Jean-Michel Weber

Leptin: an essential regulator of lipid metabolism

This paper reviews the general mechanisms by which leptin acts as a regulator of lipid reserves through changes in food intake, energy expenditure and fuel selection, with an emphasis on its direct effects on cellular lipid metabolism. Briefly, when leptin levels increase, food consumption decreases via modulation of hypothalamic neuropeptides. As well, normal decreases in energy expenditures (e.g. with diurnal cycles or reduced caloric intake) do not occur. This is probably caused by an increase in mitochondrial proton leak mediated by leptin via increases in sympathetic nervous system stimulation and thyroid hormone release. The decrease in caloric input coupled with relatively higher energy expenditure, therefore, leads to negative energy balance. Leptin also changes the fuel source from which ATP is generated. Fuel preference switches from carbohydrate (glucose) to lipid (fatty acids). This effect arises through stimulation of triacylglycerol catabolism by leptin. In vitro studies show that leptin is a potent stimulator of lipolysis and fatty acid oxidation in adipocytes and other cell types. Consequently, leptin is also a regulator of cellular triacylglycerol content. Hormonal regulation of leptin, as well as its role in fasting and seasonal weight gain and energy expenditure are also briefly discussed.

Metabolic fuels: regulating fluxes to select mix

Summary Animals must regulate the fluxes of multiple fuels to support changing metabolic rates that result from variation in physiological circumstances. The aim of fuel selection strategies is to exploit the advantages of individual substrates while minimizing the impact of disadvantages. All exercising mammals share a general pattern of fuel selection: at the same they oxidize the same ratio of lipids to carbohydrates. However, highly aerobic species rely more on intramuscular fuels because energy supply from the circulation is constrained by trans-sarcolemmal transfer. Fuel selection is performed by recruiting different muscles, different fibers within the same muscles or different pathways within the same fibers. Electromyographic analyses show that shivering humans can modulate carbohydrate oxidation either through the selective recruitment of type II fibers within the same muscles or by regulating pathway recruitment within type I fibers. The selection patterns of shivering and exercise are different: at the same , a muscle producing only heat (shivering) or significant movement (exercise) strikes a different balance between lipid and carbohydrate oxidation. Long-distance migrants provide an excellent model to characterize how to increase maximal substrate fluxes. High lipid fluxes are achieved through the coordinated upregulation of mobilization, transport and oxidation by activating enzymes, lipid-solubilizing proteins and membrane transporters. These endurance athletes support record lipolytic rates in adipocytes, use lipoprotein shuttles to accelerate transport and show increased capacity for lipid oxidation in muscle mitochondria. Some migrant birds use dietary omega-3 fatty acids as performance-enhancing agents to boost their ability to process lipids. These dietary fatty acids become incorporated in membrane phospholipids and bind to peroxisome proliferator-activated receptors to activate membrane proteins and modify gene expression.

Changes in the fatty acid composition of phospholipids in tissues of farmed sea bass (Dicentrarchus labrax) during an annual cycle. Roles of environmental temperature and salinity.

We quantified seasonal effects on fatty acid composition of tissue phospholipids in farmed sea bass. Major changes in percent phosphatidylethanolamine and phosphatidylcholine were observed in all tissues between February and March, and the phosphatidylcholine/phosphatidylethanolamine ratio was drastically reduced at this time. Different changes in the fatty acid composition of total phospholipids were observed in all tissues examined. Fish fed all year on the same commercial diet showed a significant correlation between water salinity and percentage of 22:6n-3 in muscle, liver and gill phospholipids, but no correlation was found between percent 22:6n-3 of phospholipids and water temperature. In each tissue, we observed annual variation in the 20:5n-3/20:4n-6 ratio in phospholipids, but maximum and minimum values occurred at different times in each organ. From these results, we conclude that salinity can play a significant role in modulating the activities of enzymes acting on lipid metabolism during their natural circannual cycles.

Relationship between n-3 PUFA content and energy metabolism in the flight muscles of a migrating shorebird: evidence for natural doping.

