RESUMO
Menopause is associated with increased visceral adiposity and disrupted glucose homeostasis, but the underlying molecular mechanisms related to these metabolic changes are still elusive. Brown adipose tissue (BAT) plays a key role in energy expenditure that may be regulated by sexual steroids, and alterations in glucose homeostasis could precede increased weight gain after ovariectomy. Thus, the aim of this work was to evaluate the metabolic pathways in both the BAT and the liver that may be disrupted early after ovariectomy. Ovariectomized (OVX) rats had increased food efficiency as early as 12 days after ovariectomy, which could not be explained by differences in feces content. Analysis of isolated BAT mitochondria function revealed no differences in citrate synthase activity, uncoupling protein 1 expression, oxygen consumption, ATP synthesis, or heat production in OVX rats. The addition of GDP and BSA to inhibit uncoupling protein 1 decreased oxygen consumption in BAT mitochondria equally in both groups. Liver analysis revealed increased triglyceride content accompanied by decreased levels of phosphorylated AMP-activated protein kinase and phosphorylated acetyl-CoA carboxylase in OVX animals. The elevated expression of gluconeogenic enzymes in OVX and OVX + estradiol rats was not associated with alterations in glucose tolerance test or in serum insulin but was coincident with higher glucose disposal during the pyruvate tolerance test. Although estradiol treatment prevented the ovariectomy-induced increase in body weight and hepatic triglyceride and cholesterol accumulation, it was not able to prevent increased gluconeogenesis. In conclusion, the disrupted liver glucose homeostasis after ovariectomy is neither caused by estradiol deficiency nor is related to increased body mass.
Assuntos
Tecido Adiposo Marrom/metabolismo , Estradiol/fisiologia , Fígado/metabolismo , Menopausa/metabolismo , Aumento de Peso , Animais , Feminino , Glucose/metabolismo , Obesidade/etiologia , Obesidade/metabolismo , Ovariectomia , Ratos , Ratos Wistar , Termogênese , Triglicerídeos/metabolismoRESUMO
The UCP1 [first UCP (uncoupling protein)] that is found in the mitochondria of brown adipocytes [BAT (brown adipose tissue)] regulates the heat production, a process linked to non-shivering thermogenesis. The activity of UCP1 is modulated by GDP and fatty acids. In this report, we demonstrate that respiration and heat released by BAT mitochondria vary depending on the respiratory substrate utilized and the coupling state of the mitochondria. It has already been established that, in the presence of pyruvate/malate, BAT mitochondria are coupled by faf-BSA (fatty-acid-free BSA) and GDP, leading to an increase in ATP synthesis and mitochondrial membrane potential along with simultaneous decreases in both the rates of respiration and heat production. Oleate restores the uncoupled state, inhibiting ATP synthesis and increasing the rates of both respiration and heat production. We now show that in the presence of succinate: (i) the rates of uncoupled mitochondria respiration and heat production are five times slower than in the presence of pyruvate/malate; (ii) faf-BSA and GDP accelerate heat and respiration as a result and, in coupled mitochondria, these two rates are accelerated compared with pyruvate/malate; (iii) in spite of the differences in respiration and heat production noted with the two substrates, the membrane potential and the ATP synthesized were the same; and (iv) oleate promoted a decrease in heat production and respiration in coupled mitochondria, an effect different from that observed using pyruvate/malate. These effects are not related to the production of ROS (reactive oxygen species). We suggest that succinate could stimulate a new route to heat production in BAT mitochondria.
Assuntos
Tecido Adiposo Marrom/metabolismo , Ácidos Graxos/metabolismo , Guanosina Difosfato/metabolismo , Mitocôndrias/metabolismo , Trifosfato de Adenosina/biossíntese , Animais , Respiração Celular/efeitos dos fármacos , Ácidos Graxos/farmacologia , Peróxido de Hidrogênio/metabolismo , Malatos/metabolismo , Malatos/farmacologia , Masculino , Ácido Oleico/metabolismo , Ácido Oleico/farmacologia , Ácido Pirúvico/metabolismo , Ácido Pirúvico/farmacologia , Ratos , Ratos Wistar , Rotenona/farmacologia , Ácido Succínico/metabolismo , Ácido Succínico/farmacologia , Termogênese/efeitos dos fármacos , Desacopladores/farmacologiaRESUMO
Brown adipose tissue (BAT) mitochondria thermogenesis is regulated by uncoupling protein 1 (UCP 1), GDP and fatty acids. In this report, we observed fusion of the endoplasmic reticulum (ER) membrane with the mitochondrial outer membrane of rats BAT. Ca(2+)-ATPase (SERCA 1) was identified by immunoelectron microscopy in both ER and mitochondria. This finding led us to test the Ca(2+) effect in BAT mitochondria thermogenesis. We found that Ca(2+) increased the rate of respiration and heat production measured with a microcalorimeter both in coupled and uncoupled mitochondria, but had no effect on the rate of ATP synthesis. The Ca(2+) concentration needed for half-maximal activation varied between 0.08 and 0.11 microM. The activation of respiration was less pronounced than that of heat production. Heat production and ATP synthesis were inhibited by rotenone and KCN. Liver mitochondria have no UCP1 and during respiration synthesize a large amount of ATP, produce little heat, GDP had no effect on mitochondria coupling, Ca(2+) strongly inhibited ATP synthesis and had little or no effect on the small amount of heat released. These finding indicate that Ca(2+) activation of thermogenesis may be a specific feature of BAT mitochondria not found in other mitochondria such as liver.
