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J Nedergaard

Publications and source records attributed to J Nedergaard.

At least 37 records · Page 2Linked to original sources
PubMed1999

UCP1: the original uncoupling protein--and perhaps the only one? New perspectives on UCP1, UCP2, and UCP3 in the light of the bioenergetics of the UCP1-ablated mice.

The availability of a UCP1-ablated mouse has enabled critical studies of the function of UCP1, UCP2, and UCP3. Concerning UCP1, its presence in brown-fat mitochondria is associated with innate uncoupling, high GDP-binding capacity, and GDP-inhibitable Cl- permeability and uncoupling--but the high fatty acid sensitivity found in these mitochondria is observed even in the absence of UCP1. The absence of UCP1 leads to low cold tolerance but not to obesity. UCP1 ablation also leads to an augmented expression of UCP2 and UCP3 in brown adipose tissue, making this tissue probably the one that boasts the highest expression of these UCPs. However, these very high expression levels are not associated with any inherent uncoupling, or with a specific GDP-binding capacity, or with a GDP-sensitive Cl- permeability, or with any effect of GDP on mitochondrial membrane potential, or with an increased basal metabolism of cells, or with the presence of norepinephrine- or fatty acid-induced thermogenesis in cells, and not with a cold-acclimation recruited, norepinephrine-induced thermogenic response in the intact animal. Therefore, it can be discussed whether any uncoupling effect is associated with UCP2 or UCP3 when they are endogenously expressed and, consequently, whether (loss of) uncoupling (thermogenic) effects of UCP2 or UCP3 can be invoked to explain metabolic phenomena, such as obesity.

Adipose Tissue, Brown↗
PubMed1999

Benidipine induces thermogenesis in brown adipose tissue by releasing endogenous noradrenaline: a possible mechanism for the anti-obesity effect of calcium antagonists.

BACKGROUND: Anti-obesity effects of calcium antagonists such as benidipine and nifedipine have been described in rodent obesity models, but the mode of action of the calcium antagonists as anti-obesity agents has not been established. OBJECTIVE: To examine whether the anti-obesity effects of calcium antagonists (here benidipine) could be ascribed to a direct stimulation of brown adipose tissue (BAT) thermogenesis. METHODS: Examination of the ability of benidipine to induce thermogenesis (increased rate of oxygen consumption) in isolated brown-fat cells from rats, mice and hamsters--and in intact cold-acclimated rats. RESULTS: Benidipine itself, or in combination with any dose of noradrenaline (NA), was totally unable to induce or augment thermogenesis in isolated brown-fat cells of any species tested. However, it markedly induced thermogenesis in intact animals (approx 60% increase over resting metabolic rate). This effect could be fully inhibited by propranolol. CONCLUSION: Benidipine is itself without thermogenic effect. The thermogenic response in-vivo (and thus presumably the anti-obesity effect) is probably secondary to a previously described general side-effect of calcium antagonists: a release of NA from sympathetic nerves, here most likely directly from nerves in the BAT. The anti-obesity effect of benedipine is thus probably not due to its calcium channel blocking effect. PERSPECTIVES: It is probable that the anti-obesity effects of calcium antagonists reported in several models of genetically obese rodents (MSG-obese and agouti mice, SHHF/Mcc-fa(cp) and JCR:LA-corpulent rats) are mediated via an indirect stimulation of BAT. To what extent calcium antagonists may induce similar effects in a clinical situation, is currently unknown.

Adipose Tissue, Brown↗
PubMed1999

beta1 to beta3 switch in control of cyclic adenosine monophosphate during brown adipocyte development explains distinct beta-adrenoceptor subtype mediation of proliferation and differentiation.

