A Mutant of Synechococcus PCC7942 Incapable of Adapting to Low CO(2) Concentration

In isolated phloem segments of celery (Apium graveolens L.), a tissue highly specific for sucrose and mannitol uptake, glucose uptake occurs at very low rates and exhibits biphasic kinetics. Nonpenetrating inhibitors such as parachloromercuribenzene sulfonic acid did not inhibit glucose uptake. However, uptake was greatly inhibited by penetrating inhibitors such as N-ethylmaleimide and carbonylcyanide-m-chlorophenyl hydrazone. Carbonylcyanide-m-chlorophenyl hydrazone inhibition of uptake was reversed by washing and addition of thiol reagents to uptake solutions. Phlorizin, a competitive inhibitor of glucose caused moderate inhibition of uptake only after 3 hours of tissue exposure. Low pH, fusicoccin, and low turgor which enhance H⁺-sugar cotransport did not alter uptake rates. Furthermore, glucose did not induce alkalinization of the uptake media. Efflux analysis indicated that the presence of 50 millimolar unlabeled glucose in the wash media enhanced exchange of the labeled glucose across the tonoplast. Results indicate that the glucose carrier is not located at the plasmalemma but appears to be present at the membrane of an intracellular compartment, most likely the tonoplast. Carrier-mediated glucose transport in this tissue is proposed to be a facilitated diffusion.     

Non-Invasive,Simultaneous Quantification of Vascular Oxygenation and Glucose Uptake in Tissue

We report the development of non-invasive, fiber-based diffuse optical spectroscopy for simultaneously quantifying vascular oxygenation (SO₂) and glucose uptake in solid tumors in vivo. Glucose uptake was measured using a fluorescent glucose analog, 2-[N-(7-nitrobenz-2-oxa-1,3-diaxol-4-yl)amino]-2-deoxyglucose (2-NBDG). Quantification of label-free SO₂ and 2-NBDG-fluorescence-based glucose uptake 60 minutes after administration of the tracer (2-NBDG₆₀) was performed using computational models of light-tissue interaction. This study was carried out on normal tissue and 4T1 and 4T07 murine mammary tumor xenografts in vivo. Injection of 2-NBDG did not cause a significant change in optical measurements of SO₂, demonstrating its suitability as a functional reporter of tumor glucose uptake. Correction of measured 2-NBDG-fluorescence for the effects of absorption and scattering significantly improved contrast between tumor and normal tissue. The 4T1 and 4T07 tumors showed significantly decreased SO₂, and 4T1 tumors demonstrated increased 2-NBDG₆₀ compared with normal tissue (60 minutes after the administration of 2-NBDG when perfusion-mediated effects have cleared). 2-NBDG-fluorescence was found to be highly sensitive to food deprivation-induced reduction in blood glucose levels, demonstrating that this endpoint is indeed sensitive to glycolytic demand. 2-NBDG₆₀ was also found to be linearly related to dose, underscoring the importance of calibrating for dose when comparing across animals or experiments. 4T1 tumors demonstrated an inverse relationship between 2-NBDG₆₀ and SO₂ that was consistent with the Pasteur effect, particularly when exposed to hypoxic gas breathing. Our results illustrate the potential of optical spectroscopy to provide valuable information about the metabolic status of tumors, with important implications for cancer prognosis.     

Effect of hyperglycemia on d-glucose transport across the brush-border and basolateral membrane of rat small intestine

Experimental hyperglycemia leads to an increase in the capacity of the rat small intestine to absorb glucose. This effect occurs within hours from the onset of hyperglycemia and is thought to involve an induction of glucose transport in the brush-border and/or basolateral membrane of the intestinal epithelium. We devised a protocol for the simultaneous preparation of brush-border vesicles and basolateral vesicles from rat small intestine to determine the locus for the inductioof glucose transporter in hyperglycemic rats. A 6 h period of intravenous infusion with a 30% glucose solution had no effect on the initial rate of glucose uptake across jejunal or ileal brush-border vesicles when measured in the absence of a Na⁺ gradient, suggesting that enhanced glucose uptake is not dependent on an increase in the number of Na⁺-dependent secondary active glucose transporters in the brush-border. Hyperglycemia did not effect the rate of glucose uptake across ileal basolateral vesicles but did cause a 78% increase in the initial rate of carrier-mediated d-glucose uptake across jejunal basolateral vesicles. The induction of glucose transport in the jejunal basolateral membrane was characterized by a rapid rate of glucose equilibration across the vesicles ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{= 46}\text{s}$ sorbitol infused controls, 18 s hyperglycemia) and a 75% increase in the V_max for carrier-mediated glucose uptake with no significant change in K_t. When the rats were pretreated with cycloheximide prior to intravenous infusion, the initial rate of d-glucose uptake dropped to 13% of that seen in jejunal basolateral vesicles prepared from untreated rats. These results suggest a rapid turnover rate for the Na⁺-independent glucose transporter in the basolateral membrane of the enterocyte. An increase in the number of functioning glucose transporters in the basolateral membrane may play an important role in the short-term induction of glucose absorption by the jejunum of the hyperglycemic animal.     

