Energy-rich phosphates and transintestinal transport in rat intestine incubated in vitro at different temperatures.

In the present work, the transported fluid and the tissue content of ATP, ADP and AMP has been evaluated in the jejunum rat intestine which was everted and incubated in vitro both at 28 degrees C and at 38 degrees C for 1 h. The energy-rich phosphates have been measured in the tissue at the beginning and at the end of the experiment as well as in vivo. These determinations have been made in the total intestine and in the scraped mucosa. ATP and ADP content are higher in vivo and lower but constant at 28 degrees C in vitro; on the contrary, at 38 degrees C in vitro, the initial and final content of these adenilic nucleotides are both lower than at 28 degrees C. Under all these conditions the AMP content does not vary appreciably. Wet weight to dry weight ratios ahve been reported for mucosal and submucosal tissues in unincubated and incubated intestines. In some experiments, fluid transport (measured as an actual serosal volume increase) was determined every 20 min during a 1-h incubation. At 28 degrees C, fluid transport is constant throughout the time of the experiment, but at 38 degrees C, there is a progressive decrease of the transported fluid. Fluid transport and ATP content of the intestine seem to be directly related. The transport activity which is lower at 38 degrees C than at 28 degrees C, seems to be due to a low availability of energy-rich phosphates.     

The rat and hamster jejunum during transintestinal transport in vitro.

By using the jejunal tract of rat and hamster intestine, net fluid transintestinal transport in the everted sac incubated in vitro at 28 degrees C and at 38 degrees C has been determined. In the scraped mucosa wet weight/dry weight ratio, cell water, sodium and potassium concentration have been detected in vivo and in vitro, throughout the incubation time, at the two different temperatures. Under all these conditions ATP, ADP and AMP levels have been determined in total intestine and in scraped mucosa. In rat jejunum incubated in vitro at 38 degrees C transintestinal fluid transport continuously decreases during 1-h experiment; in the same time the enterocyte gains sodium, dilutes cell potassium and swells, whereas its energy charge is dramatically reduced. All these parameters are constant in rat jejunum incubated at 28 degrees C and in the hamster incubated both at 28 degrees C and at 38 degrees C throughout the experiment. An inadequate diffusion of oxygen into the enterocyte, could tentatively explain experimental results obtained on rat jejunum at 38 degrees C. Under all tested conditions, energy charge and intracellular potassium are lower in conditions in vitro than in those in vivo; the contrary happens for cell sodium and swelling. The oxygenation of the intestine in vitro, lower than the one in vivo, could explain the different behaviour found in the two experimental conditions.     

Transport of AMP by Rickettsia prowazekii.

Rickettsia prowazekii possesses an exchange transport system for AMP. Chromatographic analysis of the rickettsiae demonstrated that transported AMP appeared intracellularly as AMP, ADP, and ATP, and no hydrolytic products appeared in either the intracellular or extracellular compartments. The phosphorylation of AMP to ADP and ATP was prevented by pretreatment of the cells with 1 mM N-ethylmaleimide without inhibiting the transport of AMP. Although no efflux was demonstrable in the absence of nucleotide in the medium, the intracellular adenine nucleotide pool could be exchanged with external unlabeled adenine nucleotides. Both ADP and ATP were as effective as AMP at inhibiting the uptake of [3H]AMP. Although this transport system was inhibited by low temperature (0 degrees C) and partially inhibited by the protonophore carbonyl cyanide-m-chlorophenyl hydrazone (1 mM), it was relatively insensitive to KCN (1 mM). The uptake of AMP at 34 degrees C had an apparent Kt for influx of 0.4 mM and a Vmax of 354 pmol min-1 per mg. At 0 degrees C there was a very rapid and unsaturable association of AMP with these organisms. Correction of the uptake data at 34 degrees C for the 0 degrees C component lowered the apparent Kt to 0.15 mM. Both magnesium and phosphate ions are required for optimal transport activity. Chemical measurements of the total intracellular nucleotide pools demonstrated that this system was not a net adenine nucleotide transport system, but that uptake of AMP was the result of an exchange with internal adenine nucleotides.     