SUMMARY During their fall migration from the Arctic to South America, semipalmated sandpipers Calidris pusilla stop in the Bay of Fundy (east coast of Canada) before flying non-stop for ∼4500 km across the ocean. Refueling birds double their body mass by feeding on Corophium volutator, an amphipod containing high amounts of n-3 polyunsaturated fatty acids (n-3 PUFA), particularly eicosapentaenoic (20:5) and docosahexaenoic acid (22:6). In mammals, high dietary intake of n-3 PUFA is known to increase capacity for oxidative metabolism. Therefore, we hypothesized that tissue incorporation of n-3 PUFA would be associated with increases in the activity of key muscle enzymes to upregulate energy metabolism for prolonged exercise. Birds were collected at various stages of fat loading to monitor changes in lipid composition and flight muscle enzymes simultaneously. Enzymes were measured to assess oxidative capacity [citrate synthase (CS)],β -oxidation [carnitine palmitoyl transferase (CPT) and 3-hydroxyacyl dehydrogenase (HOAD)] and glycolytic capacity [lactate dehydrogenase (LDH)]. Changes in the fatty acid composition of muscle membranes (phospholipids) and fuel reserves (neutral lipids) were measured separately to distinguish between membrane-related and systemic effects of n-3 PUFA. Results show that muscle CS and HOAD are stimulated during refueling and that their activities are correlated with n-3 PUFA content in phospholipids (22:6 for CS, 20:5 for HOAD) and in neutral lipids (20:5 for CS). This suggests that 20:5 and 22:6 have different effects on energy metabolism and that they act via changes in membrane structure and systemic mechanisms. CPT and LDH did not change during refueling, but LDH activity was significantly related to the n-3 PUFA content of fuel reserves. This study shows that oxidative capacity increases rapidly during refueling and supports the idea that dietary n-3 PUFA are used as molecular signals to prime flight muscles of some long-distance migrants for extreme exercise.

Pathways for metabolic fuels and oxygen in high performance fish

Abstract This paper gives an overview of oxidative fuel metabolism in swimming fish, and known or potential modifications occurring in high-performance species are explored. Carbohydrate catabolism is the only source of ATP for sprint swimming where locomotory muscles operate as closed systems. In contrast, this substrate only plays a very minor role in prolonged swimming. Glucose fluxes have been measured in vivo in several species, but mainly at rest and with somewhat questionable methodologies. High-performance species may be able to sustain higher maximal glucose fluxes that their sedentary counterparts by: a) upregulating gluconeogenesis, b) increasing glucose transporter density or V max of individual transporters, c) storing larger amounts of glycogen in liver and muscle, and d) increasing muscle hexokinase activity. Even though lipids represent a much more important source of energy for sustained swimming, their fluxes have not been measured in vivo , even at rest, probably because of their diversity and complex chemistry. Except for elasmobranchs who do not possess plasma proteins for lipid transport, high-performance fish should be able to sustain high maximal lipid fluxes by: a) elevating lipolytic capacity, b) increasing rates of circulatory lipid transport through modified plasma proteins, c) augmenting intramuscular lipid reserves, and d) upregulating capacity for lipid oxidation in locomotory muscle mitochondria. The quantitative assessment of amino acid oxidation in swimming fish is a priority for future research because protein is probably a dominant metabolic fuel in most swimming fish. Finally, we predict that high-performance species should use proportionately more proteins/lipids and less carbohydrates than low-aerobic fish. Also, and similarly to endurance-adapted mammals, high-performance fish should increase their relative reliance on intramuscular fuel reserves and decrease their relative use of circulatory fuels.

Partitioning oxidative fuels during cold exposure in humans: muscle glycogen becomes dominant as shivering intensifies

The effects of changes in shivering intensity on the relative contributions of plasma glucose, muscle glycogen, lipids and proteins to total heat production are unclear in humans. The goals of this study were: (1) to determine whether plasma glucose starts playing a more prominent role as shivering intensifies, (2) to quantify overall changes in fuel use in relation to the severity of cold exposure, and (3) to establish whether the fuel selection pattern of shivering is different from the classic fuel selection pattern of exercise. Using a combination of indirect calorimetry and stable isotope methodology, fuel metabolism was monitored in non‐acclimatized adult men exposed for 90 mins to 10°C (low‐intensity shivering (L)) or 5°C (moderate‐intensity shivering (M)). Results show that plasma glucose oxidation is strongly stimulated by moderate shivering (+122% from L to M), but the relative contribution of this pathway to total heat generation always remains minor (< 15% of total heat production). Instead, muscle glycogen is responsible for most of the increase in heat production between L and M. By itself, the increase in CHO oxidation is responsible for the 100 W increase in metabolic rate observed between L and M, because rates of lipid and protein oxidation remain constant. This high reliance on CHO is not compatible with the well known fuel selection pattern of exercise, when considering the relatively low metabolic rates elicited by shivering (∼30% for M). We conclude that shivering and exercise of similar energy requirements appear to be supported by different fuel mixtures. Investigating the physiological mechanisms underlying why a muscle producing only heat (shivering), or significant movement (exercise), shows a different pattern of fuel selection at the same power output strikes us as a fascinating area for future research.