Uncoupler-Resistant Glucose Uptake by the Thermophilic Glycolytic Anaerobe Thermoanaerobacter thermosulfuricus (Clostridium thermohydrosulfuricum)

The transport of glucose across the bacterial cell membrane of Thermoanaerobacter thermosulfuricus (Clostridium thermohydrosulfuricum) Rt8.B1 was governed by a permease which did not catalyze concomitant substrate transport and phosphorylation and thus was not a phosphoenolpyruvate-dependent phosphotransferase. Glucose uptake was carrier mediated, could not be driven by an artificial membrane potential (Δψ) in the presence or absence of sodium, and was not sensitive to inhibitors which dissipate the proton motive force (Δp; tetrachlorosalicylanilide, N,N-dicyclohexylcarboiimide, and 2,4-dinitrophenol), and no uptake of the nonmetabolizable analog 2-deoxyglucose could be demonstrated. The glucokinase apparent K_m for glucose (0.21 mM) was similar to the K_t (affinity constant) for glucose uptake (0.15 mM), suggesting that glucokinase controls the rate of glucose uptake. Inhibitors of ATP synthesis (iodoacetate and sodium fluoride) also inhibited glucose uptake, and this effect was due to a reduction in the level of ATP available to glucokinase for glucose phosphorylation. These results indicated that T. thermosulfuricus Rt8.B1 lacks a concentrative uptake system for glucose and that uptake is via facilitated diffusion, followed by ATP-dependent phosphorylation by glucokinase. In T. thermosulfuricus Rt8.B1, glucose is metabolized by the Embden-Meyerhof-Parnas pathway, which yields 2 mol of ATP (G. M. Cook, unpublished data). Since only 1 mol of ATP is used to transport 1 mol of glucose, the energetics of this system are therefore similar to those found in bacteria which possess a phosphotransferase.     

Effects of ketamine on glucose uptake by glucose transporter type 3 expressed in Xenopus oocytes: The role of protein kinase C

We investigated the effects of ketamine on the type 3 facilitative glucose transporter (GLUT3), which plays a major role in glucose transport across the plasma membrane of neurons. Human-cloned GLUT3 was expressed in Xenopus oocytes by injection of GLUT3 mRNA. GLUT3-mediated glucose uptake was examined by measuring oocyte radioactivity following incubation with 2-deoxy-d-[1,2-³H]glucose. While ketamine and S(+)-ketamine significantly increased GLUT3-mediated glucose uptake, this effect was biphasic such that higher concentrations of ketamine inhibited glucose uptake. Ketamine (10 μM) significantly increased V_max but not K_m of GLUT3 for 2-deoxy-d-glucose. Although staurosporine (a protein kinase C inhibitor) increased glucose uptake, no additive or synergistic interactions were observed between staurosporine and racemic ketamine or S(+)-ketamine. Treatment with ketamine or S(+)-ketamine partially prevented GLUT3 inhibition by the protein kinase C activator phorbol-12-myrisate-13-acetate. Our results indicate that ketamine increases GLUT3 activity at clinically relevant doses through a mechanism involving PKC inhibition.     