To explain the distinctive pharmacological profiles observed for adrenergic stimulation of cell proliferation (beta1) and cell differentiation (beta3), the adrenergic control of cAMP accumulation was investigated during brown adipocyte development. In preadipocytes, norepinephrine (NE) increased cAMP levels but the beta3-agonists BRL-37344 and CGP-12177 did not; in contrast, when the cells had differentiated into mature brown adipocytes, a large cAMP response to the beta3-agonists had emerged and was now double that to NE (although the affinity of NE had increased 10-fold). Beta1-messenger RNA (mRNA) levels were high in both pre- and mature brown adipocytes; beta3-mRNA did not appear until maturation but then abruptly. Although beta1-receptors remained detectable by [3H]CGP-12177 binding in the mature brown adipocytes, the cAMP response to NE (based on propranolol inhibitory potency) switched from beta1 to beta3. Even the established beta1-agonist dobutamine acted through beta3-receptors in the mature brown adipocytes. The increases in cAMP levels could adequately explain the increased cell proliferation in NE-stimulated preadipocytes and the NE-induced UCP1 gene expression in mature brown adipocytes. The distinctive adrenergic profiles for stimulation of proliferation and of differentiation were thus not due to the existence of additional pathways but to a switch in the type of beta-receptor mediating the NE response, coordinated with an alteration in the nuclear response to increased cAMP levels. The study implies that full recruitment of brown adipose tissue cannot be induced by exclusive beta3-stimulation.

Adipocytes↗
PubMed1998

Carteolol is a weak partial agonist on beta 3-adrenergic receptors in brown adipocytes.

The ability of the beta 1/beta 2 partial agonist carteolol to act as an agonist on beta 3-adrenergic receptors was investigated by studying its ability to stimulate thermogenesis (oxygen consumption) in brown fat cells isolated from hamsters. Carteolol was able to induce thermogenesis, with an EC50 of 5 microM, but it was only a partial (40%) agonist. D,L-Propranolol had a pKB of 5.1 as an antagonist against carteolol in this system, indicating that the carteolol effect was probably mediated via beta 3-receptors. Also in mouse and rat cells, carteolol was a partial agonist with EC50 values around 1 microM. Being a partial agonist, carteolol acted as an antagonist against norepinephrine-, BRL-37344-, or CGP-12177-stimulated thermogenesis with a pKB of approximately 5. The partial agonist effects of carteolol are discussed in relation to the absence of agonist effect of this compound on guinea-pig taenia cecum beta 3-receptors and in relation to the possible plurality of beta 3-receptors.

Adipose Tissue, Brown↗
PubMed1998

Apparent thermogenic effect of injected glucagon is not due to a direct effect on brown fat cells.

To examine the significance of brown adipose tissue for the thermogenic response to glucagon, we injected glucagon intraperitoneally into rats (that have glucagon-sensitive brown fat cells) and into hamsters (that have glucagon-insensitive brown fat cells). Although a thermogenic response to glucagon injection was apparently observed in rats, this response was not augmented by cold acclimation and was not dose dependent. Similar observations were made in hamsters. The thermogenic response could be fully blocked by prior injection of the beta-adrenergic blocker propranolol. Thus no direct thermogenic response to injected glucagon could be demonstrated, and the thermogenic response observed was fully due to vehicle injection. However, glucagon injection was able to unmask mitochondrial [3H]GDP binding. As expected, isolated brown fat cells from rats and mice responded thermogenically to glucagon but brown fat cells from hamsters were unresponsive. The EC50 of the rat brown fat cells was high (5 nM); these cells also responded to secretin, with an EC50 of 22 nM. It was concluded that, in contrast to earlier observations, no thermogenic response to injected glucagon could be observed; this may be related to differences in glucagon preparations. Brown fat cells from certain species are, however, glucagon sensitive. It is uncertain whether glucagon is the endogenous agonist for these receptors, but the presence of the glucagon-responsive receptor indicates alternative means to norepinephrine for stimulation of brown adipose tissue thermogenesis and, probably, of recruitment.

Adipose Tissue, Brown↗
PubMed1998

Thermogenesis is beta3- but not beta1-adrenergically mediated in rat brown fat cells, even after cold acclimation.

