RESUMO
Mitochondrial proton leak in rat muscle is responsible for approx. 15% of the standard metabolic rate, so its modulation could be important in regulating metabolic efficiency. We report in the present paper that physiological concentrations of AMP (K(0.5)=80 microM) increase the resting respiration rate and double the proton conductance of rat skeletal-muscle mitochondria. This effect is specific for AMP. AMP also doubles proton conductance in skeletal-muscle mitochondria from an ectotherm (the frog Rana temporaria), suggesting that AMP activation is not primarily for thermogenesis. AMP activation in rat muscle mitochondria is unchanged when uncoupling protein-3 is doubled by starvation, indicating that this protein is not involved in the AMP effect. AMP activation is, however, abolished by inhibitors and substrates of the adenine nucleotide translocase (ANT), suggesting that this carrier (possibly the ANT1 isoform) mediates AMP activation. AMP activation of ANT could be important for physiological regulation of metabolic rate.
Assuntos
Monofosfato de Adenosina/farmacologia , Monofosfato de Adenosina/fisiologia , Atractilosídeo/análogos & derivados , Mitocôndrias/metabolismo , Músculo Esquelético/metabolismo , Animais , Atractilosídeo/metabolismo , Proteínas de Transporte/metabolismo , Relação Dose-Resposta a Droga , Eletrofisiologia , Feminino , Privação de Alimentos , Canais Iônicos , Cinética , Potenciais da Membrana , Mitocôndrias Hepáticas/metabolismo , Translocases Mitocondriais de ADP e ATP/metabolismo , Proteínas Mitocondriais , Consumo de Oxigênio , Isoformas de Proteínas , Ranidae , Ratos , Ratos Wistar , Temperatura , Proteína Desacopladora 3RESUMO
Uncoupling protein-3 (UCP-3) is a recently identified member of the mitochondrial transporter superfamily that is expressed predominantly in skeletal muscle. However, its close relative UCP-1 is expressed exclusively in brown adipose tissue, a tissue whose main function is fat combustion and thermogenesis. Studies on the expression of UCP-3 in animals and humans in different physiological situations support a role for UCP-3 in energy balance and lipid metabolism. However, direct evidence for these roles is lacking. Here we describe the creation of transgenic mice that overexpress human UCP-3 in skeletal muscle. These mice are hyperphagic but weigh less than their wild-type littermates. Magnetic resonance imaging shows a striking reduction in adipose tissue mass. The mice also exhibit lower fasting plasma glucose and insulin levels and an increased glucose clearance rate. This provides evidence that skeletal muscle UCP-3 has the potential to influence metabolic rate and glucose homeostasis in the whole animal.
Assuntos
Proteínas de Transporte/fisiologia , Músculo Esquelético/fisiologia , Tecido Adiposo/metabolismo , Animais , Animais Geneticamente Modificados , Glicemia/metabolismo , Proteínas de Transporte/genética , Metabolismo Energético , Feminino , Humanos , Hiperfagia/genética , Canais Iônicos , Imageamento por Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Proteínas Mitocondriais , Fenótipo , Magreza , Proteína Desacopladora 3RESUMO
Many animals across the phylogenetic scale are routinely capable of depressing their metabolic rate to 5-15% of that at rest, remaining in this state sometimes for years. However, despite its widespread occurrence, the biochemical processes associated with metabolic depression remain obscure. We demonstrate here the development of an isolated cell model for the study of metabolic depression. The isolated cells from the hepatopancreas (digestive gland) of the land snail (Helix aspersa) are oxygen conformers; i.e., their rate of respiration depends on pO(2). Cells isolated from estivating snails show a stable metabolic depression to 30% of control (despite the long and invasive process of cell isolation) when metabolic rate at the physiological pH and pO(2) of the hemolymph of estivating snails is compared with metabolic rate at the physiological pH and pO(2) of the hemolymph of control snails. When the extrinsic effects of pH and pO(2) are excluded, the intrinsic metabolic depression of the cells from estivating snails is still to below 50% of control snails. The in vitro effect of pO(2) on metabolic rate is independent of pH and state (awake or estivating), but the effects of pH and state significantly interact. This suggests that pH and state change affect metabolic depression by similar mechanisms but that the metabolic depression by hypoxia involves a separate mechanism.