Net hepatic release of glucose from precursor supply in ruminants: a meta-analysis

For their glucose supply, ruminants are highly dependent on the endogenous synthesis in the liver, but despite the numerous studies that evaluated hepatic glucose production, very few simultaneously measured hepatic glucose production and uptake of all precursors. As a result, the variability of precursor conversion into glucose in the liver is not known. The present study aimed at investigating by meta-analysis the relationships between hepatic glucose net release and uptake of precursors. We used the FLuxes of nutrients across Organs and tissues in Ruminant Animals database, which gathers international results on net nutrient fluxes at splanchnic level measured in catheterized animals. Response equations were developed for intakes up to 41 g DM intake/kg BW per day of diets varying from 0 to 100 g of concentrate/100 g DM in the absence of additives. The net hepatic uptake of propionate, α-amino-N and l-lactate was linearly and better related to their net portal appearance (NPA) than to their afferent hepatic flux. Blood flow data were corrected for lack of deacetylation of the para-aminohippuric acid, and this correction was shown to impact the response equations. To develop response equations between the availability of precursors (portal appearance and hepatic uptake) and net glucose hepatic release, missing data on precursor fluxes were predicted from dietary characteristics using previously developed response equations. Net hepatic release of glucose was curvilinearly related to hepatic supply and uptake of the sum of precursors, suggesting a lower conversion rate of precursors at high precursor supply. Factors of variation were explored for the linear portion of this relationship, which applied to NPA of precursors ranging from 0.99 to 9.60 mmol C/kg BW per h. Hepatic release of glucose was shown to be reduced by the portal absorption of glucose from diets containing bypass starch and to be increased by an increased uptake of β-hydroxybutyrate indicative of higher body tissue mobilization. These relationships were affected by the physiological status of the animals. In conclusion, we established equations that quantify the net release of glucose by the liver from the net availability of precursors. They provide a quantitative overview of factors regulating hepatic glucose synthesis in ruminants. These equations can be linked with the predictions of portal absorption of nutrients from intake and dietary characteristics, and provide indications of glucose synthesis from dietary characteristics.     

Sink Metabolism in Tomato Fruit : II. Phloem Unloading and Sugar Uptake

Analysis of [³H]-(fructosyl)-sucrose translocation in tomato (Lycopersicon esculentum Mill.) indicates that phloem unloading in the fruit occurs, at least in part, to the apoplast followed by extracellular hydrolysis. Apoplastic sucrose, glucose, and fructose concentrations were estimated as 1 to 7, 12 to 49, and 8 to 63 millimolar, respectively in the tomato fruit pericarp tissue. Hexose concentrations were at least four-fold greater than sucrose at all developmental stages. Short-term uptake of [¹⁴C]sucrose, -glucose, and -fructose in tomato pericarp disks showed first order kinetics over the physiologically relevant concentration range. The uptake rate of [¹⁴C]-(glucosyl)-1′-fluorosucrose was identical to the rate of [¹⁴C]sucrose uptake, suggesting sucrose may be taken up directly without prior extracellular hydrolysis. Short-term uptake of all three sugars was insensitive to 10 micromolar carbonyl cyanide m-chlorophenylhydrazone and to 10 micromolar p-chloromercuribenzene sulfonic acid. However, long-term accumulation of glucose was sensitive to carbonyl cyanide m-chlorophenylhydrazone. Together these results suggest that although sucrose is at least partially hydrolyzed in the apoplast, sucrose may enter the metabolic carbohydrate pool directly. In addition, sugar uptake across the plasma membrane does not appear to be energy dependent, suggesting that sugar accumulation in the tomato fruit is driven by subsequent intracellular metabolism and/or active uptake at the tonoplast.     

Sugar transport in isolated corn root protoplasts

Isolated corn (Zea mays L.) root protoplasts were used to study sucrose and hexose uptake. It is found that glucose was preferentially taken up by the protoplasts over sucrose and other hexoses. Glucose uptake showed a biphasic dependence on external glucose concentration with saturable (K_m of 7 millimolar) and linear components. In contrast, sucrose uptake only showed a linear kinetic curve. Sucrose and glucose uptake were linear over a minimum of 1 hour at pH 6.0 and 1 millimolar exogenous sugar concentration. Glucose uptake showed a sharp 42°C temperature optimum, while sucrose uptake showed a lower temperature sensitivity which did not reach a maximum below 50°C. Uptake of both sugars was sensitive to several metabolic inhibitors and external pH. Differences between sucrose and glucose uptake in two different sink tissue (i.e. protoplasts from corn roots and soybean cotyledons) are discussed.     