To examine if acclimation of rats to cold led to alterations in the coupling between different beta-receptor subtypes and thermogenesis in brown fat cells, we investigated the adrenergic response patterns in brown fat cells isolated from warm-acclimated (28 degreesC) and cold-acclimated (4 degreesC) rats. In the cells from warm-acclimated rats, the relative affinities (EC50) for different agonists (isoprenaline, BRL-37344, norepinephrine, CGP-12177, dobutamine, and salbutamol) were those expected from their interaction with a beta3-receptor. The response to norepinephrine was competitively inhibited by propranolol with a pA2 of approximately 6, implying interaction at the beta3-receptor. No evidence for a beta1-receptor-mediated response to the beta1-selective agonist dobutamine could be obtained; the low-affinity response observed was most likely through the beta3-receptor. The beta1-antagonist ICI-89406 could not inhibit a specific fraction of the thermogenic response to norepinephrine. Thus beta3-receptors were the only beta-receptors involved in the control of thermogenesis in brown fat cells from warm-acclimated rats. A modified method of preparation was developed to isolate functional cells from cold-acclimated animals. Also in these cells, the beta-receptor coupled to thermogenesis was the beta3-receptor, although the response was desensitized with an approximately sevenfold shift in EC50 values. The pA2 for propranolol inhibition of norepinephrine-induced thermogenesis was also 6 here, and that for ICI-89406 was 5.5, also implying interaction at the beta3-receptor. Thus acclimation to cold did not alter the beta-adrenergic receptor subtype (beta3) involved in the control of thermogenesis.

Acclimatization↗
PubMed1998

Regulation of the activity of 27 pS nonselective cation channels in excised membrane patches from rat brown-fat cells.

The regulation of the activity of the approximately 30 pS nonselective cation channel (NSC channel) was studied by the patch-clamp technique in inside-out patches obtained from rat brown-fat cells. NSC channel activity was induced by excision; reduced redox state induced by dithiothreitol accelerated the kinetics in the excised state. The NSC channels were inhibited by the fenamates flufenamic acid and mefenamic acid but not by NS-1619 or SKF-96365. The channels were inhibited by purine nucleotides but not by polyamines. No evidence for protein kinase C, CaM kinase or protein kinase A activation of the NSC channel was obtained. NSC-channel activity was stimulated in a concentration-dependent manner by Ca2+ but the EC50 was very high (0.81 mM), in comparison to expected cytosolic Ca2+ levels. In the presence of ATP, even higher Ca2+ levels were necessary for comparable NSC-channel activation. The increase in Po was not associated with an increase in open-time constants. We conclude that although high Ca2+ levels can experimentally activate the NSC channel, a further mediatory step must probably be postulated in order to link alpha1-adrenergic stimulation to NSC-channel activation.

Adipose Tissue, Brown↗
PubMed1997

alpha1-Adrenergic stimulation potentiates the thermogenic action of beta3-adrenoreceptor-generated cAMP in brown fat cells.

The relationship between cAMP levels and thermogenesis was investigated in brown fat cells from Syrian hamsters. Irrespective of whether the selective beta3-, beta2-, and beta1-agonists BRL 37344, salbutamol, and dobutamine or the physiological agonist norepinephrine was used to stimulate the cells, increases in cAMP levels were mediated via the beta3-receptor, as were the thermogenic effects. However, the relationship "thermogenesis per cAMP" was much lower for agents other than norepinephrine. Similarly, forskolin, although more potent than norepinephrine in elevating cAMP, was less potent in inducing thermogenesis. The selective alpha1-agonist cirazoline was in itself without effect on cAMP levels or thermogenesis, but when added to forskolin-stimulated cells, potentiated thermogenesis, up to the norepinephrine level, without affecting cAMP. This potentiation could not be inhibited by chelerythrine, but could be mimicked by Ca2+ ionophores. It was apparently not mediated via calmodulin-dependent protein kinase and was not an effect on mitochondrial respiratory control. The ability of all cAMP-elevating agents to induce thermogenesis in brown fat cells has earlier been interpreted to mean that it is only through the beta-receptors and the resulting increase in cAMP levels that thermogenesis is induced. However, it is here concluded that the thermogenic response to norepinephrine involves two interacting parts, one mediated via beta-receptors and cAMP and the other via alpha1-receptors and increases in cytosolic Ca2+ levels.

Adipose Tissue, Brown↗
PubMed1997

Cig30, a mouse member of a novel membrane protein gene family, is involved in the recruitment of brown adipose tissue.

