon response to hypoglycemia, displayed an increased hepatic energy state compared with C57BL/6 mice in which glucagon was elevated. Taken together, these data demonstrate in vivo that the hepatic energy state is sensitive to glucagon receptor activation and requires PEPCK-C, thus providing new insights into liver metabolism. IntroductionThe energy state in the cell is defined by adenine nucleotide levels and is critically coupled to nearly all metabolic processes (1). In the cell, the adenine nucleotides ATP, ADP, and AMP are tied directly or indirectly to all energetic pathways and allosterically control numerous regulatory enzymes (2-6). Changes in adenine nucleotides typically occur such that ATP and AMP deviate in reciprocal directions, while ADP remains constant (1). Such changes are the basis for using the ratio AMP/ATP (1, 7) or the equation for cellular energy charge ([ATP + (0.5 × ADP)] / [ATP + ADP + AMP]) (8, 9) as indices of the metabolic environment. Metabolic stress is thus characterized by a rise in AMP paired with a fall in ATP levels, reflecting a decrease in energy state. A fall in energy state is of considerable importance, in part due to the regulatory role of AMP/ATP ratios on AMPK activity (10). AMPK is a metabolic switch sensitive to high AMP/ATP ratios and functions to protect the energy state by inhibiting ATP-consuming processes while stimulating ATP-producing processes. (10).In most tissues, the environment is controlled to maintain a high energy state (low AMP/ATP ratio). In skeletal muscle, for example, creatine kinase limits reductions in ATP during conditions such as exercise, when energy utilization is accelerated (11,12). The liver, in contrast, lacks creatine kinase, and exercise has been shown to markedly decrease the hepatic energy state and increase the phosphorylation of AMPK (11, 13). The regulatory...

Hepatic glucagon action increases in response to accelerated metabolic demands and is associated with increased whole body substrate availability, including circulating lipids. The hypothesis that increases in hepatic glucagon action stimulate AMP-activated protein kinase (AMPK) signaling and peroxisome proliferator-activated receptor-α (PPARα) and fibroblast growth factor 21 (FGF21) expression in a manner modulated by fatty acids was tested in vivo. Wild-type (gcgr(+/+)) and glucagon receptor-null (gcgr(-/-)) littermate mice were studied using an 18-h fast, exercise, and hyperglucagonemic-euglycemic clamps plus or minus increased circulating lipids. Fasting and exercise in gcgr(+/+), but not gcgr(-/-) mice, increased hepatic phosphorylated AMPKα at threonine 172 (p-AMPK(Thr(172))) and PPARα and FGF21 mRNA. Clamp results in gcgr(+/+) mice demonstrate that hyperlipidemia does not independently impact or modify glucagon-stimulated increases in hepatic AMP/ATP, p-AMPK(Thr(172)), or PPARα and FGF21 mRNA. It blunted glucagon-stimulated acetyl-CoA carboxylase phosphorylation, a downstream target of AMPK, and accentuated PPARα and FGF21 expression. All effects were absent in gcgr(-/-) mice. These findings demonstrate that glucagon exerts a critical regulatory role in liver to stimulate pathways linked to lipid metabolism in vivo and shows for the first time that effects of glucagon on PPARα and FGF21 expression are amplified by a physiological increase in circulating lipids.

OBJECTIVEExercise is an effective intervention to treat fatty liver. However, the mechanism(s) that underlie exercise-induced reductions in fatty liver are unclear. Here we tested the hypothesis that exercise requires hepatic glucagon action to reduce fatty liver.RESEARCH DESIGN AND METHODSC57BL/6 mice were fed high-fat diet (HFD) and assessed using magnetic resonance, biochemical, and histological techniques to establish a timeline for fatty liver development over 20 weeks. Glucagon receptor null (gcgr−/−) and wild-type (gcgr+/+) littermate mice were subsequently fed HFD to provoke moderate fatty liver and then performed either 10 or 6 weeks of running wheel or treadmill exercise, respectively.RESULTSExercise reverses progression of HFD-induced fatty liver in gcgr+/+ mice. Remarkably, such changes are absent in gcgr−/− mice, thus confirming the hypothesis that exercise-stimulated hepatic glucagon receptor activation is critical to reduce HFD-induced fatty liver.CONCLUSIONSThese findings suggest that therapies that use antagonism of hepatic glucagon action to reduce blood glucose may interfere with the ability of exercise and perhaps other interventions to positively affect fatty liver.