Multiphasic absorption of glucose and 3-o-methyl glucose by aged potato slices

The isotherm for glucose absorption by aged potato (Solanum tuberosum var. Russet Burbank) discs shows four distinct phases in the concentration ranges 1.0 to 75 μm, 75 μm to 1.5 mm, 1.5 to 15 mm, and 15 to 100 mm, respectively. Each segment of the multiphasic isotherm, when plotted reciprocally by the method of Lineweaver and Burk or of Hofstee, without regard for uptake in earlier phases, indicates absorption rate to be a hyperbolic function of concentration. The observations suggest that glucose uptake is carrier-mediated, and that the transport barrier undergoes a series of all-or-none transformations at critical external concentrations, yielding successive new and higher values for the parameters Km and V_max 3-O-Methyl glucose, a nonmetabolizable analogue of glucose, shows the same multiphasic absorption isotherm, with Km values essentially similar to those for glucose uptake, and V_max values somewhat lower than those for glucose absorption. Whereas the first three phases of the absorption isotherm are taken to reflect passage across the plasma membrane, the fourth phase may reflect kinetics of glucose or 3-O-methyl glucose transport to the vacuole.     

Hydrogen Peroxide-Dependent Uptake of Iodine by Marine Flavobacteriaceae Bacterium Strain C-21

The cells of the marine bacterium strain C-21, which is phylogenetically closely related to Arenibacter troitsensis, accumulate iodine in the presence of glucose and iodide (I⁻). In this study, the detailed mechanism of iodine uptake by C-21 was determined using a radioactive iodide tracer, ¹²⁵I⁻. In addition to glucose, oxygen and calcium ions were also required for the uptake of iodine. The uptake was not inhibited or was only partially inhibited by various metabolic inhibitors, whereas reducing agents and catalase strongly inhibited the uptake. When exogenous glucose oxidase was added to the cell suspension, enhanced uptake of iodine was observed. The uptake occurred even in the absence of glucose and oxygen if hydrogen peroxide was added to the cell suspension. Significant activity of glucose oxidase was found in the crude extracts of C-21, and it was located mainly in the membrane fraction. These findings indicate that hydrogen peroxide produced by glucose oxidase plays a key role in the uptake of iodine. Furthermore, enzymatic oxidation of iodide strongly stimulated iodine uptake in the absence of glucose. Based on these results, the mechanism was considered to consist of oxidation of iodide to hypoiodous acid by hydrogen peroxide, followed by passive translocation of this uncharged iodine species across the cell membrane. Interestingly, such a mechanism of iodine uptake is similar to that observed in iodine-accumulating marine algae.     

Munc18c is not rate-limiting for glucose and long-chain fatty acid uptake in the heart

The role of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)- and SNARE-associated proteins have not yet been assessed in regulation of cardiac glucose uptake, nor in the regulation of long-chain fatty acid (LCFA) uptake in any tissue. Munc18c is a SNARE-associated protein that regulates GLUT4 translocation in skeletal muscle and adipose tissue. Using cardiomyocytes from Munc18c^?/+ mice (with 56% reduction of Munc18c protein expression), we investigated whether this syntaxin4-associated protein is involved in regulation of cardiac substrate uptake. Basal, insulin- and oligomycin (a 5′ AMP-activated protein kinase-activating agent)-stimulated glucose and LCFA uptake were not altered significantly in Munc18c^?/+ cardiomyocytes compared to wild-type cells. We conclude, therefore, that Munc18c is not rate-limiting for cardiac substrate uptake, neither under basal conditions nor when maximally stimulated metabolically.     

Assessing Glucose Uptake through the Yeast Hexose Transporter 1 (Hxt1)

The transport of glucose across the plasma membrane is mediated by members of the glucose transporter family. In this study, we investigated glucose uptake through the yeast hexose transporter 1 (Hxt1) by measuring incorporation of 2-NBDG, a non-metabolizable, fluorescent glucose analog, into the yeast Saccharomyces cerevisiae. We find that 2-NBDG is not incorporated into the hxt null strain lacking all glucose transporter genes and that this defect is rescued by expression of wild type Hxt1, but not of Hxt1 with mutations at the putative glucose-binding residues, inferred from the alignment of yeast and human glucose transporter sequences. Similarly, the growth defect of the hxt null strain on glucose is fully complemented by expression of wild type Hxt1, but not of the mutant Hxt1 proteins. Thus, 2-NBDG, like glucose, is likely to be transported into the yeast cells through the glucose transport system. Hxt1 is internalized and targeted to the vacuole for degradation in response to glucose starvation. Among the mutant Hxt1 proteins, Hxt1^N370A and HXT1^W473A are resistant to such degradation. Hxt1^N370A, in particular, is able to neither uptake 2-NBDG nor restore the growth defect of the hxt null strain on glucose. These results demonstrate 2-NBDG as a fluorescent probe for glucose uptake in the yeast cells and identify N370 as a critical residue for the stability and function of Hxt1.     