In vitro adenine nucleotide catabolism in African catfish spermatozoa

It has been shown recently that African catfish (Clarias gariepinus) spermatozoa possess relatively low ATP content and low adenylate energy charge (AEC). One of the possible explanations for this phenomenon is that the spermatozoa actively catabolize adenine nucleotides. A relatively high rate of such catabolism could then contribute to the low ATP concentration and low adenylate energy charge observed in the spermatozoa in vitro. To check this hypothesis, we investigated ATP content and adenine nucleotide catabolism in African catfish spermatozoa stored at 4 °C in the presence of glycine as an energetic substrate. Our results indicate that the storage of African catfish sperm at 4 °C in the presence of glycine causes time-dependent ATP depletion. In contrast to ATP, the AMP content increases significantly during the same period of sperm storage, while the ADP increases only slightly. Moreover, a significant increase of inosine and hypoxanthine content was also found. Hypoxanthine was accumulated in the storage medium, but xanthine was found neither in spermatozoa nor in the storage medium. It indicates that hypoxanthine is not converted to xanthine, probably due to lack of xanthine oxidase activity in catfish spermatozoa. Present results suggest that adenine nucleotides may be converted to hypoxanthine according to the following pathway: ATP→ADP→AMP (adenosine/IMP)→inosine→hypoxanthine. Moreover, hypoxanthine seems to be the end product of adenine nucleotide catabolism in African catfish spermatozoa. In conclusion, our results suggest that a relatively high rate of adenine nucleotide catabolism contributes to the low ATP concentration and low adenylate energy charge observed in African catfish spermatozoa in vitro.     

The relation of sodium and potassium ion transport to the respiration and adenine nucleotide content of liver slices treated with inhibitors of respiration

1. The dependence of the net transport of Na(+) and K(+) by rat liver on the respiration has been determined by incubating slices in the presence of varying concentrations of respiratory inhibitors. 2. Neither the rate of net transport nor the total amount of each ion transported was inhibited unless the rate of endogenous respiration was decreased below a critical value of about 330mmol of O(2)/h per kg of protein (i.e. 50% of the total endogenous respiration). 3. The uninhibited rate of respiration could be varied over a twofold range (380-770mmol of O(2)/h per kg of protein) by the use of different substrates, but the critical value for the onset of transport inhibition was quite constant (290-360mmol/h per kg of protein) under these different conditions. 4. Slices incubated at 38 degrees C without inhibitors showed an increase of their ATP content and the concentration ratio ATP/ADP. The final ATP content and concentration ratio, ATP/ADP, of slices treated with different concentrations of inhibitors were closely related to the rate of respiration. 5. The increased ATP content of the control slices during incubation was equal to the increase of total adenine nucleotides. At increasing degrees of respiratory inhibition the relative contributions of ADP and AMP to the total adenine nucleotide content increased. 6. The critical rate of respiration for the onset of inhibition of ion transport and the corresponding contents of adenine nucleotides provide estimates of the maximal values of certain parameters of energy metabolism required for the support of alkali-cation transport in the liver slices.     

Adenine nucleotide and lysine transport in Chlamydia psittaci.

Isolated reticulate bodies of Chlamydia psittaci were found to transport ATP and ADP by an ATP-ADP exchange mechanism. ATP uptake activity was not detected in elementary bodies. The apparent Km of transport for both ATP and ADP was approximately 5 microM, and the calculated Vmax for both was about 1 nmol of nucleotide transported per min per mg of protein. ADP competitively inhibited ATP transport with a Ki of 4.5 microM. Other nucleotides tested had no effect on the uptake of ATP. A magnesium-dependent, oligomycin-sensitive ATPase (ATP phosphohydrolase, EC 3.6.1.3) was associated with reticulate bodies, and most of the transported ATP was hydrolyzed to ADP, which was exchanged for additional, extracellular nucleotide. Some ADP was hydrolyzed to AMP, which exited the cells slowly. Lysine was transported against the electrochemical gradient by reticulate bodies in the presence of ATP. Oligomycin and carbonyl cyanide p-trifluoromethoxyphenylhydrazone inhibited ATP-dependent lysine transport. Lysine exited reticulate bodies when the reticulate bodies were incubated in the presence of ADP, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, or a reduced concentration of ATP. The results support the concept that chlamydiae are energy parasites which are capable of drawing upon the adenine nucleotides of their hosts, hydrolyzing ATP, and establishing an energized membrane.     