Fuel selection during intense shivering in humans: EMG pattern reflects carbohydrate oxidation

The thermogenic response of humans depends critically on the coordination of muscle fibre recruitment and oxidative fuel metabolism. The primary goal of this study was to determine whether the electromyographic (EMG) pattern of muscle recruitment could provide metabolic information on oxidative fuel selection during high‐intensity shivering. EMG activity (of 8 large muscles) and fuel metabolism were monitored simultaneously in non‐acclimatized adult men during high‐intensity shivering. Even though acute cold exposure elicited similar changes in metabolic rate among subjects, lipid and carbohydrate use was very different. Depending on the subject, the cold‐induced increase in carbohydrate (CHO) oxidation ranged between 2‐ and 8‐fold, with CHO accounting for 33–78% of total heat production , and lipids for 14–60%. This high variability in fuel selection was primarily explained by differences in ‘burst shivering’ rate, indicating that the recruitment of type II fibres plays a key role in orchestrating fuel selection. This study is the first to show that the pattern of muscle recruitment can provide quantitative information on energy metabolism. Future work should focus on the study of shivering bursts that may provide essential clues on what limits human survival in the cold.

Plasma and muscle phospholipids are involved in the metabolic response to long-distance migration in a shorebird.

Abstract. We studied: (1) concentrations and fatty acid compositions of plasma non-esterified fatty acids, neutral lipids, and phospholipids, and (2) fatty acid composition of flight muscle phospholipids in wintering, premigratory, and spring and fall migrating western sandpipers (Calidris mauri). Plasma neutral lipid and phospholipid levels were elevated in migrants, reflecting high rates of fat deposition. An important role of phospholipids in fattening is suggested by the fact that the amount of fatty acids in plasma phospholipids was similar to, or in spring as much as twice, that of neutral lipids. Changes in the ratio of plasma neutral lipids to phospholipids may indicate seasonal changes in triacylglycerol stores of invertebrate prey. Monounsaturation and total unsaturation of plasma neutral lipids and phospholipids increased during migration. Muscle phospholipids were more monounsaturated in spring and fall, but total unsaturation was reduced in fall. Arachidonic acid [20:4(n-6)] was especially abundant in muscle phospholipids in winter (29%) and declined during migration (19–22%), contributing to a decline in the ratio of n-6 to n-3 fatty acids. The abundance of plasma phospholipids and variability of neutral lipid to phospholipid ratio indicates that measurement of plasma phospholipids will improve methods for assessment of fattening rates of birds. The functional significance of changes in muscle phospholipids is unclear, but may relate to depletion of essential n-6 fatty acids during exercise.

Mimicking the natural doping of migrant sandpipers in sedentary quails: effects of dietary n-3 fatty acids on muscle membranes and PPAR expression

SUMMARY Wild semipalmated sandpipers (Calidris pusilla) eat n-3 fatty acids to prime their muscles for long migrations. Sedentary bobwhite quails (Colinus virginianus) were used as a model to investigate the mechanisms for this natural doping. Our goal was to characterize the stimulating effects of n-3 eicosapentaenoic acid (EPA) and n-3 docosahexaenoic acid (DHA) on oxidative capacity. Mechanisms linked to changes in membrane composition and in gene expression for peroxisome proliferator-activated receptors (PPAR) were investigated. Dietary n-3 fatty acids stimulated the activities of oxidative enzymes by 58–90% (citrate synthase, cytochrome oxidase, carnitine palmitoyl transferase and hydroxyacyl dehydrogenase), and sedentary quails showed the same changes in membrane composition as sandpipers preparing for migration. EPA and DHA have the same doping effect. The substitution of n-6 arachidonic acid by n-3 EPA in membrane phospholipids plays an important role in mediating the metabolic effects of the diet, but results provide no significant support for the involvement of PPARs (as determined by changes in gene expression). The fatty acid composition of mitochondrial membranes and sarcoplasmic reticulum can be monitored by measuring total muscle phospholipids because all phospholipids are equally affected by diet. Only extreme regimes of endurance training can lead to increments in oxidative capacity matching those induced here by diet. As they prepare for long migrations, semipalmated sandpipers improve their physical fitness by eating! Choosing n-3 fatty acid doping over endurance training strikes us as a better strategy to boost aerobic capacity when rapid storage of energy is critical.

Effects of hypoxia and low temperature on substrate fluxes in fish: plasma metabolite concentrations are misleading

Oxygen levels and temperature can fluctuate rapidly in aquatic environments. Even though the effects of environmental stresses on fish metabolism have been studied extensively, information on fuel kinetics is extremely limited because it relies almost exclusively on changes in substrate concentrations. The turnover rate of nonesterified fatty acids (NEFA) has never been measured in fish. Therefore, our goal was to quantify glucose and NEFA fluxes in rainbow trout acutely exposed to severe hypoxia (25% O2 saturation) or low temperature (6°C for fish acclimated to 15°C) by performing continuous infusions of 6-[3H]glucose and 1-[14C]palmitate in vivo. Results show that hypoxia causes a 53% decrease in NEFA turnover rate, together with a transient increase in hepatic glucose production, whereas a rapid drop in temperature induces equivalent declines in glucose, NEFA, and oxygen fluxes [temperature coefficient ≅ 2]. More importantly, kinetic changes in glucose and NEFA fluxes are not accompanied by interpretable changes in the plasma concentrations of these metabolites. Thus using concentration changes to draw conclusions about fluxes must be avoided.