Dietary melatonin supplementation alters uteroplacental amino acid flux during intrauterine growth restriction in ewes

Dietary melatonin supplementation during mid- to late-gestation increased umbilical artery blood flow and caused disproportionate fetal growth. This melatonin-induced increase in umbilical artery blood flow may alter nutrient availability to the fetus, which may lead to alterations in fetal size. The objectives of the current experiment were to determine amino acid (AA) and glucose concentrations as well as AA and glucose flux across the uteroplacenta using a mid- to late-gestation model of intrauterine growth restriction supplemented with dietary melatonin as a 2 × 2 factorial design. At day 50 of gestation, 32 ewes were supplemented with 5 mg of melatonin (MEL) or no melatonin (CON) and were allocated to receive 100% (adequate; ADQ) or 60% (restricted; RES) of nutrient requirements. On day 130 of gestation, uterine and umbilical blood flows were determined via Doppler ultrasonography during a non-survival surgery. Blood samples were collected under general anesthesia from the maternal saphenous artery, gravid uterine vein, umbilical artery, and umbilical vein for AA analysis and glucose. Total α-AA concentrations in maternal artery and gravid uterine vein were decreased (P < 0.05) in RES v. ADQ fed ewes. Maternal arterial − venous difference in total α-AA was increased (P ⩽ 0.01) in RES v. ADQ fed ewes, while total uterine α-AA flux was not different (P > 0.40) across all treatment groups. Fetal venous − arterial difference in total α-AA as well as uteroplacental flux of total α-AA were decreased (P < 0.05) in CON-RES v. CON-ADQ, and similar (P > 0.20) in MEL-RES v. CON-ADQ. Maternal concentrations and uterine flux of branched-chain AA (BCAA) were not different across all treatment groups; however, fetal uptake of BCAA was decreased (P < 0.05) in CON-RES v. CON-ADQ, and similar (P > 0.20) in MEL-RES v. CON-ADQ. Uterine uptake of glucose was not different (P ⩾ 0.08) across all treatment groups, while uteroplacental uptake of glucose was increased (P ⩽ 0.05) in RES v. ADQ ewes. In conclusion, maternal nutrient restriction increased maternal arterial − venous difference in total α-AA, while total uterine α-AA flux was unaffected by maternal nutrient restriction. Melatonin supplementation did not impact maternal serum concentrations or uterine flux of glucose or AA; however, melatonin did improve fetal BCAA uptake during maternal nutrient restriction.     

Sucrose and Glucose Uptake into Beta vulgaris Leaf Tissues : A Case for General (Apoplastic) Retrieval Systems

Concentration curves for sugar and amino acid uptake by Beta vulgaris L. leaf tissues contained both a saturable and a linear component. Similarly shaped curves were obtained for influx of sucrose, glucose, and 3-O-methyl glucose by leaf discs, whole petiole slices, petiole segments containing pith tissue only, and petiole segments containing vascular bundles, although the tissues took up the various sugars via different proportions of saturable versus linear uptake. Two millimolar p-chloromercuribenzenesulfonic acid selectively inhibited the saturable component of sucrose uptake, but had almost no effect on the linear component. Uptake of glucose and 3-O-methyl glucose remained unaffected by p-chloromercuribenzenesulfonic acid treatment. Anoxia was found to inhibit the linear component of both sucrose and 3-O-methyl glucose influx, while the saturable component remained unaffected. The linear component of sucrose uptake was also competitively inhibited by maltose, as well as being selectively promoted by certain exposures to 5 millimolar N-ethylmaleimide, 2 micrograms per milliliter cycloheximide, and high levels of mannitol acting as osmoticum. These results support the proposal that the linear component is due to a process more complex than simple, or exchange, diffusion. It would also appear that the linear transport component utilizes a separate energy source than does the saturable component of sucrose influx.