We have identified a previously uncharacterized gene that is implicated in the thermogenic function of brown adipose tissue of mice. This gene, termed Cig30, is the first mammalian member of a novel gene family comprising several nematode and yeast genes, such as SUR4 and FEN1, mutation of which is associated with highly pleiotropic phenotypes. It codes for a 30-kDa plasma membrane glycoprotein with five putative transmembrane domains. The Cig30 mRNA was readily detected only in brown fat and liver. When animals were exposed to a 3-day cold stress, the Cig30 expression was selectively elevated in brown fat more than 200-fold. Similar increases were brought about in two other conditions of brown fat recruitment, namely during perinatal development and after cafeteria diet. The magnitude of Cig30 mRNA induction in the cold could be mimicked by chronic norepinephrine treatment in vivo. However, in primary cultures of brown adipocytes, a synergistic action of norepinephrine and dexamethasone was required for full expression of the gene, indicating that both catecholamines and glucocorticoids are required for the induction of Cig30. We propose that the CIG30 protein is involved in a pathway connected with brown fat hyperplasia.

Acetyltransferases↗
PubMed1997

Kinetics of the inhibition of mitochondrial respiration by NO.

The kinetics of the inhibition of mitochondrial respiration by NO was examined in isolated mitochondria (here obtained from rat brown adipose tissue). The Ki of NO for the inhibition was approximately 27 nM; the IC50 of NO increased in proportion to the square of an increase in O2 tension. The Km of O2 for respiration was approximately 16 microM; in the presence of NO, the dependence of respiration on O2 tension had a Hill coefficient of approximately 2. The unusual kinetics is probably related to the ability of cytochrome c oxidase to use 2 NO or 1 O2 as electron acceptor. The interaction between NO and O2 in the control of respiration could be described by the formula VO2(O2, NO) = VO2max x ([O2]2/((16 microM x (1 + [NO]/27 nM))2 + [O2]2)). Thus, the kinetics is such that respiration in the presence of physiological levels of NO is very sensitive to decreasing O2 tension.

Adipose Tissue, Brown↗
PubMed1997

Contrasting adrenergic effects on lipoprotein lipase gene expression in the brown adipose tissue of intact mice and in cultured brown adipocytes from mice.

To examine the regulation of lipoprotein lipase (LPL) gene expression, LPL mRNA levels in the brown adipose tissue of intact mice and in mouse brown adipocyte cultures were examined. In intact mice, exposure to cold resulted in a rapid, transient, 5-fold increase in LPL mRNA level. Norepinephrine (NE) injection could fully mimic the effect of acute exposure to cold, and LPL mRNA and enzymatic activity were increased in parallel after NE injection. These results indicated positive adrenergic control of LPL gene expression in the brown adipose tissue of intact mice. In cultured mouse brown adipocytes, the level of spontaneously expressed LPL mRNA decreased in parallel with the progression of brown adipocyte differentiation. NE treatment of undifferentiated cells led to a decrease in LPL mRNA levels. In brown adipocytes that had reached a mature state, NE had a small negative or no effect on LPL mRNA levels, irrespective of whether the experiment was performed in the presence or absence of insulin or of newborn-calf serum. It was concluded that LPL gene expression in brown adipose tissue in intact mice is under adrenergic control but that this gene is not under positive adrenergic control in cultured brown adipocytes from mice, although these cells are otherwise adrenergically sensitive. The presence of additional factors may be necessary to confer adrenergic sensitivity to the LPL gene in the cultured brown adipocytes; alternatively, cells other than the mature brown adipocytes may confer the positive adrenergic sensitivity to the brown adipose tissue depots in situ.

Adipocytes↗
PubMed1997

Analysis of the cellular mechanism for halothane inhibition of brown adipose tissue thermogenesis.

In the present studies, halothane is demonstrated to have a general inhibitory effect on norepinephrine-induced cAMP accumulation, lipolysis and rate of oxygen consumption, but no obvious effect on the respiratory capacity of isolated mitochondria. Further studies are necessary to elucidate these effects of halothane on brown adipose tissue nonshivering thermogenesis. Such studies may contribute to the understanding of the molecular effects of volatile anesthetics.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1997

Differential regulation of the expression of alpha1-adrenergic receptor subtype genes in brown adipose tissue.