Assuntos
Tecido Adiposo Marrom/metabolismo , Tecido Adiposo/metabolismo , Retículo Endoplasmático/metabolismo , Membranas Intracelulares/metabolismo , Mitocôndrias/metabolismo , Trifosfato de Adenosina/química , Animais , Calorimetria/métodos , Cianatos/química , Masculino , Mitocôndrias Hepáticas/metabolismo , Consumo de Oxigênio , Ratos , Ratos Wistar , Rotenona/farmacologiaRESUMO
Short-term response to cold promotes a small but significant rise in serum T3 in euthyroid rabbits, where the heart is an important target of T3 action. In this work, we measured changes in sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA2a) and phospholamban (PLB) in hearts of hypo- and hyperthyroid rabbits and compared them with modifications induced by short- and long-term cold exposure. Short-term cold exposure promotes a small increase in T3 and, similar to hyperthyroidism, induces an increase of heart SERCA2a expression. The total PLB content does not change in hyperthyroidism, but short-term cold exposure promotes a significant decrease in total PLB and an increase in the ratio between phosphorylated and total PLB. The temperature of a given tissue depends on the balance between the heat provided by blood circulation and the rate of heat production by the tissue. In an attempt to evaluate the heat contribution of cardiac tissue, we measured mitochondrial respiration in permeabilized cardiac muscle and heat produced by cardiac sarcoplasmic reticulum (SR) during Ca(2+) transport. We observed that there was an increase in oxygen consumption and heat production during Ca(2+) transport by cardiac SR in both hyperthyroidism and short-term cold exposure. In contrast, both the mitochondrial respiration rate and heat derived from Ca(2+) transport were decreased in hypothyroid rabbits. The heart changes in oxygen consumption, SERCA2a-PLB ratio, and Ca(2+)-ATPase activity detected during short-term cold exposure were abolished after cold adaptation. We hypothesize that the transient rise in serum T3 contributes to the short-term response to cold exposure.
Assuntos
Regulação da Temperatura Corporal/fisiologia , Proteínas de Ligação ao Cálcio/metabolismo , Miocárdio/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Tri-Iodotironina/metabolismo , Adaptação Fisiológica/fisiologia , Animais , Cálcio/metabolismo , Citrato (si)-Sintase/metabolismo , Temperatura Baixa , Metabolismo Energético/fisiologia , Temperatura Alta , Hipertireoidismo/metabolismo , Hipotireoidismo/metabolismo , Masculino , Mitocôndrias Cardíacas/metabolismo , Consumo de Oxigênio/fisiologia , Coelhos , Tiroxina/metabolismo , Tiroxina/farmacologiaRESUMO
In this work, we compared the effect of K+ on vesicles derived from the longitudinal (LSR) and terminal cisternae (HSR) of rabbit white muscle. In HSR, K+ was found to inhibit both the Ca2+ accumulation and the heat released during ATP hydrolysis by the Ca2+-ATPase (SERCA1). This was not observed in LSR. Valinomycin abolished the HSR Ca2+-uptake inhibition promoted by physiological K+ concentrations, but it did not modify the thermogenic activity of the Ca2+ pump. The results with HSR are difficult to interpret, assuming that a single K+ is binding to either the ryanodine channel or to the Ca2+-ATPase. It is suggested that an increase of K+ in the assay medium alters the interactions among the various proteins found in HSR, thus modifying the properties of both the ryanodine channel and SERCA1.
Assuntos
Sinalização do Cálcio , Cálcio/metabolismo , Fibras Musculares de Contração Rápida/metabolismo , Potássio/metabolismo , Retículo Sarcoplasmático/metabolismo , Animais , Temperatura Alta , Hidrólise , Coelhos , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismoRESUMO
Brown adipose tissue (BAT) is involved in rat and mice thermoregulation, and heat produced by BAT depends on the concerted action of thyroid hormones and catecholamines. Little is known about cold-induced thermogenesis in mammals that have little or no BAT, such as rabbits. In these animals, thermogenesis primarily occurs in skeletal muscle. In this work, we have studied the effect of cold acclimation (4 C for 10 d) in normal and hypothyroid rabbits. It is known that hypothyroid rats die after a few hours of cold exposure. We now show that, different from rats, hypothyroid rabbits sustain their body temperature and survive after 10 d cold exposure. When compared with rabbits kept at room temperature, the muscles of cold-exposed rabbits showed a dark red color characteristic of oxidative muscle fibers. According to this pattern, we observed that in both normal and hypothyroid rabbits, cold exposure promotes an increase in oxygen consumption by skeletal muscle mitochondria. Moreover, in red muscle, cold acclimation induces an increase in the expression and activity of sarcoplasmic reticulum Ca(2+) ATPase isoform 1 (SERCA1), one of the muscle enzymes involved in heat production. We conclude that rabbit cold tolerance is probably related to increased muscle oxidative metabolism and heat production by SERCA1 and that these changes are not completely dependent on normal thyroid function.