Ouabain-insensitive transintestinal transport in the rat jejunum incubated in vitro

The jejunal tract of rat intestine, everted and incubated in vitro at 28 degrees C for 2 hr in Krebs-Ringer bicarbonate solution, was used to test the existence of a ouabain-insensitive sodium pump. Cell water, Na, and K together with Na, fluid, K, and lactate transported into the serosal compartment were determined and, under control conditions, the tested parameters were found constant in time. By blocking the Na-K pump with 20 mM ouabain in the serosal compartment, the enterocyte lost K and gained Na, but the cell volume did not vary. Moreover, the transport of Na, fluid, and lactate, although lower, was constant for 2 hr. When ethacrynate was added or when the ATP supply was blocked by adding 2,4-dinitrophenol plus iodoacetate, the cell swelled and the transport of Na and fluid stopped. These results are interpreted as suggesting the existence of a ouabain-insensitive Na pump, in addition to the well-known Na-K pump.     

Molecular recognition of nucleotides by means of ionic interaction in hydrophobic media

Adenosine phosphates, AMP, ADP and ATP were found to be recognized and extracted from aqueous to an organic phase by the newly prepared lipophilic diammonium salt, N,N'-distearyldiammonium dichloride of 1,4-diazabicyclo [2.2.2] octane 1. ADP and ATP were specifically bound by 1 under the condition of no appreciable binding of AMP. The conventional phase transfer reagent, trioctylmethylammonium chloride 2 was far less effective and lacked selectivity fort the binding of adenosine phosphates. This diammonium salt was used as a specific carrier of ADP in the passive transport through a liquid membrane. A high selectivity was observed in the transport rate of ADP relative to that of AMP.     

Mechanism of action of ATP on intestinal epithelial cells. Cyclic AMP-mediated stimulation of active ion transport

ATP, ADP and AMP but not adenosine increased cyclic AMP in dispersed enterocytes prepared from guinea pig small intestine. This action of ATP was augmented by IBMX and was reproduced by App(NH)p or App(CH2)p. ATP also increased the formation of cyclic [14C]AMP in enterocytes that had been preincubated with [14C]adenine. Gpp(NH)p and NaF each caused persistent activation of adenylate cyclase in plasma membranes from enterocytes and ATP caused significant augmentation of this persistent activation. In addition to increasing cellular cyclic AMP and augmenting Gpp(NH)p and NaF-stimulated persistent activation of adenylate cyclase, ATP increased the Isc across mounted strips of small intestine and inhibited net absorption of fluid and electrolytes in segments of everted small intestine. These results indicate that intestinal epithelial cells possess a receptor that interacts with ATP and other adenine nucleotides and that receptor occupation by ATP causes activation of adenylate cyclase, increased cyclic AMP and changes in active ion transport across intestinal mucosa.     

Mechanism of action of ATP on intestinal epithelial cells: Cyclic AMP-mediated stimulation of active ion transport

ATP, ADP and AMP but not adenosine increased cyclic AMP in dispersed enterocytes prepared from guinea pig small intestine. This action of ATP was augmented by IBMX and was reproduced by App(NH)p or App(CH₂)p. ATP also increased the formation of cyclic [¹⁴C]AMP in enterocytes that had been preincubated with [¹⁴C]adenine. Gpp(NH)p and NaF each caused persistent activation of adenylate cyclase in plasma membranes from enterocytes and ATP caused significant augmentation of this persistent activation. In addition to increasing cellular cyclic AMP and agumenting Gpp(NH)p and NaF-stimulated persistent activation of adenylate cyclase, ATP increased the I_sc across mounted strips of small intestine and inhibited net absorption of fluid and electrolytes in segments of everted small intestine. These results indicate that intestinal epithelial cells possess a receptor that interacts with ATP and other adenine nucleotides and that receptor occupation by ATP causes activation of adenylate cyclase, increased cyclic AMP and changes in active ion transport across intestinal mucosa.     

Adenosine Phosphates in Germinating Radish (Raphanus sativus L.) Seeds

Changes in concentrations of adenosine phosphates (AMP, ADP, and ATP), oxygen utilization, and fresh weights were measured during the first 48 hours after imbibition of water by quiescent radish seeds (Raphanus sativus L.) at 22.5 C. The changes in ATP concentrations, oxygen utilization, and fresh weights followed a triphasic time course, characterized by a rapid initial increase, which extended from 0 to approximately 1.5 hours, a lag phase from 1.5 to 16 hours, and a sharp linear increase from 16 to 48 hours. In unimbibed seeds, the concentrations of ATP, ADP, and AMP were <0.1, 0.9, and 2.2 nmoles/seed, respectively. After imbibition of water by the quiescent seeds, for 1 hour, the ATP concentration had increased to 2.5, and ADP and AMP concentrations had decreased to 0.3 and 0.1 nmole/seed, respectively. These early changes occurred also in seeds maintained under anaerobic conditions (argon), or when treated with either 5 mm fluoroacetate, or 5 mm iodoacetate. The concentrations of ADP and AMP did not change significantly from 1 to 48 hours. The termination of the lag phase at 16 hours correlated with radicle emergence. Cell division in the radicles was initiated at approximately 28 hours. ATP concentrations in seeds maintained under argon or treated with fluoroacetate remained relatively constant from approximately 2 to 48 hours. In contrast, the ATP concentration of iodoacetate-treated seeds decreased curvilinearly from 4 to 48 hours. Oxidative phosphorylation was estimated to have contributed 15, 20, and 65% of the pool ATP at 1.5, 16, and 48 hours, respectively.     