The physiological control of the expression of the genes for the alpha1-adrenoceptor subtypes was examined in rat brown adipose tissue by analysing Northern blots of poly(A)-enriched RNA with oligonucleotide probes. In control rats, alpha1B-receptor gene expression was much lower in brown adipose tissue than in liver, but the expression of both alpha1A and alpha1D was higher than in the heart, making brown adipose tissue one of the mammalian tissues with the highest expression of these subtypes. During acute exposure to cold, alpha1B-receptor gene expression was essentially unchanged, alpha1A-receptor gene expression was increased and alpha1D-receptor gene expression was transiently decreased. Noradrenaline injection could mimic these effects of acute cold exposure, indicating that the physiologically induced up- and down-regulations were due to the interaction of noradrenaline with cells within the tissue. In chronically cold-acclimated animals, alpha1B-receptor gene expression was decreased but that of the alpha1A-receptor gene remained at a level twice that of controls. alpha1D-Receptor gene expression was also somewhat decreased. It is suggested that the enhanced expression of the alpha1A-receptor gene explains the increased alpha1-receptor density in recruited brown adipose tissue reported previously. The intricate and differential regulation of alpha1-receptor gene expression and the markedly enhanced expression of the alpha1A-receptor may imply that alpha1-receptors are important for the recruitment process or for maintenance of the recruited state in this tissue.

Acclimatization↗
PubMed1997

Adrenergic stimulation of lipoprotein lipase gene expression in rat brown adipocytes differentiated in culture: mediation via beta3- and alpha1-adrenergic receptors.

In order to investigate whether the positive effect of adrenergic stimulation on lipoprotein lipase (LPL) gene expression in brown adipose tissue is a direct effect on the brown adipocytes themselves, the expression of the LPL gene was investigated by measuring LPL mRNA levels in brown adipocytes, isolated as precursors from the brown adipose tissue of rats and grown in culture in a fully defined medium before experimentation. Addition of noradrenaline led to an enhancement of LPL gene expression; the mRNA levels increased as a linear function of time for at least 5 h and were finally approx. 3 times higher than in control cells, an increase commensurate with that seen in vivo in both LPL mRNA levels and LPL activity during physiological stimulation. The increase was dependent on transcription. The effect of noradrenaline showed simple Michaelis-Menten kinetics with an EC50 of approx. 11 nM. beta3-Agonists (BRL-37344 and CGP-12177) could mimic the effect of noradrenaline; the beta1-agonist dobutamine and the beta2-agonist salbutamol could not; the alpha1-agonist cirazoline had only a weak effect. The effect of noradrenaline was fully inhibited by the beta-antagonist propranolol and was halved by the alpha1-antagonist prazosin; the alpha2-antagonist yohimbine was without effect. An increase in LPL mRNA level similar to (but not significantly exceeding) that caused by noradrenaline could also be induced by the cAMP-elevating agents forskolin and cholera toxin, and 8-Br-cAMP also increased LPL mRNA levels. The increase in LPL gene expression was not mediated via an increase in the level of an intermediary proteinaceous factor. It is concluded that the physiologically induced increase in LPL gene expression is a direct effect of noradrenaline on the brown adipocytes themselves, mediated via a dominant beta3-adrenergic pathway and an auxiliary alpha1-adrenergic pathway which converge at a regulatory point in transcriptional control.

Adipose Tissue, Brown↗
PubMed1996

Down-regulation of beta3 adrenoreceptor gene expression in brown fat cells is transient and recovery is dependent upon a short-lived protein factor.

The regulation of the expression of the beta3 adrenoreceptor gene was examined in the brown adipose tissue of intact mice and in murine brown fat primary cell cultures. Both in vivo and in vitro, high levels of beta3 receptor mRNA were observed. Acute cold exposure of mice resulted in a marked and rapid down-regulation of beta3 gene expression; this down-regulation was, however, transient. Similarly, in brown fat cell cultures, norepinephrine addition led to down-regulation of beta3 gene expression, with a lag phase of 30 min and with an apparent half-life of beta3 mRNA of approximately 30 min. This down-regulation was stimulated via the beta3 receptors themselves and mediated via cAMP; the apparent affinity of norepinephrine was extremely high (<1 nM). The degradation rate after actinomycin was identical to that after norepinephrine and was not affected by the presence of norepinephrine; thus, the down-regulation was due to cessation of transcription but not to an increased rate of degradation. Notably, inhibition of protein synthesis by cycloheximide also led to down-regulation. The norepinephrine-induced down-regulation was transient; spontaneous recovery occurred after approximately 18 h and was not due to depletion of adrenergic agent. Recovery did not occur in the presence of cycloheximide. After recovery, the cells showed a functional desensitization of the down-regulation process itself (EC50 now approximately 10 nM). It is concluded that a down-regulated state cannot explain the functional desensitization of beta3 adrenergic responsiveness observed in brown fat cells isolated from cold-acclimated animals (i.e. physiologically chronically adrenergically stimulated brown fat cells); since the beta3 receptor is not subject to desensitization via phosphorylation processes, no satisfactory explanation for the functional desensitization exists as yet. A model is presented for the down-regulation/recovery process, involving the participation of a phosphorylatable short-lived transcription factor.