Evidence for phloem loading from the apoplast was re-examined with respect to the present findings. Saturable sucrose uptake by minor vein tissues may represent retrieval of solute from the free space, which could explain the `apoplastic loading' phenomenon.

     

VDAC1 serves as a mitochondrial binding site for hexokinase in oxidative muscles

Voltage-dependent anion channels (VDACs), also known as mitochondrial porins, are the main pathway for metabolites across the mitochondrial outer membrane and may serve as binding sites for kinases, including hexokinase. We determined that mitochondria-bound hexokinase activity is significantly reduced in oxidative muscles (heart and soleus) in vdac1^−/− mice. The activity data were supported by western blot analysis using HK2 specific antibody. To gain more insight into the physiologic mean of the results with the activity data, VDAC deficient mice were subjected to glucose tolerance testing and exercise-induced stress, each of which involves tissue glucose uptake via different mechanisms. vdac1^−/− mice exhibit impaired glucose tolerance whereas vdac3^−/− mice have normal glucose tolerance and exercise capacity. Mice lacking both VDAC1 and VDAC3 (vdac1^−/−/vdac3^−/−) have reduced exercise capacity together with impaired glucose tolerance. Therefore, we demonstrated a link between VDAC1 mediated mitochondria-bound hexokinase activity and the capacity for glucose clearance.     

Development of an in vitro technique for cytological investigations of slices of Fasciola hepatica: Evaluation by physiological criteria

In order to check the via bilityof Fasciola hepatica slices, maintained in M199 under defined physico-chemical conditions, their rates of consumption of oxygen and glucose were determined, throughout a 12-h period, using polarographic and spectrophotometric techniques. The results, when compared with similar determinations on intact animals, indicated that the slices remained as viable as incubated whole flukes, for at least 12 h. This finding supports ultrastructural evidence detailed in a previous publication. The slices were shown to consume more oxygen and less glucose than whole animals, and possible reasons for this are discussed. Studies on the effects of DNP and iodoacetate on the uptake behaviour of slices suggested that in Fasciola hepatica, as in aerobes, most of the oxygen and glucose consumed is involved with energy metabolism. Hence, oxygen and glucose uptake rates are probably valid criteria of tissue viability for the slices.     

Similar and opposite effects of dibutyryl cyclic AMP and insulin on glucose metabolism in adipose tissue

The effects of N⁶-2′-O-dibutyryl cyclic AMP on glucose metabolism and lipolysis in fragments of rat epididymal adipose tissue were studied. Measurements were made of glucose uptake, conversion of glucose carbon to CO₂ and tissue fatty acids and glyceride-glycerol, lactate production, and glycerol release. Low concentrations of dibutyryl cyclic AMP (0.1–0.5 mM) increased all parameters of glucose metabolism and inhibited glycerol release in tissue from both normally fed and fasted rats. Higher concentrations of dibutyryl cyclic AMP (3–5 mM) diminished glucose utilization and greatly accelerated lipolysis. Insulin, 50 μunits/ml, accelerated glucose metabolism in the presence of either low or high concentrations of dibutyryl cyclic AMP though the effect of insulin was greatly reduced by 3 mM dibutyryl cyclic AMP. Tissue exposed to concentrations of dibutyryl cyclic AMP which inhibited glucose metabolism (5 mM), then rinsed and reincubated without dibutyryl cyclic AMP, displayed increased glucose utilization. The results of these experiments emphasize the need for caution in interpretation of the effects of dibutyryl cyclic AMP on adipose tissue metabolism and the need for further research to elucidate the role of cyclic AMP in the regulation of glucose metabolism.     

Ghrelin is a signal to facilitate the utilization of fatty acids and save glucose by the liver,skeletal muscle,and adipose tissues in chicks