Activation of the Alternative Oxidase of Yarrowia lipolytica by Adenosine Monophosphate

The study of the effect of nucleoside phosphates on the activity of cyanide-resistant oxidase in the mitochondria and the submitochondrial particles of Yarrowia lipolytica showed that adenosine-5-monophosphate (AMP) did not stimulate the respiration of the intact mitochondria. The incubation of the mitochondria at room temperature (25°) for 3–5 h or their treatment with ultrasound, phospholipase A, and the detergent Triton X-100 at a low temperature inactivated the cyanide-resistant alternative oxidase. The inactivated alternative oxidase could be reactivated by AMP. The reactivating effect of AMP was enhanced by azolectin. Some other nucleoside phosphates also showed reactivating ability, in the following descending order: AMP = GMP > GDP > GTP > XMP > IMP. The apparent reaction rate constant K _m for AMP upon the reactivation of the alternative oxidase of mitochondria treated with Triton X-100 or incubated at 25°C was 12.5 and 20 M, respectively. The K _m for AMP upon the reactivation of the alternative oxidase of submitochondrial particles was 15 M. During the incubation of yeast cells under conditions promoting the development of alternative oxidase, the content of adenine nucleotides (AMP, ADP, and ATP) in the cells and their respiration tended to decrease. The subsequent addition of cyanide to the cells activated their respiration, diminished the intracellular content of ATP by three times, and augmented the content of AMP by five times. These data suggest that the stimulation of cell respiration by cyanide may be due to the activation of alternative oxidase by AMP.     

Activation of the alternative oxidase of Yarrowia lipolytica by adenosine 5'-monophosphate

The study of the effect of nucleoside phosphates on the activity of cyanide-resistant oxidase in the mitochondria and the submitochondrial particles of Yarrowia lipolytica showed that adenosine monophosphate (5'-AMP, AMP) did not stimulate the respiration of the intact mitochondria. The incubation of the mitochondria at room temperature (25 degrees C) for 3-5 h or their treatment with ultrasound, phospholipase A, and detergent Triton X-100 at a low temperature inactivated the cyanide-resistant alternative oxidase. The inactivated alternative oxidase could be reactivated by AMP. The reactivating effect of AMP was enhanced by azolectin. Some other nucleoside phosphates also showed reactivating ability in the following descending order. AMP = GMP > GDP > GTP > XMP > IMP. The apparent reaction rate constant Km for AMP upon the reactivation of the alternative oxidase of mitochondria treated with Triton X-100 or incubated at 25 degrees C was 12.5 and 20 microM, respectively. The Km for AMP upon the reactivation of the alternative oxidase of submitochondrial particles was 15 microM. During the incubation of yeast cells under conditions promoting the development of alternative oxidase, the content of adenine nucleotides (AMP, ADP, and ATP) in the cells and their respiration tended to decrease. The subsequent addition of cyanide to the cells activated their respiration, diminished the intracellular content of ATP three times, and augmented the content of AMP five times. These data suggest that the stimulation of cell respiration by cyanide may be due to the activation of alternative oxidase by AMP.     

Temperature acclimation in brook trout muscle: Adenine nucleotide concentrations, phosphorylation state and adenylate energy charge