Assuntos
Sistema Digestório/metabolismo , Estivação , Caracois Helix/fisiologia , Animais , Concentração de Íons de Hidrogênio , Concentração Osmolar , Oxigênio/metabolismoRESUMO
There is a futile cycle of pump and leak of protons across the mitochondrial inner membrane. The contribution of the proton cycle to standard metabolic rate is significant, particularly in skeletal muscle, and it accounts for 20% or more of the resting respiration of a rat. The mechanism of the proton leak is uncertain: basal proton conductance is not a simple biophysical leak across the unmodified phospholipid bilayer. Equally, the evidence that it is catalysed by homologues of the brown adipose uncoupling protein, UCP1, is weak. The yeast genome contains no clear UCP homologue but yeast mitochondria have normal basal proton conductance. UCP1 catalyses a regulated inducible proton conductance in brown adipose tissue and the possibility remains open that UCP2 and UCP3 have a similar role in other tissues, although this has yet to be demonstrated.
Assuntos
Proteínas de Membrana Transportadoras , Mitocôndrias/fisiologia , Proteínas Mitocondriais , Força Próton-Motriz/fisiologia , Animais , Metabolismo Basal , Proteínas de Transporte/metabolismo , Catálise , Humanos , Membranas Intracelulares/metabolismo , Canais Iônicos , Proteínas de Membrana/metabolismo , Proteínas/metabolismo , Proteína Desacopladora 1 , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
Proton pumping across the mitochondrial inner membrane and proton leak back through the natural proton conductance pathway make up a futile cycle that dissipates redox energy. We measured respiration and average mitochondrial membrane potential in perfused rat hindquarter with maximal tetanic contraction of the left gastrocnemius-soleus-plantaris muscle group, and we estimate that the mitochondrial proton cycle accounted for 34% of the respiration rate of the preparation. Similar measurements in rat hepatocytes given substrates to cause a high rate of gluconeogenesis and ureagenesis showed that the proton cycle accounted for 22% of the respiration rate of these cells. Combining these in vitro values with literature values for the contribution of skeletal muscle and liver to standard metabolic rate (SMR), we calculate that the proton cycle in working muscle and liver may account for 15% of SMR in vivo. Although this value is less than the 20% of SMR we calculated previously using data from resting skeletal muscle and hepatocytes, it is still large, and we conclude that the futile proton cycle is a major contributor to SMR.
Assuntos
Fígado/metabolismo , Mitocôndrias Hepáticas/metabolismo , Mitocôndrias Musculares/metabolismo , Músculo Esquelético/metabolismo , Consumo de Oxigênio/fisiologia , Bombas de Próton/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Feminino , Membro Posterior , Membranas Intracelulares/fisiologia , Fígado/citologia , Masculino , Potenciais da Membrana/fisiologia , Contração Muscular/fisiologia , Músculo Esquelético/fisiologia , Bombas de Próton/fisiologia , Ratos , Ratos WistarRESUMO
The relationship between UCP2 and UCP3 expression and mitochondrial proton conductance of rat skeletal muscle was examined. Rats were starved for 24 h and the levels of UCP2 and UCP3 mRNA and UCP3 protein were determined by Northern and Western blots. Proton conductance was measured by titrating mitochondrial respiration rate and membrane potential with malonate. Starvation increased UCP2 and UCP3 mRNA levels more than 5-fold and 4-fold, respectively, and UCP3 protein levels by 2-fold. However, proton conductance remained unchanged. These results suggest either that Northern and Western blots do not reflect the levels of active protein or that these UCPs do not catalyse the basal proton conductance in skeletal muscle mitochondria.