The present study examined the circuitry of the red nucleus of the Sprague‐Dawley rat and the freshwater pond turtle, Chrysemys picta, by using intracellular cell filling combined with anterograde tract tracing. Although both species have a well‐developed cerebellorubral system, they differ in that the red nucleus of rats receives direct input from the motor areas of the cerebral cortex, whereas turtles do not. However, a direct descending projection from the hypothalamus to the red nucleus of turtles has been described. The aim of this study was to elucidate the relative functional contributions of the cerebellum and descending inputs to motor signal generation in the red nucleus. The results show that the cellular distribution of cerebellar inputs on rubrospinal neurons is similar between the rat and turtle; these projections are observed on the soma and the proximal and distal dendrites. In contrast, the hypothalamic inputs in turtles occupy mainly the more distally located dendrites, similar to the position of the cortical inputs in rats. These findings suggest that, first, the cerebellar inputs are not spatially segregated from the cortical or hypothalamic inputs in rats or turtles, as far as can be determined by light microscopy. Second, there is specificity of input from the cortex in rats and hypothalamus in turtles onto the distal portions of the dendrites. The similarity in the organizational features of the mammalian and non‐mammalian cerebellorubrospinal systems has implications for interpretations of the relative roles of the cerebellum and cerebral cortex in motor control. J. Comp. Neurol. 416:101–111, 2000. © 2000 Wiley‐Liss, Inc.

The rat and turtle differ markedly in major structural features of the corticocerebellorubrospinal circuitry. Although both species have a well-developed cerebellorubrospinal system, they differ in that a direct cerebral cortical input to the red nucleus is present only in the rat. The aim of the present study was to compare features of the soma and dendritic morphology of rubrospinal neurons that receive cortical input, as in rats, with those that do not, as in turtles. Intracellular Lucifer Yellow injections of neurons retrogradely labeled with Fast Blue in the rat or activity-dependent sulforhodamine-labeled neurons in the turtle were used to fill rubrospinal neurons in 150-200-microm-thick fixed sections. Images of filled neurons were imported into a computer to analyze quantitatively soma and dendritic morphology. The results show that rubrospinal soma size is slightly larger in the rat than in the turtle. However, analysis of the dendritic morphology, including total dendritic length, length of primary, secondary, and tertiary dendritic branches, and a Scholl analysis of dendritic branch intersections across concentric rings, demonstrated no significant differences between the two species. These findings suggest that the basic dendritic morphology of rubrospinal neurons may have been established early in phylogeny, preceding the evolution of cortical inputs. Alternatively, similar dendritic morphologies may have arisen due to the presence of other synapses in the turtle that occupy the sites of the cortical input in the rat. This comparative approach provides insights into the information processing capabilities of cortically versus subcortically controlled motor systems.

No abstract

Evidence suggests that glucagon (GN) has persistent effects on the liver that we hypothesize are related to AMPK, PPARα, and FGF‐21. These proteins are known to promote hepatic oxidative processes. We aimed to define signaling pathways downstream of the hepatic GN receptor in vivo. The role of GN stimulation was tested using wild type (WT) and glucagon receptor null (KO) mice catheterized in a carotid artery and jugular vein 5d prior to study. In 5h fasted mice, GN (10ng/kg/min) or saline (SAL) was infused for 2h and the interaction of GN with circulating fat levels was tested in separate studies in which intralipid (IL) was also infused. Pfloridzin was infused in all studies to prevent high glucose and insulin levels. Euglycemia (150mg/dL) was maintained using a variable glucose infusion. Elevated GN (∼5‐fold) in WT increased glucose production and caused increases in AMP:ATP (7.7±0.5 fold), p‐AMPK (2.1±0.3 fold), and PPARα and FGF‐21 mRNA (2.0±0.3 and 2.0±0.4 fold) in liver compared to SAL. IL alone had no effects. However, IL led to marked potentiation of GN‐induced stimulation of FGF‐21 mRNA (12.3±4.2‐fold) in WT mice. Effects of GN with and without IL were eliminated in KO mice. We show in vivo that glucagon drops the liver to a low energy state which corresponds to increased activation of AMPK and induction of PPARα and FGF‐21 mRNA. Effects of GN on FGF‐21 are potentiated by IL. In conclusion, GN not only acutely regulates glucose homeostasis, but also causes adaptations in processes that improve the ability of the liver to oxidize fat. Supported by DK 50277.