Summary Cold acclimation in fish is associated with an elevation in metabolic rate. The present study investigates the role of adenine nucleotides and related compounds in metabolic regulation following temperature acclimation. Brook trout (Salvelinus fontinalis) were acclimated for 10 weeks to either +4°C or +24°C. Both groups of fish were exercised at 2.5 body lengths s^–1 for 2 weeks prior to sacrifice in order to control for differences in spontaneous activity.Concentrations of ATP, ADP, AMP, P _i and PC were approximately 2-fold higher in white than red muscles. Temperature acclimation had little effect on total adenine nucleotide concentration in either muscle type. In white fibres acclimation to 4°C results in a 39% increase in [ADP] and [AMP], a 35% decrease in [PC] (phosphorylcreatine), and no significant change in [P _i ]. In contrast temperature has little effect on concentrations of these compounds in red muscle.Parameters of metabolic control — adenylate energy charge ([ATP]+0.5 [ADP]/[ATP]+[ADP]+[AMP]), phosphorylation state ([ATP]/[ADP]·[P _i ]), and the ratios [ATP][ADP] and [ATP][AMP] — were significantly lower in cold- than warm-acclimated white muscle. The observed changes in phosphorylation state and [ATP][AMP] are consistent with an increase in mitochondrial respiration and glycolysis, respectively.In conclusion, changes in metabolites may be an important factor in producing an enhanced metabolic rate in cold-acclimated fish.     

Adenine nucleotide contents in callus-forming potato tuber discs with different respiratory characteristics

ATP content (per g fresh weight) and energy charge were higher during incubation at 8°C than at 28°C in the early stages of callus induction of potato tuber discs ( Solanum luberosum L. cv. Bintje). After a transfer from 28°C to 8°C, ATP content increased while a sharp decline in ATP content was observed after a transfer from 8°C to 28°C. ADP and AMP pools did not increase correspondingly. When the callus discs had entered the logarithmic growth phase, the energy charge was maintained within relatively narrow limits (0.77–0.80) at all culture temperatures.     

Alteration of 1,2-diacylglycerol content in ischemic and reperfused heart

Human term placenta contains an ATP diphosphohydrolase activity which hydrolyses ATP to ADP and inorganic phosphate and ADP to AMP and a second mole of inorganic phosphate. The activity has a pH optimum between 8.0 and 8.5. Magnesium or calcium ions are required for maximum activity. Other nucleoside phosphates, p-nitrophenyl phosphate or sodium pyrophosphate, are not hydrolysed. The activity is not due to ATPases, or to myokinase, as determined by the use of inhibitors. NaF and NaN₃ were found to inhibit strongly the activity thus identifying it as an ATP diphosphohydrolase.A sensitive enzymatic assay for measurement of AMP, one of the products of the reaction, was established, based on the strong inhibition of muscle fructose 1,6-biphosphatase by AMP. The range of the assay was 0.05–0.8 µM AMP. ATP diphosphohydrolase was found to have a rate of AMP production from ADP twice the rate from ATP. Under the same conditions, the assay for Pi release, on the other hand, gave velocities similar to each other for the two substrates.The activity appears to be identical to the ADP-hydrolysing activity in placenta reported by others.Abbreviations Ap₅A P¹ - P⁵-di(adenosine-5) Pentaphosphate - ATP-DPH ATP Diphosphohydrolase - DCCD N,N Dicyclohexycarbodiimide - Fru-P₂ase Fructose 1,6-biphosphatase - SDS Sodium Dodecyl Sulfate - TLC Thin Layer Chromatography     

Active transport of ATP and presence of a vanadate-sensitive membrane-bound ATPase in Mycobacterium leprae.

It is not known how Mycobacterium leprae obtains energy for survival and growth in the host tissues; the organism does not grow in vitro. In the studies reported here, M. leprae incorporated labelled ATP, which was blocked by cyanide, unlabelled ATP or ADP, but not by adenosine or Pi. It seems that the organism takes up unhydrolysed ATP by an active transport process. The bacterium contained a membrane-bound, vanadate-sensitive E1 E2-ATPase (which creates a transmembrane potential driving transport of solutes into cells). The enzyme was not inhibited by N-ethylmaleimide, suggesting that it is not an F0F1-ATPase which catalyses ATP synthesis. Apparently, M. leprae derives energy-rich compounds from the host cell.     