Assuntos
Proteínas de Transporte/metabolismo , Proteínas de Membrana Transportadoras , Mitocôndrias/metabolismo , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Proteínas/metabolismo , Prótons , Desacopladores/metabolismo , Animais , Northern Blotting , Western Blotting , Proteínas de Transporte/genética , Eletrofisiologia , Feminino , Privação de Alimentos , Canais Iônicos , Cinética , Proteínas/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
We measured the proton leak across the inner membrane of liver mitochondria isolated from six different vertebrate species and from obese and control Zucker rats. Proton leak at 37 degrees C was similar in rat and pigeon, and in obese and control Zucker rats. Compared to rat, it was lower in cane toad, shingleback lizard, and the Madeiran lizard Lacerta dugessi. Proton leak at 20 degrees C was similar in xenopus toad and higher in rainbow trout, compared to rat. In general, proton permeability and substrate oxidation activity were greater in liver mitochondria from endotherms than those from ectotherms. Analysis of this and previous data showed that proton leak per milligram of mitochondrial protein correlated with standard metabolic rate, and proton leak per milligram of inner membrane phospholipid correlated with 11 phospholipid fatty acid compositional parameters, including unsaturation index.
Assuntos
Regulação da Temperatura Corporal/fisiologia , Membranas Intracelulares/metabolismo , Mitocôndrias Hepáticas/metabolismo , Obesidade/metabolismo , Ratos Zucker/metabolismo , Vertebrados/metabolismo , Animais , Respiração Celular , Ácidos Graxos/análise , Potenciais da Membrana/fisiologia , Mitocôndrias Hepáticas/ultraestrutura , Permeabilidade , Fosfolipídeos/química , Prótons , Ratos , Especificidade da EspécieRESUMO
Radioallergosorbent testing was used to look for the presence of specific IgE antibody against a para-tolyl-mono-isocyanate human serum albumin conjugate in sera from five groups of subjects. The first three groups consisted of individuals exposed to toluene diisocyanate (TDI) who had been shown by bronchial provocation testing with levels of TDI below the threshold limit value of 0.02 parts/10(6), to have immediate asthmatic reactions, late asthmatic reactions or no respiratory changes at all. The two control groups consisted of atopic and non-atopic individuals who had no respiratory symptoms and no known exposure to TDI. Although RAST showed high ct/min in some of the sera from patients with proven TDI-induced respiratory disease, these levels were not significantly different from controls and appeared to reflect the presence in these sera of high levels of total IgE (greater than 100 mu ml--1). There is no evidence from this study for the presence of specific IgE antibody against a para-tolyl mono isocyanate human serum albumin conjugate in patients with TDI-induced respiratory disease. This finding may reflect absence of antibodies, or that the techniques for their detection are not always effective even when performed by experienced persons, and there is a potential source of error in the interpretation of results when sera contain large amounts of IgE.
Assuntos
Formação de Anticorpos , Asma/etiologia , Cianatos/efeitos adversos , Isocianatos , Doenças Profissionais/imunologia , Tolueno 2,4-Di-Isocianato/efeitos adversos , Especificidade de Anticorpos , Asma/imunologia , Humanos , Imunoglobulina E , Doenças Profissionais/etiologia , Radioimunoensaio , Albumina Sérica/imunologiaRESUMO
One hundred and twenty-two patients with intrinsic asthma, extrinsic atopic asthma, or asthma with allergic bronchopulmonary aspergillosis were tissue typed for the HLA A, B, and C loci. No associations were found with any of the clinical groups, or with serum total IgE concentrations. Sixty-eight members of 10 families where more than one member was affected by asthma were studied. The segregation of haplotypes in siblings of the propositi who were or were not affected by asthma did not differ from the predicted segregation, and there were no differences when atopy or serum total IgE were considered. No biologically important association between HLA and asthma has been shown.