Assuntos
Temperatura Baixa , Hipotireoidismo/metabolismo , Mitocôndrias Musculares/metabolismo , Músculo Esquelético/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Animais , Western Blotting , Peso Corporal , Ingestão de Alimentos , Hipotireoidismo/sangue , Hipotireoidismo/fisiopatologia , Masculino , Mitocôndrias Musculares/fisiologia , Músculo Esquelético/fisiologia , Consumo de Oxigênio/fisiologia , Coelhos , Termogênese , Tiroxina/sangue , Fatores de Tempo , Tri-Iodotironina/sangueRESUMO
This report is divided in two parts. The first section shows that vesicles derived from the sarcoplasmic reticulum of rats skeletal muscle can cleave ATP at a faster rate and produce more heat that the vesicles derived from rabbit skeletal muscle. In the second part, we compared the rates of Ca(2+) transport and ATP hydrolysis by rats and rabbits heart sarcoplasmic reticulum. It is shown that the two vesicles preparations are able to use glucose 6-phosphate and hexokinase as an ATP regenerative system. The rates of Ca(2+)-uptake and ATP hydrolysis measured with glucose 6-phosphate and hexokinase is four to six times slower than that measured with phosphoenolpyruvate and pyruvate kinase as ATP regenerative system.
Assuntos
Trifosfato de Adenosina/metabolismo , ATPases Transportadoras de Cálcio/metabolismo , Metabolismo Energético/fisiologia , Glucose-6-Fosfato/metabolismo , Músculo Esquelético/fisiologia , Retículo Sarcoplasmático/fisiologia , Termogênese/fisiologia , Animais , Células Cultivadas , Técnicas In Vitro , Coelhos , Ratos , Ratos WistarRESUMO
The sarcoplasmic reticulum Ca(2+) ATPase 1 (SERCA 1) is able to handle the energy derived from ATP hydrolysis in such a way as to determine the parcel of energy that is used for Ca(2+) transport and the fraction that is converted into heat. In this work we measured the heat production by SERCA 1 in the two sarcoplasmic reticulum (SR) fractions: the light fraction (LSR), which is enriched in SERCA and the heavy fraction (HSR), which contains both the SERCA and the ryanodine Ca(2+) channel. We verified that although HSR cleaved ATP at faster rate than LSR, the amount of heat released during ATP hydrolysis by HSR was smaller than that measured by LSR. Consequently, the amount of heat released per mol of ATP cleaved (DeltaH(cal)) by HSR was lower compared to LSR. In HSR, the addition of 5 mM Mg(2+) or ruthenium red, conditions that close the ryanodine Ca(2+) channel, promoted a decrease in the ATPase activity, but the amount of heat released during ATP hydrolysis remained practically the same. In this condition, the DeltaH(cal) values of ATP hydrolysis increased significantly. Neither Mg(2+) nor ruthenium red had effect on LSR. Thus, we conclude that heat production by SERCA 1 depends on the region of SR in which the enzyme is inserted and that in HSR, the DeltaH(cal) of ATP hydrolysis by SERCA 1 depends on whether the ryanodine Ca(2+) channel is opened or closed.
Assuntos
Músculo Esquelético/enzimologia , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Retículo Sarcoplasmático/enzimologia , Termogênese/fisiologia , Animais , Western Blotting , Cálcio/metabolismo , Músculo Esquelético/fisiologia , Coelhos , Retículo Sarcoplasmático/fisiologiaRESUMO
Previous data from our laboratory showed that the reticulum of the sea cucumber smooth muscle body wall retains both a sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) and a sulfated polysaccharide. In this invertebrate, the transport of Ca(2+) by the SERCA is naturally inhibited by these endogenous sulfated polysaccharides. The inhibition is reverted by K(+) leading to an enhancement of the Ca(2+) transport rate. We now show that vesicles derived from the endoplasmic reticulum of unfertilized eggs from the sea urchin Arbacia lixula retain a SERCA that is able to transport Ca(2+) at the expense of ATP hydrolysis. As described for the sea cucumber SERCA isoform, the enzyme from the sea urchin is activated by K(+) but not by Li(+) and is inhibited by thapsigargin, a specific inhibitor of SERCA. A new sulfated polysaccharide was identified in the sea urchin eggs reticulum composed mainly by galactose, glucose, hexosamine and manose. After extraction and purification, this sulfated polysaccharide was able to inhibit the mammal SERCA isoform found in rabbit skeletal muscle and the inhibition is reversed by K(+). These data suggest that the regulation of the SERCA pump by K(+) and sulfated polysaccharides is not restricted to few marine invertebrates but is widespread.