Balanced contribution of glycolytic and adenylate pool in supply of metabolic energy in platelets

When platelets are treated with H2O2 the metabolic ATP content decreases sharply (Holmsen, H., and Robkin, L. (1977) J. Biol. Chem. 252, 1752-1757). Here we report that the loss of metabolic energy is fully recovered in phosphorylated glycolytic intermediates. A mixture of antimycin A/2-deoxy-D-glucose/D-gluconic acid-1,5-lactone blocks mitochondrial ATP resynthesis and prevents the entry of sugars into the glycolytic sequence. The energy-rich phosphates in the adenylate and the glycolytic pool are then consumed in a specific order. First, the glycolytic pool is consumed at a rate of 4.5 mumol of ATP equivalents/min/10(11) cells, and metabolic ATP and ADP are kept stable; then the consumption of the glycolytic pool decreases and metabolic ATP and ADP are consumed, together keeping up with the same rate of energy consumption. Thrombin stimulation increases the energy consumption to about 17 mumol of ATPeq/min/10(11) cells which is now furnished by both the glycolytic and the adenylate pool, again with a preferential consumption of the former. The results show that H2O2 triggers a shift of energy-rich phosphates from the adenylate to the glycolytic pool and that the latter remains rapidly accessible to energy consumption thereby stabilizing the level of metabolic ATP. The adenylate energy charge is independent of the distribution of energy among the two pools, which extends its importance to the regulation of energy supply and demand beyond the adenylate pool.     

Zeitverlauf der Änderungen an energiereichen Phosphorverbindungen im Meerschweinchenherzen nach Asphyxie und Wiederbeatmung

Zusammenfassung 1. Nach verschieden langer Dauer von Asphyxie und anschließender Wiederbeatmung wurde im Meerschweinchenherzen ATP, ADP, AMP, Kreatinphosphat und anorganisches Phosphat bestimmt.2. Bereits 1 Minute nach Asphyxie kam es zu einem deutlichen Abfall von Kreatinphosphat und Anstieg von anorganischem Phosphat; 5 Minuten nach Asphyxie war Kreatinphosphat nicht mehr nachweisbar.3. ATP fiel 2 Minuten nach Asphyxie ab und zeigte einen weiteren Abfall bis 15 Minuten nach Asphyxie, während AMP zuerst einen Anstieg, später einen Abfall zeigte.4. Bei Asphyxiedauer von 1 Minute regenerierten sich die energiereichen Phosphorverbindungen im Meerschweinchenherzen nach Widerbeatmung von 1/2 Minute und bei 5 Minuten Asphyxiedauer nach 9 Minuten Wiederbeatmung.5. Dauert die Asphyxie 7 Minuten, so ist keine vollständige Regeneration der energiereichen Phosphorverbindungen mehr erreichbar.6. War bei der Wiederbeatmung eine 50- bis 60prozentige Regeneration des Kreatinphosphatanteiles eingetreten, so verschwanden im EKG regelmäßig die Rhythmusstörungen.

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Time course of changes in energy-rich phosphate compounds in the heart of the guinea pig following asphyxia and renimation

In the myocardium of guinea pigs the levels of ATP, ADP, AMP, creatine phosphate and inorganic phosphate, respectively, were determined following different periods of asphyxia and subsequent reanimation. Depending upon the duration of asphyxia, all energy-rich phosphate compounds decreased, creatine phosphate demonstrating this effect most markedly. Following reanimation, the energy-rich phosphate compounds underwent partial or complete regeneration, unless asphyxia lasted longer than 5 to 7 minutes.

     

Brown adipose tissue dynamics in wild-type and UCP1-knockout mice: in vivo insights with magnetic resonance

We used noninvasive magnetic resonance imaging (MRI) and magnetic resonance spectroscopy to compare interscapular brown adipose tissue (iBAT) of wild-type (WT) and uncoupling protein 1 (UCP1)-knockout mice lacking UCP1-mediated nonshivering thermogenesis (NST). Mice were sequentially acclimated to an ambient temperature of 30°C, 18°C, and 5°C. We detected a remodeling of iBAT and a decrease in its lipid content in all mice during cold exposure. Ratios of energy-rich phosphates (ATP/ADP, phosphocreatine/ATP) in iBAT were maintained stable during noradrenergic stimulation of thermogenesis in cold- and warm-adapted mice and no difference between the genotypes was observed. As free fatty acids (FFAs) serve as fuel for thermogenesis and activate UCP1 for uncoupling of oxidative phosphorylation, brown adipose tissue is considered to be a main acceptor and consumer of FFAs. We measured a major loss of FFAs from iBAT during noradrenergic stimulation of thermogenesis. This mobilization of FFAs was observed in iBAT of WT mice as well as in mice lacking UCP1. The high turnover and the release of FFAs from iBAT suggests an enhancement of lipid metabolism, which in itself contributes to the sympathetically activated NST and which is independent from uncoupled respiration mediated by UCP1. Our study demonstrates that MRI, besides its potential for visualizing and quantification of fat tissue, is a valuable tool for monitoring functional in vivo processes like lipid and phosphate metabolism during NST.