Ghrelin, classically known as a central appetite-stimulating hormone, has recently been recognized to play an important role in peripheral tissue energy metabolism. In chicken, contrary to mammal, ghrelin acts as an anorexia signal, increased by fasting and further elevated after refed. In the present study, the effect of ghrelin on glucose/lipid utilization by peripheral tissues was investigated. Injection of exogenous acyl ghrelin reduced plasma triglyceride and glucose levels of chickens at both fasting and fed status. In the in vitro cultured chicken primary hepatocytes, adipocytes, and myoblasts, ghrelin suppressed glucose uptake, stimulated fatty acids uptake and oxidation, and decreased TG content. In hepatocyte, ghrelin increased the activities of LPL and HL, and upregulated the expression levels of gene ACC, CPT1, and PPARα. Ghrelin treatment markedly increased the protein level of p-ACC, PPARγ, PGC1α, and CPT1 in hepatocytes, adipocytes and myoblasts. Inhibition of AMPK activity by Compound C had no influence on glucose uptake by hepatocyte, adipocyte, and myoblast, but further amplified the stimulated fatty acid uptake of adipocyte by ghrelin. The present result demonstrates that ghrelin facilitates the uptake and oxidation of fatty acid and cut down the utilization of glucose by the liver, muscle, and adipose tissues. The result suggests that ghrelin functions as a signal of fatty acid oxidation. The study provides a vital framework for understanding the intrinsic role of ghrelin as a crucial factor in the concerted regulation of metabolic substrate of hepatocytes, adipocytes, and myoblasts.     

The regulation of sugar uptake and accumulation in bean pod tissue

The identity, localization and physiological significance of enzymes involved in sugar uptake and accumulation were determined for endocarp tissue of pods of Kentucky Wonder pole beans (Phaseolus vulgaris). An intracellular, alkaline invertase (pH optimum, 8) was assayed in extracted protein, as well as enzymes involved in sucrose synthesis, namely, uridinediphosphate (UDP-glucose pyrophosphorylase and UDP-glucose-fructose transglucosylase). Indirect evidence indicated the presence also of hexokinase, phosphohexoseisomerase and phosphoglucomutase. The data suggested that sucrose synthesis occurred in the cytoplasm, and that both sugar storage and an alkaline invertase occurred in the vacuole. The latter functions to hydrolyze accumulated sucrose. An outer space invertase (pH optimum, 4.0) was detected, but was variable in occurrence. Although its activity at the cell surface enhanced sucrose uptake, sucrose may be taken up unaltered.

Over a wide range of concentrations of exogenous glucose the sucrose/reducing sugar ratio of accumulated sugars remained unchanged at about 20. Synthesis of sucrose appears to be requisite to initial accumulation from glucose or fructose, as free hexoses do not increase at the apparent saturating concentration for uptake. Sucrose accumulation from exogenous hexose represents a steady-state value, in which sucrose is transported across the tonoplast into the vacuole at a rate equivalent to its rate of synthesis. Evidence indicates that this component of the accumulation process involves active transport of sucrose against a concentration gradient. The ratio of sucrose/reducing sugars in the accumulated sugars immediately after a period of uptake was inversely related to the level of inner space invertase. Within 16 hours after a period of accumulation, practically all of the sugar occurs as glucose and fructose.

The absence of competition among hexoses and sucrose indicated that a common carrier was not involved in their uptake. From a series of studies on the kinetics of uptake of glucose and fructose, including competition studies, the effects of inhibitors, radioactive assay of accumulated sugars and the distribution of label in accumulated sucrose it appeared that rate limitation for glucose or fructose uptake resides in the sequence of reactions leading to sucrose synthesis, rather than in a process mediated by a carrier protein.

     

Lysophosphatidylserine regulates blood glucose by enhancing glucose transport in myotubes and adipocytes

Lysophosphatidylserine (LPS) is known to have diverse cellular effects, but although LPS is present in many biological fluids, its in vivo effects have not been elucidated. In the present study, we investigated the effects of LPS on glucose metabolism in vivo, and how skeletal muscle cells respond to LPS stimulation. LPS enhanced glucose uptake in a dose- and time-dependent manner in L6 GLUT4myc myotubes, and this effect of LPS on glucose uptake was mediated by a Gα_i and PI 3-kinase dependent signal pathway. LPS increased the level of GLUT4 on the cell surface of L6 GLUT4myc myotubes, and enhanced glucose uptake in 3T3-L1 adipocytes. In line with its cellular functions, LPS lowered blood glucose levels in normal mice, while leaving insulin secretion unaffected. LPS also had a glucose-lowering effect in STZ-treated type 1 diabetic mice and in obese db/db type 2 diabetic mice. This study shows that LPS-stimulated glucose transport both in skeletal muscle cells and adipocytes, and significantly lowered blood glucose levels both in type 1 and 2 diabetic mice. Our results suggest that LPS is involved in the regulation of glucose homeostasis in skeletal muscle and adipose tissue.