Adipose Tissue, Brown↗
PubMed1996

Induction of uncoupling protein in brown adipose tissue. Synergy between norepinephrine and pioglitazone, an insulin-sensitizing agent.

Insulin resistance and obesity in rodent models of non-insulin-dependent diabetes mellitus have been correlated with ablated or defective brown adipose tissue (BAT) function. The mitochondrial uncoupling protein (UCP) allows BAT to perform its unique role in facultative energy expenditure. In this study, we observed an increase in both BAT mass and the expression of UCP mRNA in BAT from obese diabetic mice and their lean littermates following treatment with the thiazolidinedione pioglitazone, a novel insulin-sensitizing agent. Thus, we wanted to ascertain if pioglitazone directly induces BAT differentiation. We found that treatment for 48 hr with pioglitazone caused a 32-fold increase in UCP mRNA, whereas a 7-hr treatment with norepinephrine caused a 24-fold increase in expression. Cells treated with pioglitazone for 48 hr, with norepinephrine added during the last 7 hr, demonstrated a 59-fold increase in UCP mRNA. However, simultaneous treatment with pioglitazone and repeated treatment norepinephrine for 48 hr yielded a greater than 200-fold increase in UCP mRNA. Examination of UCP protein levels demonstrated a similar time-dependent increase with pioglitazone and/or norepinephrine treatment, as well as a synergistic increase with concurrent pioglitazone and norepinephrine treatment. This study shows that pioglitazone exerts a direct effect on BAT cells in vitro by increasing UCP mRNA and protein levels, and that it also synergizes with norepinephrine perhaps by inducing and stabilizing UCP mRNA and/or preventing proteolysis of UCP protein.

Adipocytes↗
PubMed1996

Cold-induced reduction in Gi alpha proteins in brown adipose tissue. Effects on the cellular hypersensitization to noradrenaline caused by pertussis-toxin treatment.

The significance of Gi proteins for the physiological desensitization phenomena observed in brown-fat cells from cold-acclimated hamsters was investigated. For this purpose, pertussis toxin (the inhibitor of Gi function) was injected into control and cold-acclimated hamsters. After 3 days the thermogenic response to noradrenaline injection was monitored in the intact animals. It was found that the pertussis-toxin pretreatment did not affect the thermogenic response to noradrenaline. Nonetheless, the pertussis toxin pretreatment had a dramatic effect on the noradrenaline-sensitivity of isolated brown-fat cells (measured the following day as the respiratory response): a 250-fold-increased sensitivity to noradrenaline was observed in cells from control animals that had been pertussis-toxin pretreated. However, only a 20-fold increase was observed in cells from cold-acclimated hamsters, implying a lower complement of the Gi system in these cells. Therefore the content of Gi proteins was determined by quantitative immunoblotting of purified plasma-membrane proteins. Cold acclimation resulted in a nearly 50% reduction in the content of Gi 1 alpha and Gi 2 alpha, as well as of the beta-subunit, both when expressed on a protein basis and when related to the content of forskolin-stimulated adenylyl cyclase; when expressed per unit of [3H]ouabain-binding (NA+/K+-ATPase), the reduction was even higher. In view of the magnitude of the pertussis-toxin effect, it was concluded that Gi proteins must play a substantial role in the regulation of the response of brown-fat cells to noradrenaline. As the capacity of the Gi pathway is reduced rather than augmented during cold acclimation, Gi activity cannot be responsible for the desensitization to noradrenaline observed in cells from cold-acclimated animals. However, the reduced Gi content may explain the earlier observed desensitization to adenosine that occurs after acclimation to cold.

Acclimatization↗